UNIVERSITAS DIPONEGORO

JUDUL
Extraction and Characterization of Saponin from Sapindus rarak
DC as Plant-Derived Surfactant in Micellar-Enhanced
Ultrafiltration Process

TESIS

AININU NAFIUNISA
NIM. 21030116420027

FAKULTAS TEKNIK
PROGRAM MAGISTER TEKNIK KIMIA

SEMARANG
FEBRUARI 2019

i
 UNIVERSITAS DIPONEGORO

JUDUL
Extraction and Characterization of Saponin from Sapindus rarak
DC as Plant-Derived Surfactant in Micellar-Enhanced
Ultrafiltration Process

TESIS

Diajukan sebagai salah satu syarat untuk memperoleh gelar

Magister Teknik (M.T.)

AININU NAFIUNISA
NIM. 21030116420027

FAKULTAS TEKNIK
PROGRAM MAGISTER TEKNIK KIMIA

SEMARANG
FEBRUARI 2019

i
 HALAMAN PERNYATAAN ORISINALITAS

Tesis ini adalah hasil karya saya sendiri,

dan semua sumber baik yang dikutip maupun yang dirujuk
telah saya nyatakan dengan benar.

NAMA : Aininu Nafiunisa

NIM : 21030116420027

Tanda Tangan : ......................................

Tanggal : Februari 2019

ii
 HALAMAN PENGESAHAN

Skripsi ini diajukan oleh

NAMA : AININU NAFIUNISA
NIM : 21030116420027
Jurusan/Program Studi : PROGRAM STUDI MAGISTER TEKNIK KIMIA
Judul Tesis : Extraction and Characterization of Saponin from Sapindus rarak
DC as Plant-Derived Surfactant in Micellar-Enhanced
Ultrafiltration Process

Telah berhasil dipertahankan di hadapan Tim Penguji dan diterima sebagai bagian
persyaratan yang diperlukan untuk memperoleh gelar Magister Teknik pada
Jurusan/ Program Studi Magister Teknik Kimia, Fakultas Teknik, Universitas
Diponegoro.

TIM PENGUJI

Pembimbing : Nita Aryanti, ST., MT., Ph.D (…………………….)

Penguji : Prof. Dr. Tutuk Djoko Kusworo, ST., M.Eng. (…………………….)

Penguji : Prof. Dr. Ir. Bambang Pramudiono, MS. (…………………….)

Penguji : Prof. Dr. Andri Cahyo Kumoro, ST., M.Eng. (…………………….)

Semarang, Maret 2019

Ketua Departemen Teknik Kimia

Dr. techn. Siswo Sumardiyono, ST., MT.

NIP. 19750915 200012 1 001

iii
 HALAMAN PERNYATAAN PERSETUJUAN PUBLIKASI
TUGAS AKHIR UNTUK KEPENTINGAN AKADEMIS

Sebagai sivitas akademika Universitas Diponegoro, saya yang bertanda tangan di bawah
ini :

Nama : Aininu Nafiunisa

NIM : 21030116420027
Jurusan/Program Studi : Prodi. Magister Teknik Kimia
Departemen : Teknik Kimia
Fakultas : Teknik
Jenis Karya : Tesis

demi pengembangan ilmu pengetahuan, menyetujui untuk memberikan kepada

Universitas Diponegoro Hak Bebas Royalti Noneksklusif (None-exclusive Royalty Free
Right) atas karya ilmiah saya yang berjudul :
“Extraction and Characterization of Saponin from Sapindus rarak DC as Plant-
Derived Surfactant in Micellar-Enhanced Ultrafiltration Process”

beserta perangkat yang ada (jika diperlukan). Dengan Hak Bebas Royalti/Noneksklusif
ini Universitas Diponegoro berhak menyimpan, mengalihmedia/formatkan, mengelola
dalam bentuk pangkalan data (database), merawat dan memublikasikan tugas akhir saya
selama tetap mencantumkan nama saya sebagai penulis/pencipta dan sebagai pemilik Hak
Cipta.

Demikian pernyataan ini saya buat dengan sebenarnya.

Dibuat di : Semarang
Pada Tanggal : Februari 2019

Yang menyatakan

( Aininu Nafiunisa)

iv
 ABSTRACT

Micellar-Enhanced Ultrafiltration Membrane (MEUF) is a promising technology

to separate low molecular weight substances such as dyes. However, the recent
study of MEUF is only focused on the synthethic surfactant which is not
environmentally friendly. For that reason, saponin from Sapindus rarak DC as a
natural surfactant is applied to substitute the synthethic surfactant in MEUF
process. In this research, extraction of saponin from Sapindus rarak and its
surfactant characteristic were investigate. Then, the saponin extract was applied
as surfactant on MEUF process. To achieve the research ultimate goal, the
extraction performed using ultrasonic-assisted (UAE) and maceration extraction
(ME). The characterization of surfactant comprised FTIR, CMC, HLB number, dye
solubilization as well as its potential in the MEUF application for Remazol dyes
separation. It was fixed that extraction by UAE at temperature of 30oC, and solvent
to solute ratio of 10 mL/gr, for 40 minutes gives highest saponin yield. The FTIR
analysis for both pure saponin and saponin extract identified –OH, C-H, C=C, and
C-O-C, the CMC and HLB number were determined. Addition of more saponin in
dye solution increased the solubilized dye. The solubilization power (SP) of
Sapindus rarak extract is similar with the pure saponin for both remazol dye. The
ΔG of Sapindus rarak saponin and pure commercialized saponin for remazol red
and blue showing a negative value, confirms that the process was spontaneous.
Application of saponin extract as surfactant in MUEF system shows that the highest
flux profile was achieved without any addition of saponin. However, with no
surfactant addition the rejection was very low. On the other hand, the addition of
surfactant resulted on decrease of permeate flux until a certain point, but the
%rejection of dye was high. The highest dye rejection was achieved at saponin
concentration of 2 times CMC, having %rejection of 97.02% and 99.42% for
remazol red and blue respectively. A very low Lm was obtained for the solution
having saponin concentration below CMC, for both remazol dye. At CMC, the
micelle loading was increased significantly. However, further addition of saponin
leads to decrease the Lm into 0.0352 mM/mM and 0.0428 mM/mM for remazol red
and blue respectively. Evaluation of model blocking mechanism in the UF process
was standard blocking, UF process with addition of saponin below CMC shows a
cake formation blocking, and the addition of saponin above CMC shows complete
blocking mechanism.

Keyword : Saponins, Micellar-Enhanced Ultrafiltration, Sapindus rarak DC,

Ultrasound-Assisted Extraction

v
 PREFACE

The authors would like to show the highest gratitude to the Almighty God
Allah SWT. By Allah SWT’s graces, this research and its reports entitled
“Extraction and Characterization of Saponin from Sapindus rarak DC as Plant-
Derived Surfactant in Micellar-Enhanced Ultrafiltration Process” has completed.
On this occasion, the author would like to thank Dr. Nita Aryanti, ST., MT. for her
guidance, the author’s parents Mr. Kastollah KP. and Mrs. Sri Miyati, PT Panasonic
Gobel for the financial supports through the Panasonic Gobel Scholarship Program
2017, Prof. Dr. Andri Cahyo Kumoro as the Head of Chemical Engineering
Magister Program UNDIP, the examiner team and all of the supporting parties.
The purpose of this research reports is to explain the final results of the
research study. Of course, this research report still have many shortcomings. For
this reason, the authors expect a constructive criticism and suggestions from various
parties. Finally, the author would like to apologizes if there any mistake in writing
this research report.

Best regards, Author.

vi
 TABLE OF CONTENT

COVER .................................................................................................................... i
INSIDE COVER ...................................................................................................... i
HALAMAN PERNYATAAN ORISINALITAS .................................................... ii
HALAMAN PENGESAHAN ................................................................................ iii
HALAMAN PERNYATAAN PERSETUJUAN PUBLIKASI ............................. iv
ABSTRACT ............................................................................................................ v
PREFACE .............................................................................................................. vi
TABLE OF CONTENT ........................................................................................ vii
LIST OF TABLE ................................................................................................... ix
LIST OF PICTURE ................................................................................................ x
CHAPTER I INTRODUCTION ............................................................................. 1
1.1 Background .............................................................................................. 1
1.2 Problem Statement ................................................................................... 5
1.3 Research Objective ................................................................................... 7
CHAPTER II LITERATURE REVIEW ................................................................. 8
2.1 Surfactant in General ................................................................................ 8
2.2 Characteristics of plant-derived saponin ................................................ 10
2.3 Saponin as plant derived surfactant ........................................................ 14
2.4 Sapindus rarak DC ................................................................................. 17
2.5 Saponin Extraction ................................................................................. 18
2.6 Reactive Dye .......................................................................................... 21
2.7 Polyethersulfone Ultrafiltration Membrane ........................................... 23
2.8 Fundamentals and Application of Micellar-Enhanced Ultrafiltration
Membrane (MEUF) ........................................................................................... 24
CHAPTER III RESEARCH METHODS ............................................................. 29
3.1. Research Design ......................................................................................... 29
3.2. Variables .................................................................................................... 31
3.3. Response and Observation ......................................................................... 32
3.4. Research Implementation ........................................................................... 33
3.5. Figure ......................................................................................................... 34

vii
 3.6. Research Procedure .................................................................................... 35
CHAPTER IV RESULT AND DISCUSSION ..................................................... 45
4.1. The investigation of saponin content extracted from Sapindus rarak DC . 45
4.2. Characterization of saponin as surfactant .................................................. 52
4.3. Dye Solubilization...................................................................................... 59
4.4. Permeate flux profile of surfactant-enhanced ultrafiltration system for dye
removal.............................................................................................................. 65
4.5 Rejection of dye and saponin residue.......................................................... 69
4.6 Performance of Surfactant in MEUF system: Micelle Loading an
Equilibrium Distribution Constant .................................................................... 72
4.7 Model of Fouling Mechanism ..................................................................... 75
CHAPTER V CONCLUSION AND SUGESTION ............................................. 81
5.1 Conclusions ................................................................................................. 81
5.2 Sugestions ................................................................................................... 82
REFFERENCE ...................................................................................................... 83

viii
 LIST OF TABLE

Table 1.1. Selected Saponin Extraction Method and Their Application 6

Table 2.1 High Saponin Content Derived from Plant 14

Table 2.2 Physical Structure of Remazol Dye 22

Table 2.3 Membrane Specification 23

Table 2.4. Application of MEUF for Dye Removal 28

Table 3.1 Linearisation equation of blocking/fouling models based on

Hermia’s model 44

Table 4.1 The optimized parameter of the saponin extraction by UAE and ME 51

Table 4.2 Common HLB value and its application 58

Table 4.3 The surfactant characteristic and properties of saponin extract from
Sapindus rarak and commercialized pure saponin 59

Table 4.4 Molar solubilization power (SP), ln Km and ΔG of solubilization of

pure saponin and extract of S.rarak for remazol dye, at 27oC 65

Table 4.5 Concentration of dye and saponin on the permeate after membrane
separation 70

Table 4.6 The effect of saponin concentration to the equilibrium distribution

constant and micelle loading for remazol red and blue 73

Table 4.7 Mathematical model parameter of UF and MEUF blocking

phenomena on indigo sol dye removal 76

ix
 LIST OF PICTURE

Figure 2.1 Structures of some representative surfactants 8

Figure 2.2 Structure of aglycones 12
Figure 2.3 Structural classification of saponin 12
Figure 2.4 Structural classification of saponin extracts in respect to,
(a) aglycone structure, (b) number of sugas chain 13
Figure 2.5 Basic structures of various saponins 15
Figure 2.6 (A) The tree of Sapindus rarak DC, (B) The fruits of Sapindus
rarak DC 17
Figure 2.7 Schematic representation of MEUF 25
Figure 2.8 Experimental parameters that influence the performance of MEUF 27
Figure 3.1 Schematic diagram for the whole study 29
Figure 3.2 Schematic diagram of saponin extraction from Sapindus rarak 30
Figure 3.3 Schematic diagram of MEUF process 31
Figure 3.4. The schematic diagram of the saponin extraction 34
Figure 3.5. Schematic Picture of Membrane Ultrafiltration 34
Figure 4.1 Effects of extraction time on the saponin content of the Sapindus
rarak extracts, at temperature of 30oC and solid-liquid ratio of 2 mL/gram 46
Figure 4.2 The effect of solvent to material ratio on the saponin content of the
Sapindus rarak extracts at 30oC for 120 minutes 47
Figure 4.3 The effect of extraction temperature on the saponin content of the
Sapindus rarak extracts at solvent to material ratio of 10 mL/g, for 40 min 49
Figure 4.4 Schematic illustration of (a) Saponin in plant tissue, (b) ME
mechanism and (c) UAE mechanism 51
Figure 4.5 The FTIR spectra of pure saponin and extract of saponin from
Sapindus rarak DC. 53
Figure 4.6 Molecular structure of Sapindus rarak (monodesmosidic oleanane
triterpenoid saponin) 54
Figure 4.7 The micelle formation in water solution, (A)Below CMC, (B)Right
at CMC, and (C) Above CMC 55
Figure 4.8 Surface tension of (a) Saponin extract in aqueous solution, (b) Pure
saponin in aqueous solution, at various concentration 55

x
Figure 4.9 The value of total initial foam (V0) and total foam after 6 hours
(V6) of saponin extract and pure saponin 58
Figure 4.10 Part of surfactant micelle in water 60
Figure 4.11 Effect of saponin concentration (in mM) on the solubility of (A)
Remazol red RB, and (B) Remazol Blue TQ 61
Figure 4.12 Solubilization of remazol dye in saponin micelle 64
Figure 4.13 Variation of the observed permeate flux of remazol red RB and Tq
Blue with time at room temperature and pressure of 1.5 bar 66
Figure 4.14 Rejection of remazol dye at various CMC, (A) rejection of dye,
(B)rejection of saponin 72
Figure 4.15 Schematic illustration of mechanisms of four blocking filtration
laws,adapted for cross-flow filtration 75
Figure 4.16 Illustration of membrane blocking based on the formation of
surfactant micelle, (a) without saponin, (b) below CMC, (c) at CMC,(d)above
CMC 78
Figure 4.17. FTIR Spectra of clean PES membrane and fouled membrane. 79
Figure 4.18 SEM figure of (a) clean membrane, and fouled membrane (b)
without saponin, (c) with saponin under CMC, (d) with saponin above
saponin 80

xi
 CHAPTER I
INTRODUCTION

1.1 Background
Micellar-enhanced ultrafiltration (MEUF) is one of the methods to remove
traces of organic pollutants from aqueous streams (Puasa et al., 2011). MEUF
utilizes a filtration process by membrane and micellization of surfactant. The basic
idea of MEUF is that a surfactant forms large amphiphilic micelles aggregates when
added to an aqueous streams at the concentration higher than its critical micelle
concentration (CMC). CMC is the concentration where micelle of surfactant started
to form. Surfactant have the tendency to spontaneously aggregate to form micelles
at concentration above the critical micellar concentration (CMC) in water (Acero et
al., 2017). This micellar formation generate a solubilization effect before the
ultrafiltration process. The process begins with the mixture of surfactants in the
effluent, allows its interaction with the dye. This interaction occurs through micellar
solubilization, where the solute, according to its affinity for aqueous medium, is
positioned within the micelle. Thus, its concentration in the form of free monomer
is decreased. When submitted to stirring, this precipitate aggregates to form flocs
which maintain the interaction of the surfactant with the dye. In this way, surfactant
floc acts as an adsorbent surface, retaining the dye inside the micelles (Zaghbani et
al., 2008).

Micelles containing solubilized solute are larger in size, with aggregation

number range from 50 to 100 (Bade and Lee, 2011). Thus, the pollutant molecule
inside the surfactant micelle can be rejected by the pores of ultrafiltration
membrane. Even if the original molecular weight could only be rejected by a lower
molecular weight cut-off (MWCO) nanofiltration (NF) or reverse osmosis (RO)
membranes. Leaving only water and a small amount of unsolubilized solutes with
free surfactants in the permeate stream, providing an effluent with less pollutant
load. The micelle solubilization ability was very dependent to the properties of the
surfactant used in the process.

In Indonesia, MEUF for wastewater treatment is highly needed, because

Indonesia is a country with a high textile demand and textile production. This

1
industry, approximately increased by 6% per year. About 75 % of the production
process is conduct in Indonesia (Saputra, 2018). The production and processing of
textile require great amount of chemical dyes. Azo dyes have been widely applied
in the textile, paper printing, plastics, and cosmetics industries, among others. Azo
dye is classified as a reactive dye and substantive dye. Some disadvantages
associated with reactive dyes are the use of high concentrations of electrolytes in
dyeing, presence of unfixed dyes in the substrate, resistance to light and resistance
to oxidizing agents. This characteristic makes reactive dye difficult to degrade once
released into aquatic systems (Melo et al., 2017). This undegradable chemical
substances used in the textile dyeing appears as mutagenic and carcinogenic
materials (Pelosi et al., 2014). Meanwhile, about 10-15% of the total dyes lost
during the dyeing process and is released in the textile industries wastewater
(Bielska and Szymanowski, 2006; and liu et al., 2017). The release of dyeing
wastewater with high intensity color causes aesthetic pollution and poses a great
threat to aqueous ecosystems (Liu et al., 2017). It affects the nature of water and
inhibits sunlight penetration, which reduce photosynthetic activity and lowering
oxygen concentration in the water. This phenomena may specifically disturb the
water ecosystem itself and human living generally. Reactive dyes also prove to be
toxic for the water environment and affect the lifespan of freshwater fish (Pelosi et
al., 2014). Hence, good wastewater treatment management is highly needed.

Membrane technology is known to be a good wastewater treatment methods.

Low molecular pollutant such as dye is widely treated by membrane nanofiltration
(NF) or reverse osmosys (RO). However, NF/RO requires high initial investment,
high operating pressure (8-12 bar), and very low permeability (Katheresan et al.,
2018). Compared to nanofiltration (NF), and reverse osmosys, MEUF possesing
high removal efficiency, high permeate flux, and lower energy consumption
(Purkait et al., 2004). The application of surfactant in MEUF process also shows
rejection profile almost similar to NF/RO membrane process (Das et al., 2008; and
Purkait et al., 2004). The operating pressure requirement is low compared to RO/NF
and membranes of higher permeability can be used. MEUF as a modified
ultrafiltration membrane separation process had been recognized as a promising
alternative. MEUF can also be chosen for its simplicity and efficiency. By far, this

2
method requires the least amount of chemicals compared to the biological or
chemical dye removal methods (Khan et al., 2018). MEUF does not deal with living
organisms hence is considered to be more predictable than the biological methods.
MEUF is able to treat textile effluent in a more beneficial way. However, the
suitable surfactant that compatible with the pollutant such as dye need to be chosen
to achieve good solubilization properties.

Surfactant-enhanced remediation technology has shown promising potential

for removing various organic and/or inorganic residual compound from the liquid
effluent. MEUF has been studied to remove heavy metal ions such as, chromate
(Bade et al., 2008), Cu2+, Cd2+, Zn2+, Pb2+ (Huang et al., 2017). It is also applied to
remove phenol and phenolic compound from wastewater (Victor-Ortega et al.,
2017; Li et al., 2011; and Luo et al., 2010). Hydrophobic organic chemicals
(HOCs), and polycyclic aromatic hydrocarbons (PAHs) also studied to be removed
from liquid effluent by MEUF system (Samal et al., 2017). As for reactive dye
removal, the method has been successfully applied for removal of Remazol dyes
(Aryanti et al., 2017), Methylene blue (Huang et al., 2009; and Khosa et al., 2013;
Bielska and Szymanowski (2006), Eriochrome Blue Black (Zaghbani et al., 2008),
a mixture of Reactive Black and Orange (Puasa et al., 2012) and eosin (Purkait et
al., 2006).
Although MEUF are known to be an effective method for removal of dye
pollutant from wastewater effluent, the potential problem about synthetic surfactant
for being toxic is still remain. Most common used surfactant is chemically
synthesized such as, hexadecylpyridinium chloride (CPC), Hexavalent trimethyl
ammonium bromide (CTAB) (Iqbal et al., 2006), Sodium dodecyl sulfate (SDS) (Li
et al., 2011; and Huang et al., 2017), Brij (Xu et al. 2007), Tween 80, Marlophen
NP5 (Bade and Lee, 2011) and many more. The degradation process of synthetic
surfactants are slow, some are persistent in nature and may cause estrogenic activity
and chronic toxicity. In MEUF, the used surfactant remains in retentate stream at a
high concentration which creates secondary pollution (Samal et al., 2017b).
Therefore, before disposing them to the water body, a post treatment process are
needed, which attributes additional cost to the process. Thus, the selection of

3
surfactants which can be easily degraded is very important. Moreover, a surfactant
with good pollutant solubilization need to be considered as well.

For the solution, plant derived biosurfactant such as saponin is suggested to

substitute the synthetic surfactant in the MEUF process as a solubilization agent.
Saponin has been classified as one of natural surfactants derived from plants that
appear to have potential to replace synthetic surfactant. The plants produce
saponaceous substances called saponins, which form lather or foam in water (Roy
et al., 1997). At concentration above CMC the saponins start to aggregate and forms
micelle. The micelles can solubilize pollutant such as dye in its micelle structure.
This process grows the molecular weight of the pollutant in the water solution,
making it easier to be rejected by the membrane pore. Unlike the synthetic
surfactant, plant derived biosurfactants such as saponin is naturally biodegradable.
Saponin has an excellent functional surfactant properties, biodegradable, renewable
and ecologically adaptable as well as environmentally safe (Roy et al., 1997; and
Schmitt et al., 2014).

Saponins are a widespread class of natural compounds found in a lot of plant

species. Various plants are found to be the source of saponin. The previous study
also found saponin on some of food based plant, such as green tea (Chen et al.,
2013; and Ye et al., 2015), soymilk, sugar beet, soy and chickpea, asparagus, marion
blackberry, strawberry, and plum fruit (Cheok et al., 2014). High concentration of
saponin also found in mostly saponaria and sapinduciae plants. Previously, several
triterpene saponins and acyclic sesquiterpene glycosides have been reported from
the pericarps of other Sapindus species, such as Sapindus mukurossi, Sapindus
emarginatus, and Sapindus delavayi, and many more (Morikawa et al., 2009).
Saponin also found in the several plant grow in Indonesia such as binahong leaf
(Rachman et al., 2016), inggu leaf (Ruta angustifolia L.) (Noer et al., 2017), white
Oyster mushroom (Zahro et al., 2013), and sprouts of green beans (Yohana et al.,
2009). One of non-food based saponin source is pericarps of lerak (Sapindus rarak
DC) (Morikawa et al., 2009). Sapindus rarak DC classified as one of the main
choice of saponin. It is a tropical plant suitable to be cultivate in Indonesia,
guaranted its availability.

4
1.2 Problem Statement
Taking into consideration about the potential of saponin as important
substances, great efforts are being made to obtain saponin-rich extracts. The
efficiency of the extraction process is usually influenced by several factors, such as
solvent concentration, solvent to solid ratio, temperature, extraction time, and also
the extraction methods (Sarvin et al., 2018). In general, the extraction techniques
employed in saponin extraction can be classified into two categories, the
conventional and the green technologies. The conventional extraction techniques
such as maceration, or the green technologies including ultrasound-assisted
extraction (UAE) (Heng et al., 2013). UAE has been successfully developed and
proposed to significantly reduced extraction times, reduce energy consumption, less
solvent requirement and higher extraction efficiency. (Vilkhu et al., 2008; Garcia-
Salas et al., 2010; and Majd et al., 2014). UAE methods to extracted bioactive plant-
derived compound has been conduct to extract anthocyanin (Oancea etal., 2013;
D’Alesandro., et al., 2013; and Alighourchi et al., 2013), antioxidant compound
(Rosello-Soto et al., 2015; and Xu et al., 2015), natural pigment (Yolmeh et al.,
2014), phenolics compound (Dai et al., 2010) and many others. However, the
application of UAE to extract saponin from Sapindus rarak DC is very limited.

The available studies of saponin extraction were only focused on the

extraction process and the phytochemical properties (Heng et al., 2014; Kurniawan
et al., 2011, Pasaribu et al., 2014). Some studies investigate saponin as an
antibacterial agents (Pasaribu et al., 2014; Wina et al., 2005) and bioactive
compound (Heng et al., 2014). However, to the author’s best knowledge, the
surfactant properties of saponin has not been investigated before. Furthermore, a
limited number of research are found to reviewing about the utilization of saponin
in the MEUF. Its ability to solubilized dye pollutant was also limited. To our best
knowledge, only one study of plant-derived biosurfactant as a surfactant in the
MEUF application is found. The plant-derived saponin, extracted from reetha
(Sapindus mukurossi) or soapnut was applied in MEUF to remove methyl violet
(Samal et al., 2017). The application of saponin extracted from Sapindus rarak DC
in the MEUF process is nowhere to be found. Table 1.1 present the previous study
of saponin extraction from various plant source and its application.

5
 Considering this condition, the present studies will provide a new scarce
research having a less or more novelty compared to the previous work. This study
was projected to be focused for investigating the new proposed method for dye
removal from wastewater effluent employing the combination of micellar-enhanced
ultrafiltration methods with saponin from Sapindus rarak DC as the plant derived
bio surfactant. Moreover, the ability of saponin micelle to solubilize dye pollutant
was also investigated to get the information of saponin micelle solubilization
power.

Table 1.1. Selected Saponin Extraction Method and Their Application

Source of Extraction Methods Application Reference

Saponin
Sapindus Maceration by water, Soil washing Roy et al.,
mukorossi methanol, ethanol, and containing 1997
benzene:methanol (1:3) Hexachlorobenzene
mixture as solvent (HCB) and
Naphthalene
Aescin, n.a Soil washing Urum and
lecithin, containing Pekdemir,
rhamnolipid, crude oil 2004
saponin and
tannin
Saponin n.a Soil washing Song et al.,
contaminated 2008
phenanthrene and
Cadmium
Sapindus Maceration with Antioxidant Tmakova et
mukorossi methanol for 6 hours compounds al., 2016
Gaertner under stirrer
Allium UAE of dry ground Antifungal and Mostafa et
nigrum L. samples for 3 times 30 aginoside compound al., 2013
min
Ziziphus UAE at high intensity Only for qualitative Guo et al.,
jujuba (40 kHz) for 30 min and quantitative 2011
analysis of saponin
Sapindus Maceration with water Removal of cationic Samal et al.,
mukorossi for 24 hours and anionic dyes 2017a

6
1.3 Research Objective
This research are design to study the process of micellar-enhanced
ultrafiltration (MEUF) with plant derived biosurfactant. To be specific, the
objectives of this research are,

1. To investigate the effect of extraction parameters on saponin extracted from

the pericarps of Sapindus rarak DC and characterize the surfactant
properties of saponin extracted from Sapindus rarak DC.

2. To investigate the ability of saponin to solubilized the dye pollutant.

3. To applied natural-based surfactant (saponin) from Sapindus rarak DC to

remove reactive dye in micellar enhanced ultrafiltration process.

7
 CHAPTER II
LITERATURE REVIEW

2.1 Surfactant in General

The word surfactant is an abbreviation for surface active agent. The term
amphiphile is sometimes also used synonymously with surfactant. The term relates
to the fact that all surfactant molecules consist of at least two parts, one part which
is soluble in a specific fluid (the lyophilic part) and one which is insoluble (the
lyophobic part). When the fluid is water one usually talks about the hydrophilic and
hydrophobic parts, respectively. The hydrophilic part is often referred to as the polar
head group and the hydrophobic part as the tail. The primary classification of
surfactants as cationic, anionic, nonionic and zwitterionic (amphoteric) is made on
the basis of the charge of the hydrophilic group, which is either ionic or nonionic
(Jonsson et al., 1998). The hydrophobic group is normally a hydrocarbon chain and
the majority are linear to meet the demands for biodegradability (Figure 2.1) (Berg,
1999).

Figure 2.1 Structures of some representative surfactants:

(a) Alkyl ethercarboxylate (Anionics have carboxylate, sulfate or phosphate
surfactants as polar head groups), (b) Fatty amine salt (Cationics are based on
amine or quanternary ammonium groups), (c) Ethoxylated fatty alcohol (Non-
ionics have either ether or polyhydroxyl as polar group), (d) Betaine
(Zwitterionics contain both an anionic and a cationic charge under normal
conditions)

A surfactant is characterized by its tendency to adsorb at surfaces and

interfaces. Examples of interfaces involving a liquid phase include suspension

8
(solid-liquid), emulsion (liquid-liquid) and foam (liquid-vapour). In many
formulated products several types of interfaces are present at the same time.
Another general and fundamental property of surface active agents is that
monomers in solutions tend to form aggregates, called micelles. Micelles are
already formed at very low surfactant concentrations in water (Bergh, 1999). The
concentration at which micelles start to form is called critical micelle concentration
(CMC). Micelle formation, or micellization, can be viewed as an alternative
mechanism to adsorption at the interfaces for removing hydrophobic groups from
contact with the water, thereby reducing the free energy of the system (Schwarze,
2017). It is an important phenomenon since surfactant molecules behave very
differently depending on whether they are present in micelles or as free monomers.
The micelles behave as large molecules and influencing the solubility of organic
hydrocarbons and oils in aqueous solution and also influencing the viscosity. The
size of the micelle is measured by the aggregation number which is the number of
surfactant molecules associated with a micelle (Pisarcik et al., 2015).

Many surfactants are available to be used in MEUF process such as SDS

(anionic), TX-100 (nonionic), CPC or CTAB (cationic). The surfactants are widely
applied in the filtration process due to the following reasons: a) the surfactants are
available in higher quality, b) the surfactants have already been well characterized,
c) their performance has already been proven with similar types of solutes, and d)
literature results are available for comparison (Schwarze, 2017).

For an MEUF process, the surfactant type, the critical micelle concentration
(CMC), and the micelle size are important factors. The surfactant type determines
which kind of solute, ions and/or organics can be removed in MEUF, the CMC
gives information about the minimum amount of surfactant that is needed to form
micelles, and the micelle size affects the membrane selection with respect to the
molecular weight cut-off (MWCO). The CMC is characterized by a spontaneous
change in the physical properties of the solution, e.g. surface tension or electrical
conductivity (Schwarze, 2017). The size of the micelles is usually obtained from
dynamic light scattering (DLS) experiments, where the diffusion coefficient is
measured and used to calculate the hydrodynamic radius “r” (Tummino and Gafni,
1993; Materna et al., 2004).

9
 If nonionic surfactants are applied in MEUF, the cloud point temperature
(CPT) that increases with the hydrophilic character of the surfactant is also an
important parameter to avoid temperature-induced phase separation during the
filtration process (Bergh, 1999). For too low CPTs, cloud point extraction (CPE) is
an alternative surfactant-based separation method for solute removal (Materna et
al., 2004). The selection of a surfactant depends mainly on the separation task. If
ions should be removed from the water, anionic surfactant with the opposite charge
is selected to form an ion-pair complex that is removed during ultrafiltration
(Mungray et al., 2011). Generally, the CMC of ionic surfactants is very high and,
during the filtration process, a surfactant concentration equal to the CMC is
discharged into the water phase. This will be a problem in water treatment plants as
the chemical or biological oxygen demand will increase due to the leached
surfactant that will become a “contaminant”. In contrast, the CMC of nonionic
surfactants is very low but they cannot form an ion-pair complex; therefore, they
are usually applied to the removal of hydrophobic non-charged solutes or where
ligands are applied to complex ions before their removal via MEUF (Schwarze,
2017). However, in some cases, ions can also be removed with nonionic surfactants
for nickel ions (Tanhaei et al., 2014).

2.2 Characteristics of plant-derived saponin

In general, surface active agents (surfactants) are a group of substance that
can accumulate at the surface of a liquid, or interface between two phases. Its role
is to change the surface tension of the interface. Surfactants are changing the surface
tension of a solution by minimizing the effective free surface energy of the
interfaces (Tmáková et al., 2015).

The molecular structure of surfactants are amphiphilic molecules consisting

of a hydrophobic chain and a hydrophilic head group (Schwarze, 2017). The
hydrophobic group has low affinity with water and the hydrophilic group has strong
affinity with the water. The hydrophillic end attract water and the hydrophobic end
repel water (Boruah and Gogoi, 2013). Its molecule is typically a long hydrocarbon
chain or an aromatic ring with a functional group acting as its head. Commonly, the
tail is derived from an unsaturated high molecular weight fatty acid or its

10
derivatives. The presence of an electrophilic or nucleophilic group is responsible
for its hydrophilic nature; consequently, the tail acts as a hydrophobic end of the
molecule (De and Mondal, 2012)

For a compound to be a surfactant, it should possess three characteristics. First

of all, the molecular structure should be composed of polar and nonpolar groups. It
also should exhibit surface activity, and it should form self-assembled aggregates
(micelles, vesicles, liquid crystalline, etc.) in liquids. Based on the type of head
group, surfactants are classified into four categories: anionic, cationic, nonionic,
and amphoteric. And based on the availability there are two types of surfactant;
synthetic and natural surfactant. Anionic and cationic surfactant are mostly
synthetics. As for natural surfactant is usually categorized as nonionic surfactant.

Natural surfactants are the naturally available substances present in plants and
animal sources (star fish, sea cucumber) (Tmakova et al., 2015) which can lower
the surface tension. One of the chemically natural surfactant substance is saponin.
Saponins are second metabolites which are widely distributed in the plant kingdom.
It is a natural phytochemicals, which occur in three out of four 4 plant species.
Saponins acts as a chemical barrier or shield in the plant defense system to counter
pathogens and herbivores. It has a bitter taste, which protects the plant from being
eaten by animals (Cheok et al., 2014). For several decades, saponins in plant
materials were considered undesirable ingredients in human and animal nutrition
because of their bitter taste and detrimental effects on the gastro-intestinal system
when consumed at high concentrations. But recently, researchers regained interest
in saponins due to their interfacial activity and unique behavior in dispersed
systems.

The empirical formula for saponin is C26H31O10. It is mainly glycoside having

glycosidic structure of surfactant molecule comprising two ends, glycone and
aglycone (sapogenin). Predominantly, the glycone structure can be occurred as one,
two or three sugar chains attached to the aglycone. Where the terms
monodesmosides, bidesmosides or tridesmosides have been given to them,
respectively (Greekdesmos=chain). The aglycones, also called sapogenins are the
nonpolar parts of the molecule. Based on its aglycone backbone, saponins also
divided into two major classes which are triterpenoid and steroid glycosides. Which

11
their structure characterization are varied by the numbers of sugar units attached at
different positions (Hostettmann & Marston, 1995).

Triterpene aglycones Steroid aglycones Diosgenin

Figure 2.2 Structure of aglycones (Glstnda et al., 2007)

Figure 2.2 shows the molecular structure of monodesmosidic saponins with a

sugar chain attached usually at C-3 of the aglycone. The bidesmosides have two
sugar chains, most often with one attached through the ether linkage at C-3 and one
attached through the ester linkage at C-28 in triterpene saponins or an ether linkage
at C-26 in steroidal saponins. The tridesmosides may have a third sugar chain linked
through an ether or ester link at one of the OH or COOH functional groups
occurring on the aglycone (Oleszek et al., 2009). The classification and occurrence
of saponins in the plant kingdom are reviewed in detail in the previous study, for
references, the structural classification of saponin are present in Figure 2.2 – 2.4
(Böttcher and Drusch, 2017).

Natural Surfactant

Aglycone
Glycone (Sapogenin)

Sugar Chain Steroids Triterpenoids

Mono- Bi- Tri-

desmosides desmosides desmosides

Figure 2.3 Structural classification of saponin

12
 (a) Aglycone Type

Triterpenoid Steroid
 Yucca schidigera
 Tribulus terrestris (TT)

Ursolic & Dammarane

Oleanane type Oleanane type type
 Quillaja  Ilex  Panax ginseng
Saponaria paraguariensis
Molina
 Camellia oleifera
Abel
 Glycyrrhiza
glabra
 Sapindus
Mukorossi

Number of
(b) Sugar Chain

Mono- Mono- & Bi-

desmosidic Bidesmosidic desmosidic
 Aesculus hippo-  Tribulus  Quillaja
castanum terrestris saponaria
 Camellia oleifera  Sapindus Molina
Abel Mukorrosi  Gypsophia
 Glycyrrhiza  Yucca schidigera species
glabra  Panax Ginseng

Figure 2.4 Structural classification of saponin extracts in respect to (a) aglycone

structure, (b) number of sugas chain

Many natural surfactants can be obtained from root, flower, leaves and fruits
from the plant. As reviewed by Boruah and Gogoi (2013), such as Aloe vera,
asparagus, balloon flower (Platycodon grandittonem), china rose (Hibiscus rosa
sinesis), Daisy, Ginseng (Panax araliaceae), Horse chestnut (Aesculus
hippocastanum), Spinach, Tea, Tomato, Soapbark (Quillaja saponaria), Soapnut/
soap herry (Sapindus mukorossi), and Soapwort (Saponaria officinalis). In more
specific, Table 2.1. lists many sources of saponin from the plant.

13
 Table 2.1 High Saponin Content Derived from Plant (summarised from Oleszek
and Hamed, 2010)
Plant Source Common Use Plant Part Concetration
Name (%)
Quillaya Quaillaya Soaps, Bark >25
saponaria Murillo foaming
agents
Balanites Heglig, lalob, Diosgenin Fruits, 22-27
aegyptiaca desert date and seeds, and
yamogenin bark
source
Chlorogalum Soaproot, Amolonin Bulb 19-22
pomeridianum California source
soap plant
Tribulus Puncturevine, Protodioscin Fruits >20
terrestris yellow vine source
and goathead
Sapindus Soapnut Hedragenin Fruits 20
mukorossi Source pericarps
and roots
Glinus lotoides Soap Jacob, Hopane and Roots, 16.5
lotus oleanane leaves,
sweetjuice source seeds
Sapindus Soapberry Hedragenin Fruits 11
saponaria Source
Yucca Mohave yucca, Soaps, Stalk and 10
schidigera Joshua tree foaming roots
agents

2.3 Saponin as plant derived surfactant

As a surfactant, the ability of a saponin to foam is caused by the combination
of the nonpolar sapogenin and water-soluble side chain. This characteristic is
similar to the structure of most synthetic surfactants having lipophilic and
hydrophilic molecular parts. In synthetic surfactants lipophiles are usually similar
from one surfactant to another (i.e. straight or branched alkyl chains) but
hydrophiles show a range of chemical types. This has been the basis for surfactant
classification as anionic, cationic, nonionic and amphoteric. In the case of saponins,
hydrophiles are built of a sugar chain, which can differ in the length, branching,
substitution and composition (glucose, galactose, rhamnose, arabinose, xylose,

14
apiose and uronic acid), while the lipophiles may have a steroidal or triterpene
structure. Hence, saponins of this composition are nonionic surfactants (Oleszek et
al., 2009).

(a) Steroidal saponin from

Yucca schidigera

(b) Hederagenin glycosides

from Sapindus
Mukorossi

(c) Soyasaponin I from

soybean (Glycine max)

(d) Triterpenoid saponin of

oleanolic acid

Figure 2.5 Basic structures of various saponins

15
 Saponins with one sugar chain have the best foaming characteristics. For
saponins with two or three sugar chains the foaming ability decreases and in some
saponins no foaming in water solution has been observed, but due to their chemical
structure they are still considered as saponins. The emulsifying properties of
saponins are due to the fact that they have a salt-free nature, making them less likely
to be affected by alkaline or acid conditions (Oleszek et al., 2009).

In water solution saponins form micelle-like aggregates above its critical

micelle concentration (CMC) (Grzegorsek and Majewska-Nowak, 2018). Below
this concentration, molecules remain unassociated. An abrupt change in physical
properties appears when the concentration surpasses the CMC and the solute starts
to form micelles. The size and structure of saponin micelles are dependent on the
type of saponin. Commercial ‘saponin white’ from Saponaria officinalis and soya
bean saponins form small micelles consisting of only two molecules, while the
aggregates of saponin of Quillaya saponaria consist of 50 molecules, and appear to
be significantly less hydrated (Oakenful, 1986). This differences are quite
unexpected as aglycones of quillaya saponin differ from the aglycone of ‘saponin
white’ only by one hydroxyl group. Presumably they aggregate by hydrophobic
interaction of their aglycones (as for other surfactants), leaving the hydrophilic
sugar groups exposed to the water.

The shape of the micelles also depends on the saponin structure. The micelles
formed by ‘saponin white’ and Quillaya saponins appear as elongated and
filamentous, while those formed by soya bean saponins appear spherical (Oleszek
and Hamed, 2009). The microscopic analysis conduct by Samal et al. (2017) also
shows a spherical structur of Sapindus saponin. The reason for this differences is
assumed to be the aglycone structures. The soya bean aglycones do not possess
carboxylic functions, unlike the aglycones of Saponaria officinalis and Quillaya
saponaria saponins, therefore they are more uniformly hydrophobic. The presence
and the location of carboxylic acid in the saponin molecule also strongly influence
the surface activity, emulsion stability or zeta potential of the emulsion droplet (De
and Mondal, 2012).

Saponins can also form mixed ‘sandwich-like’ or ‘pile of coins like’ micelles
with bile acids. This structure are much larger than the micelles of saponins alone

16
and again they differ depending on the structure of the aglycone. Saponin white and
Quillaya saponin form filamentous structures with bile acids, while the soya bean
saponins have a loose, open structure with considerable penetration of water
(Oleszek and Hamed, 2009).

2.4 Sapindus rarak DC

Lerak or Sapindus rarak DC is a subtropical dicotyledonous plant that can
grow up to 8 to 40 meters with stem diameter up to a meter. Lerak grows well in
almost all types of soil condition, from lowlands to mountains with altitudes of 450-
1500 m above sea level. This plant originated in South East Asia and is now widely
distributed in Asia and Africa. Nowadays, Sapindus rarak DC trees grows wild on
the island of Java at an altitude between 450 to 500 meters above the sea level (Wina
et al., 2005). The shape of the leaves is rounded pointed, flat-edge, short-stemmed
and green. The seeds are wrapped in skin quite hard round like marbles, if they are
ripe the color is blackish brown, the surface of the fruit is smooth and shiny. Figure
2.6 shows the appearance of Sapindus rarak DC.

(A) (B)

Figure 2.6 (A) The tree of Sapindus rarak DC, (B) The fruits of Sapindus rarak
DC.

The fruit is a hard fruit, with dicotyledonous seed. Its fruits have pericaps that
are soft and brown in color and become dark brown when they are dried. Lerak fruit
is a seasonal fruit. It is commonly preserve in dry state until used. The fruit pericarp
is widely used as a natural detergent and its extract is commercially utilized

17
as a foam stabilizing and emulsifying agent in cleansers, shampoos and
cosmetics. In Indonesia, it is also used as a substitute for soap and is mainly used
for washing batik cloth. Because of its nature, larak soap does not cause fading
(fading) on the fabric. The reason is that the pericarp is abundant in saponins, which
gives a high surface activity (Morikawa et al., 2009).

The major saponins in the pericarp of Sapindaceae plant is triterpenoid-type

(Sharma et al., 2013). Lerak fruit contains several compounds including 12%
saponin, 1% alkaloid, 0.036% steroid, and 0.029% triterpenes (Ismail et al., 2012).
The crude leaching liquor from the pericarp of other genus of sapindaceae (S.
mukorossi Gaerten.) has complex ingredients such as saponins, saccharides,
proteins and other unknown ones (Li et al., 2013).

2.5 Saponin Extraction

The recent advances in extraction of bioactive compound including saponin
from plant material have been intensively reviewed (Azmir et al., 2013). Due to the
abundance of saponins in nature, a wide range of plant materials can be used as raw
materials for commercial production of saponins. The first step in the processing of
saponins involves their extraction from the plant matrix. As in any extraction
process, the extraction solvent, extraction conditions (such as temperature, time,
pH, solvent to feed ratio), and the properties of the feed material (such as
composition and particle size) are the main factors that determine process efficiency
and the properties of the end product.

In general, the extraction techniques employed in saponin extraction can be

classified into two categories, the conventional and the green technologies. Some
conventional methods to extract saponin from the plant tissue is maceration, reflux,
and soxhlet. This conventional extraction methods is relied on the solubility of
solute from plant materials into solvent. Therefore, it often utilizes a large quantity
of solvent to extract the desired solute, even though sometimes is aided with
elevated temperature by heating, and mechanical stirring or shaking. While, the
green technologies are ultrasound-assisted, microwave-assisted, and accelerated
solvent extraction (Heng et al., 2013). The green extraction techniques involved

18
less hazardous chemical synthesis, safer chemicals used, energy efficiency, use of
renewable feedstock, and pollution prevention (Azmir et al., 2013).

The maceration extraction is a solid–liquid extraction where the bioactive

compound (solute) inside the plant material is extracted by soaking the plant
material in a specific solvent for a period of time (Takeuchi et al., 2009). The
efficiency of maceration process is determined by two main factors, solubility and
effective diffusion. The solubility is governed by basic rule of “like dissolves like”
which indicated that polar compounds dissolve in polar solvents, and nonpolar
compounds dissolve in nonpolar solvents (Reichardt & Welton, 2011). The rate of
dissolution of a solute in the extraction solvent is determined by the rate of mass
transfer of a solute from the plant material to the solvent (Takeuchi et al., 2009).
Generally, no complicated utensil and equipment are needed for the set-up of a
maceration extraction system that has made it a popular choice for researchers. The
only paramount factor to be paid attention in enhancing extractability is the
knowledge of similarity of bioactive compound interest and solvent polarity.

Reflux and soxhlet extraction is similar proces. The difference between reflux
and Soxhlet is that Soxhlet apparatus consists of a thimble to house the plant
material. Reflux and Soxhlet extraction involved distillation process which is
widely used in food and nonfood industrial and laboratories. The process involves
heating a solution to boiling and then returning the condensed vapors to the original
flask (Bart, 2011).

In contrast with the conventional methods, the green technology requires up-
to-date equipment such as ultrasound, microwave and accelerated solvent
extraction method. But, green technology is known to gives a better extraction
results. Ultrasonic technology has been widely used in food physical processing
mainly through cavitation phenomenon (Verhaagen and Rivas, 2016; Shanei and
Shanei, 2017). Ultrasound has many advantages such as avoiding of chemical use
and keeping the flavor of foods original. High heat and mass transfer improve the
extraction or degradation efficiency of the effective food components (Kielczyn’ski
et al., 2014; Jordens et al., 2014). Oscillation of cavitation bubbles occurs due to
the interaction between ultrasound and material, and the collapse of the oscillated
bubbles introduces shock waves. At the same time, high temperature and pressure

19
appear in the local area. Free radical might be introduced via ultrasonic cavitation
processing, which changes the material structure (Samani et al., 2016; Colivet et
al., 2016). The ultrasonic extraction is a typical technology of physical processing.

The main driving force for the extraction effects of sonication is acoustic
cavitation. When ultrasound propagates through any medium, it induces a series of
compressions and rarefactions in the molecules of the medium. Such alternating
pressure changes cause the formation and, ultimately, the collapse of bubbles in a
liquid medium. This phenomenon of creation, expansion, and implosive collapse of
microbubbles in ultrasound-irradiated liquids is known as “acoustic cavitation”
(Tiwari, 2015).

Cavitation bubbles formed are roughly divided into two types, namely
transient cavitation (inertial) and stable cavitation (noninertial). Stable cavities are
relatively long-lived gas bubbles and exist for many cycles of compression and
rarefaction (Park et al., 2003). Transient cavitation or inertial cavitation bubbles
exist for a very short period, sometimes less than one cycle, and collapse violently
(Fuleki et al., 2003; Oberholster et al., 2009). There are many thousands of such
bubbles in a liquid, some of which are relatively stable, but others expand further
to an unstable size and undergo violent collapse to generate temperatures of about
5000 K and pressures of the order of 50 MPa (Liang et al., 2008; Tiwari, et al.,
2015) at a minuscule level. Theoretical calculations using hydrodynamic models of
cavitational collapse have reported temperature and pressure estimates of 2000–
10,000 K and 100–1000 MPa, respectively (Corrales et al., 2009). The temperature
and pressure changes that occur from this implosions causes shear disruption,
thinning of cell membranes and cell disruption, resulting in enhanced solvent
penetration into cells and amplification of mass transfer of target compounds into
the solvent. The implosion of cavitating bubbles also generates turbulence at a
microscopic level, high-velocity inter-particle collisions and agitation in
microporous particles of the matrix, which accelerates the diffusion (Salleh-Mack
and Roberts, 2007; Tiwari et al., 2015). Ultrasonic waves also facilitate hydration
and swelling of the matrix with an enlargement of pores that increases the diffusion
of solvent into the matrix and increases mass transfer. Hydration and swelling
effects are advantageous when a dry matrix is used for UAE.

20
 The ability of ultrasound to cause cavitation depends upon its characteristics
(e.g., frequency and intensity), medium properties (e.g., viscosity and surface
tension) and ambient conditions (e.g., temperature and pressure) (Falleh et al.,
2012). The high-shear energy created by the cavitation effect is employed for
applications including particle modification in liquids, release of cellular
components and molecular structures, and de-aeration of liquids and surfaces. For
extraction applications, formation and collapse of cavitation bubbles also depend
on solvent properties. For example, vapor pressure governs intensity of collapse;
surface tension and viscosity govern the transient threshold of cavitation. Chemical
reactivity of the solvent dictates the primary and secondary sonochemical reactions
(Tiwari, 2015).

2.6 Reactive Dye

Reactive dyes are a highly successful class of modern synthetic dyes due to
their wide shade gamut, their flexibility in application and the excellent fastness
properties they offer when dyed on wool, silk, cotton and regenerated cellulosic
fibres (Ahmad et al., 2006). Reactive dyes may be loosely defined as chromophores
which contain pendant groups capable to form covalent bonds with nucleophilic
sites in fibrous substrates. This covalent bonds are strong enough to encounter the
harsh condition in laundering process resulting to the outstanding coloration
properties. Most dyes possess colour because they, 1) absorb light in the visible
spectrum (400–700 nm), 2) have at least one chromophore (colour-bearing group),
3) have a conjugated system, i.e. a structure with alternating double and single
bonds, and 4) exhibit resonance of electrons, which is a stabilizing force in organic
compounds (Abrahart, 1977). When any one of this features is lacking from the
molecular structure the colour is lost. In addition to chromophores, most dyes also
contain groups known as auxochromes (colour helpers), examples of which are
carboxylic acid, sulfonic acid, amino, and hydroxyl groups. While they are not
responsible for colour, their presence can shift the colour of a colourant and they
are most often used to influence dye solubility. Additionally, dyes are different with
pigments. With regard to their solubility, organic colourants fall into two
classes, viz. dyes and pigments (Allen 1971). The key distinction is that dyes are
soluble in water and/or an organic solvent, while pigments are insoluble in both

21
types of liquid media. Dyes are used to colour substrates to which they have affinity.
Pigments can be used to colour any polymeric substrate but by a mechanism quite
different from that of dyes, in that surface-only colouration is involved unless the
pigment is mixed with the polymer before fibre or moulded article formation.

Table 2.2 Physical Structure of Remazol Dye (Pubchem, NCBI, U.S)

Chemical Remazol Turquoise Blue, Remazol Remazol Red RB-133

Name Blue TQ
Molecular C40H27N9O17S6 C27H18ClN7Na4O15S5
Formula
Molecular 1098.062 g/mol 668.999 g/mol
Weight
Molecular
Structure

Soluble in Yes Yes

water

Reactive dye is the most common dyes textile dyeing, including remazol dye
for common cotton, naphtol for jeans, indigosol dyes for bright color and many
more. Reactive dyes is different from other types of dyes because it forms a covalent
bond with the substrate. It is normally applied in cotton dyeing. Some have
complicated aromatic structures that resist degradation in a conventional
wastewater treatment process. It is because of their stability to sunlight, oxidizing
agents and microorganism (Ahmad et al., 2006). The reactive dye used in this study

22
is Remazol dye. The molecular structure and its physical properties are present in
table 2.2

2.7 Polyethersulfone Ultrafiltration Membrane

In recent years, the synthetic polymeric membranes have been used for a wide
variety of liquid separations such as nano-filtration, reverse osmosis, ultrafiltration
(UF) and microfiltration. The UF membrane technology has found growing
applications in various industrial processes such as food, paper, chemical,
pharmaceutical, biotechnological, wastewater purification, and seawater
desalination to retain desirable and undesirable water components including
proteins, viruses, colloids, and pathogens (Kumar, et al., 2013; Urtiaga et al., 2013;
Shockravi et al., 2017).

Table 2.3 Membrane Specification (Sterlitech, U.S)

Molecular structure
(in general)

Membrane TF (thin film) ultrafiltration membrane

Pore Size (MWCO) 10.000 Da

Typical operating pressure 0.7-8.3 bar

Recommended pH 3-9

Maximum temperature 55o Celcius (131o F)

Among different polymeric membranes, polyethersulfone (PES) membranes

have been extensively utilized various separation and purification process. This
polymeric membrane have high thermal resistance, excellent hydrolytic stability,
and ability to retain mechanical properties in hot and wet environments. Other
literature states, PES have an excellent chemical stability as well as good

23
mechanical property (Marchese et al., 2003). In general, pure PES membrane
exhibits high hydrophilicity properties, which can increase the permeation and
antifouling properties of the UF membranes. However some fabricate PES indicates
a slightly hydrophobic nature (Shockravi et al., 2017). The specification of UF PES
membrane used in this research study present in the Table 2.3.

2.8 Fundamentals and Application of Micellar-Enhanced Ultrafiltration

Membrane (MEUF)
Micellar-Enhanced Ultrafiltration Membrane (MEUF) is a technology
that employs surfactant micelles to solubilize inorganic and organic pollutants
from the effluent stream. It is a membrane based separation technique mainly
investigated for wastewater treatment. This method allows dissolved matter, such
as organic molecules or ions, to be rejected by an ultrafiltration membrane due to
molecular weight enlargement using surfactants. The membrane used in this
process is ultrafiltration (UF) membrane. It is a membranes with pore sizes in the
range of 0.1 to 0.001 micron. Typically, UF membranes will remove high
molecular-weight substances, colloidal materials, soluble pollutant and organic and
inorganic polymeric molecules.

In MEUF, the surfactant is added to the aqueous stream containing

contaminants or solute (e.g. metal ion, organic materials, low molecular weight
solute) above its critical micelle concentration (CMC) (Landaburru-Aguire et
al., 2009). When the surfactant concentration exceeding the CMC value, the
surfactant monomers will assemble (Chung et al., 2009, Huang et al., 2012b) and
aggregate to form large amphiphilic transparent micelles (Luo et al., 2010). Above
the critical micelle concentration (CMC), surfactant monomers spontaneously
aggregate into micelles. The micelles are having a hydrodynamic diameter
significantly larger than the pore diameter of ultrafiltration membrane (Misra et al.,
2009; Chung et al., 2009; Huang et al., 2009). The contaminants or solute will
entrap in micelles if they tend to strongly attracted by micelle surface and will
solubilize in the micelle interior (Zaghbani et al., 2008). Micelles containing
solubilized contaminants with a larger diameter than membrane pore size will be
rejected by the membrane during ultrafiltration process leaving only water,

24
unsolubilized contaminants and surfactant monomers in permeate stream (Misra et
al., 2009; Zaghbani et al., 2008). Selection of surfactant is significant to ensure the
efficiency of MEUF process. Based on the literature study, the MEUF of
wastewater commonly can be classified into four categories; (i) MEUF using a
cationic surfactant, (ii) MEUF using an anionic surfactant, (iii) MEUF using a
nonionic surfactant, and (iv) MEUF using a mixed surfactant.

The micelle size is in the same order of magnitude as the pore sizes of
ultrafiltration membranes. By selection of an appropriate membrane for the
ultrafiltration process, the micelles can be rejected by the membrane whereby the
surfactant monomers pass through. This fundamental concept of MEUF was
already studied before for anionic surfactants and for a nonionic surfactant (Melo
et al., 2017; Grzegorzek and Majewska-Nowak, 2018). The main advantage of
MEUF over conventional ultrafiltration is that it can even rejected a low
concentrations molecules, which is usually too small to be rejected by ultrafiltration
membranes. The pollutant molecules can bind to the micelles because of ionic or
hydrophobic interactions and subsequently separated together with them in the
ultrafiltration step (Figure 2.7).

Figure 2.7 Schematic representation of MEUF

This process utilizes the high efficiency of reverse osmosis (RO) and high
permeates flux of ultrafiltration membrane (UF) (Baek et al., 2003). The main
principle of this process is to increase the size of pollutant molecules by forming a
complex with surfactant. Cationic or anionic surfactants are used for the removal
of inorganic pollutants. In this system, the surfactant forms micelles at critical

25
micelle concentration (cmc). The aggregation number ranges from 50 to 100. (Bade
and Lee, 2011). Micelle (cationic or anionic) has high electrical potential on its
surface where anionic or cationic pollutants can be bounded depending upon the
charge characteristic of the pollutants. When the solution containing micelle is
passed to the membrane ultrafiltration, micelle retains on the membrane surface.
Unbound ions and surfactant monomers pass through the ultrafiltration membrane
to the permeate side.

In general, the MEUF performance is explained by the flux profile and

%rejection. Flux is defined as the flow rate of a solution goes through the membrane
(permeate solution) per effective area of the membrane. While, rejection is the
amount of particles/substances that have been removed from the feedwater. The
performance of MEUF is not only based on simple relations but also a very complex
process that depends on a miscellaneous parameters. Solute rejection efficiency and
permeate flux depend on several parameter, such as the characteristics of added
solutes, membrane type, surfactant type, and various operating conditions. (Bade
and Lee, 2011, Schwarze et al., 2015). A schematic overview is given in Figure 2.8.
Many studies have been carried out to investigate the performance of MEUF and
how it is influenced by this different parameters. Commonly, there are no universal
experimental conditions that can be applied to MEUF experiments. In fact,
experimental parameters have to be selected based on the individual system.

26
 Material
Membrane
MWCO
Selection
Pore Size
Type
Surfactant CMC
Selection Aggregation structure & Size
Single or Mixed System

MEUF Operating Mode

TMP
Operating
pH
Condition
Temperature
Concentration
Ions
Organic Molecule
Solutes
Salt
Single or Mixed System

Figure 2.8 Experimental parameters that influence the performance of MEUF

Micellar enhanced ultrafiltration (MEUF) has been used for the removal of
various organic and/or inorganic pollutant from aqueous phase (Baek et al., 2003).
It is particularly effective for removal of single components such as Cd2+, Mn2+,
Zn2+, Cu2+, Cr3+, Pd2+, Al3+, and many other. It is also effective for simultaneous
removal of Ni2+ and Co2+, and Ni2+ and Zn2+. The application of MEUF for dye
removal has also been conducted before, Table 2.4 presents the previous study of
MEUF for dye removal.

27
 Table 2.4. Application of MEUF for Dye Removal

Membrane Surfactant Dye Result Reference

Compound
Polyethersulfone CPC 2x and Remazol COD Rejection: (Aryanti et
4x CMC Blue, 70% al., 2017)
Black, Dye Rejection
Yellow 96%
PVDF SDS (2,5 OMW (Olive Dye rejection (El-Abbassi
(Polyvinylidene kg/m3) mill 74% et al.,
fluoride) wastewater) 2011)
Hollow SDS Methylene Dye rejection (Zeng et al.,
membrane 8,27 mM blue 99,2% 2011)
(MB)
Polysulfone SDS Methylene Dye rejection (Huang et
8 mM blue 96% al., 2012)
(MB)
Cellulose CTAB Eriochrome Dye rejection (Zaghbani et
membrane 2 mM Blue 99% al., 2009)
Black r
Cellulose SDS 10- Saffarin T Rejection 99% (Zaghbani et
Membrane 10 100mM al., 2008)
kDa
Polimeric CPC RO5 and Reactive (Ahmad et
membrane 1 gr/L RO16 Black 599,7% al., 2006)
rejected,Reactive
Orange 1699,6%
rejected
Cellulose SDS Methylene SDS rejection (Zaghbani et
membrane 4 mM blue >97% al., 2008)
(MB)
Organic CPC (10 Eosin Dye rejection 70.45% (Purkait et
Polyamide kg/m3) al., 2004)

28
 CHAPTER III
RESEARCH METHODS

3.1. Research Design

This research is designed to obtain extract of saponin from Sapindus rarak with high
purity, and characterization of saponin extract as a surfactant. In this study, the
application of saponin extract to replace synthetic surfactant in the micellar-
enhanced ultrafiltration also performed. The performance of saponin to solubilize
dye was also conducted at various concentration. The schematic diagram for the
whole study was presented in Figure 3.1.
Materials Process Results
Extraction Process
Dried-ground
Raw Pericarps of Dried, Size
Sapindus rarak DC reduction pericarps of S.
rarak

Best condition*
Extraction Analysis Yield (Extract. methods,
Process temperature, time,
and ratio of L/S)
UAE & ME, t, T, ratio L/S
Saponin extract from
1st
extraction at best
Objective
condition
Characterization

Saponin Extract
Surfactant FTIR, Foam ability Specific Functional
analysis, Surf. Group, Foam ability,
Pure Commercial Characterization Tension, HLB
Saponin CMC HLB number

Dye Solubilization

Remazol Red RB, Solubilization Power

Micellar Dye Dye and Saponin 2nd
and Remazol Blue (SP), Partition Coef.,
TQ Solubilization Concentration Objective
ΔG
Various concentration

MEUF Process UF Performance Flux Pofile, %

MEUF process Analysis Rejection

Surfactant Micelle Loading and

3rd
Performance Distribution Coef.
Objective
Analysis Profile

Blocking Suitable Blocking

Mechanism Mechanism

*Best condition determined based on the higher saponin yield

Figure 3.1 Schematic diagram for the whole study

29
The research began with preparation of raw material, preliminary analysis,
extraction of saponin from Sapindus rarak, and characterization of saponin extract.
The detail of research activities are shown in Figure 3.2.

Pericarps of Sapindus rarak

Drying and Size Reduction

(50oC, 2 hours, 100 mesh)

Ready to use dry grounded

FTIR analysis
Sapindus rarak

SAPONIN EXTRACTION
Variables
Extraction methods : ME, UAE
Temperature : 30, 40, 50, 60 oC
Ratio of Solute-Solvent: 1:2, 1:5, 1:10,
1:15, 1:25, 1:50, 1:100 (gr/ml)

Filtration & Centrifugation

(10.000 rpm, 15 min.)

Supernatant solution
(Saponin Extract)

ANALYSIS OF SAPONIN SURFACTANT CHARACTERISTIC

 Total saponin analysis by UV  Foaming properties analysis
Spectrometer  CMC and HLB measurement
 FT-IR analysis  Solubilization of Dye

Figure 3.2 Schematic diagram of saponin extraction from Sapindus rarak

The further study is to apply the saponin extract in the MEUF process and
evaluation of MEUF process using saponin extract from Sapindus rarak. Figure 3.3
explain the schematic diagram of MEUF study using saponin-based surfactant.

30
 Reactive Dye Saponin Extract
Solvent
(Remazol Blue TQ, Remazol (0; 0.5; 1; 1.5; 2 times of
(Distilled water)
Red RB; 1000 ppm ) CMC)

Mixing
o
(26 C; 200 rpm; 30 min.)

Ready to use Spectrophotometric

Dye Model Solution Analysis

ULTRAFILTRATION
 Membrane : PES, 10 kDa
 Temperature : 26 0C
 Pressure : 2 bar
 Time : 120 minutes

 Analysis of permeate flux

 Analysis of dye concentration
in the permeate
 Membrane rejection
 Analysis of Micelle Loading

Figure 3.3 Schematic diagram of MEUF process

3.2. Variables

3.2.1. Variable on the extraction process

Control Variable
Drying Temperature : 50oC (Roy et al., 1997)
Drying Time : 2 hours (Roy et al., 1997)
Powder Size : 100 mesh (Sarvin et al., 2018)
Solvent : Distilled water (Sarvin et al., 2018)
Extraction Pressure : 1 atm
Max. Extraction Time : 2 hours (Ji et al., 2014; Sarvin et al., 2018)
Extraction methods : Maceration, Ultrasound-Assisted Extraction
Independent Variable
Time : 5, 10, 20, 40, 60, 90, 120 minutes
Ratio of solute-solvent : 1:2, 1:5, 1:10, 1:15, 1:25, 1:50, 1:100 (g/mL)
Temperature : 30, 40, 50, 60 oC

31
3.2.2. Variable on the Dye Solubilization

Control Variable
Temperature : room temperature (26 ±1 oC)
Pressure : 1 atm (Tehrani-Bagha et al., 2012)
Centrifugation speed : 4500 rpm (Samal et al., 2017 (b))
Centrifugation time : 1 hour (Samal et al., 2017 (b))
Dye Concentration : 10.000 ppm (Tehrani-Bagha et al., 2013)
Reactive dye : Remazol Blue Tq, Remazol Red Rb
Independent Variable
Saponin type : Pure commercialized saponin, extract saponin
Saponin concentration : 0.2 – 4 times CMC (under CMC, at CMC, above
CMC)

3.2.3. Variable on the Micellar-Enhanced Ultrafiltration (MEUF) process

Control Variable
Membrane material : Polyethersulfone (PES)
Molecular weight cut-off : 10 kDa (Sterlich, 2018)
Temperature : room temperature (26 ±1 oC)
Trans-membrane Pressure : 2 bar (Ahmad et al., 2006)
Filtration Time : 120 minutes (Huang et al., 2014)
Membrane area : 9,6 cm2
Carrier Solvent : Distilled water (Huang et al., 2014)
Dye Concentration : 300 ppm (Specification of Membrane)
Reactive dye : Remazol Blue Tq, Remazol Red Rb
Independent Variable
Saponin concentration : 0; 0.5; 1; 1,5; 2 times of CMC

3.3. Response and Observation

The response analysis and the observation in this research including, the analysis
on extraction process, the dye solubilization performance and its application as a

32
surfactant on the MEUF process. After the extraction, total saponin content analysis
was conducted to find the extraction condition having highest yield. The surfactant
characterization of saponin was performed by FTIR, surface tension analysis,
determination of CMC, and HLB number. The performance of saponin to
solubilized the dye molecule was tested and present as solubilization power (SP),
coefficient of distribution (Km), and Gibbs free energy (ΔG). The application of
saponin as a surfactant on the MEUF process is studied by flux analysis, membrane
rejection, and micelle loading (Lm) parameter. The membrane blocking analysis
was conducted by mathematical model of Hermia’s model. All of the saponin
analysis was compared with pure commercialized saponin.

3.4. Research Implementation

3.4.1. Materials

Dry pericarps of Sapindus rarak is the raw material use in this research, procured
from Sleman, Jogjakarta. The fruit is dark brown in color and globular in shape with
a diameter of 1 to 3 cm. Distilled water as solvent are produced by UPT UNDIP,
Semarang. Pure saponin in practical (P.A) grade is procured from Sigma Aldrich as
the comparative standard. Reactive dye (Remazol Red RB, and Remazol Blue TQ)
is used to make the model of dye wastewater. The dye is procured from dye
manufacturer in dry powder form. The membrane used for the MEUF process was
Polyethersulfone (PES) flat sheet membrane with MWCO 10 kDa procured from
Sterlitech.

3.4.2. Equipment

To make a dry powder of Sapindus rarak DC, oven and mechanic blender is used,
sieve of 100 mesh in size is used to make a homogeneous powder. An ultrasonic
bath (Krisbow) with temperature control is going to use for ultrasonic extraction.
The ultrasonic bath is equipped with beaker tray and lid. The supporting equipment
such as beaker glass, erlenmeyer glass, stirrer, sampling bottle, measurement flask,
graduated cylinder, etc. is used in the extraction, analysis and application process.
Some equipment is used for the post treatment process, such as filter cloth, whatman
paper filter, and centrifuge. It is equipped to separate the solid particles from crude

33
Sapindus rarak extract. The analysis process was conducted using various
apparatus such as Kruss automatic tensiometer, FTIR, and spectophotometric UV-
VIS. The MEUF process was conducted in a crossflow filtration equipped with
centrifugal pump, three tank, pressure gauge and pressure control.

3.5. Figure

Sapindus rarak DC

Centrifuge
Dried, Peeled

Ultrasonic-Assisted
Extraction SAPONIN
with controlled temperature, Analysis and
time and wavelength Kinetic Study
Grinded to powder

Figure 3.4. The schematic diagram of the saponin extraction

3 4
5

7
10

2
1 1
9 8

1. Feed Tank 6. Retentate Recycle Valve

2. Feed Pump 7. Permeate Recycle Valve
3. Feed Valve 8. Permeate Tank
4. Pressure Gauge 9. Retentate Tank
5. UF Membrane Module 10. Retentate sampling valve

Figure 3.5. Schematic Picture of Membrane Ultrafiltration

34
3.6. Research Procedure

3.6.1. Pretreatment

Before extracting the saponin from Sapindus rarak DC, pretreatment are conducted
to prepare the ready to use pericarps of Sapindus rarak DC in dry powder form. In
this pretreatment phase, some analysis also conducted to make sure the saponin
contain in the Sapindus rarak DC.

Dry-Grounded Sapindus rarak DC

The fruits of Sapindus rarak is prepared to obtain a clean and homogeneous sample
before used in the extraction process. The fruits is washed thoroughly with flowing
water to get rid of dirt. The seed and the pericarps are separated. The outer pericarps
of dry fruits are dried in an oven at 50 oC for 12 hours (Roy et al., 1997). The dry
pericarps are ground and sieved through 100 mesh sieve to obtain homogeneous
fine powder of Sapindus rarak. This ready to use powder are stored in the closed
vacuum storage by the temperature of 7 oC, silica gel are added to keep the powder
dry (Chen et al., 2013).

Preliminary Analysis

The preliminary analysis are conduct to evaluate the chemical structure and saponin
existence in the Sapindus rarak. Prior to the quantification of total saponins of a
plant source, it is propose to carry out a simple procedure to test the presence of
saponins in the sample of extracted matter. As much of 2-5 grams of the plant
material is put into a test tube filled with distilled water and vigorously shaken for
2 min (Ncube et al., 2011). The appearance of stable and persistent foam on the
liquid surface for 15 min indicated the presence of saponins. While, FT-IR analysis
is used to compare the chemical structure of pure saponin, and the dry-grounded
Sapindus rarak. For this, the FT-IR transmittance spectra of Sapindus rarak sample
are recorded and compared with the FT-IR transmittance spectra of pure saponin.

3.6.2 Extraction of Saponin from Sapindus rarak

The objective of this part of study is to obtain extract of Sapindus rarak with high
saponin concentration. Ready to use dry Sapindus rarak powder is extracted by two
methods, maceration extraction (ME) and ultrasound-assisted extraction (UAE).

35
Maceration Extraction (ME)

The maceration extraction of saponin from Sapindus rarak is performed according

to method by Samal et al., (2017). Dry powder of Sapindus rarak in various amount
ranging from 2-200 mL/g are mixed with distilled water as the solvent and
homogenized using magnetic stirrer at 30 rad/s for 1 minutes. The homogeneous
mixtures are then soaked for 2 hours at various temperature. A further post
treatments are perform after the extraction process. The suspense containing
solution is mixed using magnetic stirrer at 30 rad/s for 1 minutes to make a
homogeneous suspension again. Then the solution is filtered using filter cloth and
centrifuged at 4500 rpm for 1 hour to separate out the solid particles. The
supernatant solution is ready to use for further process and analysis.

Ultrasound-Assisted Extraction (UAE)

Ultrasound-assisted extraction was carried out in an ultrasonic bath device with a

constant power of 280 W, at the frequency of 40 kHz. The ultrasonic bath device is
equipped with digital sonication time and temperature control system. The dry
Sapindus rarak powder in the same range as in maceration extraction is placed in
capped plastic tube and mixed with distilled water as the solvent. The tube with
suspension is sonicated for maximum 2 hours in the ultrasonic device contenting 4
L water at various temperature (Ji et al., 2014; Sarvin et al., 2018). All the samples
in each extraction are placed in the center of ultrasonic bath with the depth of fifteen
centimeters and subsequent each batch of samples to be sonicated is kept the same
position. The same post treatments as conduct in the maceration extraction are
performed after extraction process.

3.6.3. Analysis of Saponin Extracted from Sapindus rarak

The spectrophotometric and chromatographic methods are usually employed to

detect and quantify the saponin compounds in the extract of Sapindus rarak.
Spectrophotometric methods give a total saponin value contain in the extract
(Cheok et al., 2014). A further analysis to evaluate the chemical structure of saponin
extract from Sapindus rarak is performed by FT-IR method.

36
 a. Spectrophotometric method

The most common methods applied for saponin quantification is using

spectrophotometer (Guo et al., 2018; Wu et al., 2018; Cheok et al., 2014). This
method is conduct based on the specific group of saponin detected by UV light of
spectrophotometer. Pure saponin purchase from Sigma, Aldrich is used as the
standard solution reference to make the calibration curves. The maximum
absorption wavelength is determined by UV-VIS Spectrophotometer. The
absorbance of 312 nm was found to be the maximum wavelength in this study (the
detailed data is presented in Appendix).

The preparation of the calibration curve is performed by analyzed the absorbance

of pure saponin solution in various concentration (5000-100 ppm).
Spectrophotometric analysis is performed by measuring the absorbance at the
maximum absorption wavelength using reagent blank as a reference. The regression
equation of the standard curve based on the concentrations (x) versus the
absorbance value (y) is made and should have a linear correlation of more than 0,95
(the detailed data is presented in Appendix).

The total saponin analysis on the extract of Sapindus rarak obtained at different
extraction conditions were dilute to suit the spectrophotometer apparatus. Then the
absorbance value (A) is measured at the appropriate wavelength. Thus the sample
concentration in a solution is calculated with the equation obtain from the
calibration curve of pure saponin standard, and it is a pure saponin equivalent
(Sarvin et al., 2018).

b. FT-IR Analysis

In this study, the chemical structure of saponin extract from different extraction
methods are analyzed by the Fourier transform infrared spectroscopy (FT-IR). The
analysis is performed using Shimadzu FT-IR spectrometer by an attenuated total
reflection method. Before the analysis, 20 mL of sample are evaporate using IKA
RV 10 Rotary Vacuum Evaporator to reduce the water content. A semi liquid
sample of saponin extract is used as the sample. The analysis is also conducted for
the pure commercialized saponin as a reference.

37
3.6.4. Surfactant Characteristic of Saponin Extract from Sapindus rarak

The saponin extract from Sapindus rarak is analyzed for its surfactant characteristic.
Two kind of analysis is performed, foaming properties analysis and dye
solubilization.

a. Foaming properties analysis

Foaming properties were measured according to methods proposed by Chen et al.

(2017). The aqueous foams were produced by hand-shaking the water dispersion of
saponin solution in an 50 mL glass vessel (inner diameter = 3.1 cm and height = 9.8
cm). The foam formation and stability were monitored by a combination of optical
(camera photos) measurements. The foam stability was determined by comparing
the foam volume after 6 hours with the initial foam volume (Behera et al., 2014).
All experiments were performed at 25 °C and at various saponin concentration. The
foaming properties of the extract saponin was also compared to the pure saponin.

b. CMC measurement of saponin from Sapindus rarak DC

The analysis of CMC is performed based on the ring tension methods. Surface
tension measurements of crude saponins extracts at various concentrations were
used to determine the CMC at (25.0 ±0.5) °C in water (Tippei et al., 2012). CMC
is defined as the concentration at which surface tension is the lowest and at which
micelles begin to form. The critical micellar concentration of aqueous Sapindus
rarak extract, was determined by measuring the surface tension with respect to
saponin concentration in the solution. The surfactant concentration at which surface
tension reached its almost constant value considered as the CMC of saponin.

c. Measurement of Hydrophile - Lipophile Balance (HLB) number

The empirical hydrophile–lipophile balance is commonly used parameter to express

the relative degrees of hydrophilic and lipophilic character possessed by
respectively hydrophilic and lipophilic parts of a surfactant molecule. Low HLB
surfactant indicating a hydrophobic surfactant type which soluble in oil phase. In
contrast, high HLB surfactant indicating a hydrophilic characteristic, and is water
soluble. In this study, the HLB numbers obtained by the acid number and
saponification number (Brahmana et al., 1998), based on equation 3.1,

38
 𝑆
𝐻𝐿𝐵 = 20 𝑥 (1 − 𝐴) (3.1)

Where, S is the saponification number and A is the acid number. The measurement
of HLB number is conducted for the saponin extract solution and pure saponin
solution at concentration of 1 CMC.

3.6.5. Solubilization of dye in saponin based surfactant

Micellar solubilization of dyes

The solubilization capability of saponin based surfactant was studied based on the
methods proposed by Samal, et al. (2017) at various concentration of saponin for
reactive synthetics dye. Remazol black, and remazol brown is use as the reactive
dye in this experiment. The saponin extract is prepared following the best extraction
condition with higher saponin concentration. The amount of saponin used as the
solubilized agent is calculate based on the CMC concentration of pure saponin and
saponin extract.

The saponin extract solution is diluted with distilled water and prepared by a
suitable range of saponin concentration (0.2 – 4 times CMC). As much of 10.000
ppm of reactive dyes are separately added to each saponin extract solution. The
suspension were kept in the orbital incubator shaker at 200 rpm agitation speed for
24 hours at room temperature (± 30 oC). After then, the solutions were centrifuged
to separate undissolved dyes. The supernatant was analyzed for its dye and saponin
concentration. The solutions were diluted in distilled water to suit the
spectrophotometer specification. The concentration of dyes in all samples were
measured at respective maximum wavelength using UV-Vis spectroscopy. The
UV-Vis spectrophotometric analysis is conduct based on calibration curve methods.

Measurement of solubilization power (SP) and micellar solubilisation

parameter

The solubilization capacity of dyes in saponin micelles was evaluated by

determining the molar solubilization power (SP). SP also known as molar

39
solubilization ratio (MSR). It is the ratio of moles of solubilized dye to moles of
micellized surfactant in solution. The solubilization power (SP) of saponin
surfactant micelles can be determined from the slope of linear fitting of solubilized
dye concentration vs saponin concentration in solution (Samal et al., 2017 (b), Luo
et al., 2010). The solubilization power of surfactant micelle can also be expressed
in terms of mass (g) of solubilized dye per mass (g) of surfactant. In this study, a
plot of solubilized dye in respect to various saponin concentration is made for
Remazol red and blue.
Another parameter, partition coefficient, Km also represents the effectiveness of
solubilization for any surfactant, which is the ratio of the mole fraction of solute in
surfactant micelles to the mole fraction of solute in the aqueous phase. The partition
coefficient (Km) can be calculated using Equation (3.2) (Samal et al., 2017):
𝑆𝑃
𝐾𝑚 = [𝑆𝐶𝑀𝐶 ] 𝑥 𝑉𝑤 𝑥 (1+𝑆𝑃)
(3.2)

Where Vw is the molar volume of water (1.805 x 10-2 L/mol at 25 oC), SCMC is the
solubilized dye at CMC. The knowledge of distribution coefficient is helpful in
determining the thermodynamic parameter Gibbs free energy (ΔG), which helps in
the better understanding of the mechanisms associated with the solubilization
process (Samal et al., 2017). The Gibbs free energy of solubilization can be
calculated from Equation (3.3).

ΔG = −R. T. Ln K m (3.3)

Where, R is the gas constant in kJ/mol.K and T is the temperature in K.

3.6.6. Application of Saponin for Micellar-Enhanced Ultrafiltration of

Reactive Dye

The extract of Sapindus rarak DC with highest concentration of saponin is chosen

to use in the MEUF study. The extract solution is used as the substitution of
synthetic surfactant. Ready to use extract is the solution obtain after post treatment
process (see 3.3.5). The feed solution is a model of dye wastewater containing
reactive dye

40
Preparation of dye wastewater model solution

The feed in the MEUF process is dye wastewater model solution prepared from
analytical grade remazol dye and distilled water. Remazol blue TQ and remazol
Red Rb are used as the reactive dye on the wastewater model solution. The remazol
dyes is added into an appropriate amount of distilled water with concentration of
300 ppm. The extract of Sapindus rarak is use to replace the surfactant, it is added
to the reactive dye wastewater by a suitable range of 0 – 2 times CMC. Then the
solution was agitated using a magnetic stirrer at 200 rpm for 30 minutes to provide
efficient mixing and kept at room temperature of 26 ± 1 oC to reach the equilibrium
(Melo et al., 2017). Then, the prepared feed solutions were used for MEUF
experiments.

Ultrafiltration and MEUF system

In this study, filtration is carried out using cross-flow filtration cell, equipped with
water pump and pressure control. A schematic flow diagram of the ultrafiltration
set-up is shown in Figure 3.4

The membrane used in this experiment is flat sheet polyethersulfone (PES)

membrane with molecular weight cut off 10 kDa (Sterlich membrane inc, USA).
The membrane utilized in this work has an effective surface area of 9,6 cm2. In each
experiment, the membrane is first placed in distilled water for 2 h before used. It is
then compacted by place it in the cell and compress up to 3 bar for approximately
one hour using distilled water (Ahmad et al., 2006). To discover the membrane
permeability, pure water fluxes were measured for each run. The weight of
permeate collected at specific time was calculated to obtain the initial membrane
characteristic as pure water flux (J0). Then, the dye wastewater model solution was
feed into the filtration instrument. To obtain a clear flux profile, the permeate fluxes
(J) were determined by weighing permeate collected every 5 minutes for 120
minutes (Huang et al., 2014). The total operation time determined in assumption
that the flux steady state condition achieved after 120 minutes. The flux was
calculated based on Equation (3.4) (Bird et al., 1960).
𝑉
𝐽 = (3.4)
𝐴𝑥𝑡

41
Where V is the permeate volume, A is the membrane area, and t is the time interval.
Ultrafiltration was operated without any addition of surfactant in the feed solution.
On the other hand, the micellar-enhanced ultrafiltration was conducted with the
addition of surfactant (model surfactant solution). The experiment was a total
recycle system where permeate and the retentate were recycled into the feed tank.
In each operation, permeate, and retentate were collected and analyzed at the time
of 0, 30, 60, 90 and 120 minutes. The samples are then combined to be an average
single solution.

Analysis of membrane rejection

Ultrafiltration and MEUF performances to remove dye from the wastewater model
solution were evaluated by dye rejection. The rejection (R) sample was collected
for each sample at time 0, 30, 60, 90, and 120 minutes. The calculation was carried
out according to Equation (3.5).
Cp
%R = 1 − x 100%
Cf
(3.5)

where, CP is permeate concentration and CF is the feed concentration respectively.

The concentration of dye was determined using Spectrophotometric UV-Vis at
maximum wavelength by calibration methods.

Micelle Loading Analysis

Ultrafiltration of micellar solutions could be considered as a research method

helpful in estimating the equilibrium distribution constant (Kd) and micelle loading
(Lm) (Huang et al., 2012; Bielska and Szymanowski, 2006). According to the law
of mass action, one definition of Kd is defined as:
𝐷𝑚
𝐾𝑑 = 𝑆 (3.6)
𝑚 − 𝐷𝑤

Lm is defined as:
𝐷𝑚 − 𝐷𝑤
𝐿𝑚 = (3.7)
𝑆𝑚 − 𝑆𝑤

Where Sm and Dm are the concentrations of surfactant and dye pollutant in retentate,
Sw and Dw are the concentrations of surfactant and dye pollutant in permeate. The
concentration of dye in both retentate and permeate is analyzed using UV-Vis

42
Spectrophotometric by calibration methods. As for saponin content was analyzed
using the same tools and methods on UV wavelength (190-400 nm).

Model of Fouling Mechanism

Hermia et al. (1982) presented a physical model to derive the so-called

"intermediate blocking" law, which had previously been considered empirical. The
model also determined a characteristic form of the blocking filtration laws for
constant-pressure filtration. The result in the form was presented in the form of
Equation 3.8.
𝑑2 𝑡 𝑑𝑡 𝑛
= 𝑘 (𝑑𝑉) (3.8)
𝑑𝑉 2

where V is volume of filtrate collected in time t, and k and n are constants

depending upon the mechanism involved. As analyses of membrane filtration are
normally performed in terms of flux it is noted that Equation 3.8 can be presented
in an alternative form. As dV/dt=AJ thus Equation 3.8 can be rewritten in a
physically more meaningful form as Equation 3.9.
1 𝑑𝐽 1 𝑛
− 𝐴2 .𝐽3 . 𝑑𝑡 = 𝑘 (𝐴𝐽) (3.9)

For complete blocking, it is assumed that each solute molecule arriving at the
membrane surface participates in blocking by means of pore sealing (Iritani, 2013).
Moreover, a molecule never settles over another molecule that has been previously
deposited on the membrane surface. Thus, the fractional reduction in permeate flux
is equal to the fractional reduction of the membrane surface area corresponding to
unblocked pores, but not inside the membrane pores (Hwang and Lin, 2002). This
mechanism implies that for the resolution of Eq. 3.9, n must take a value equal to
2. For standard blocking, this model considers that molecules enter the membrane
pores and deposit over the pore walls due to the irregularity of pore passages,
reducing the membrane pore volume. As a result, the volume of membrane pores
decreases proportionally to the filtered permeate volume (Vincent-Vela et al.,
2009). The decrease in the volume of membrane pores with time is equal to the
decrease in their cross section. In this case, flux decline can be expressed by take a
value of n to 1.5. In comparison to the complete pore blocking model, the
intermediate fouling or intermediate blocking model considers the probability that

43
some molecules may settle over others (Iritani, 2013). The non-blocked membrane
surface diminishes with time, thus the probability of a molecule blocking a
membrane pore reduces continuously with time and the n need to be take a value
by 1. As for cake fouling mechanism, the model considers that solute molecules do
not enter the membrane pores, but they form a gel layer over the membrane surface.
Flux decline is then represented by replacing n value in the Equation 3.9 in to 0.
After taking account of the n value and the condition on each fouling mechanism,
the linearised equation according to Equation (3.9) are given in Table 3.1.

Table 3.1 Linearisation equation of blocking/fouling models based on Hermia’s

model

Model of Blocking Linearize Equation Physical Concept Eq.

Mechanism Number
Complete Blocking 𝑙𝑛 𝐽 = 𝑙𝑛 𝐽0 − 𝐾𝑐 𝑡 Formation of surface (3.10)
deposit
Standard Blocking 1 1 Pore blocking and surface (3.11)
= + 𝐾𝑠 𝑡
𝐽 𝐽0
deposit
Intermediate Blocking 1 1 Pore constriction (3.12)
= + 𝐾𝑖 𝑡
𝐽 𝐽0

Gel/Cake Formattion 1 1 Pore blocking (3.13)

= 2 + 𝐾𝑒𝑓 𝑡
𝐽2 𝐽0

44
 CHAPTER IV
RESULT AND DISCUSSION

This research was designed to obtain a saponin extract from Sapindus rarak
as a natural surfactant. The extract having highest saponin yield was characterized
as a surfactant and then used in the surfactant-integrated membrane process. In this
study, the saponin extract was applied to replace synthetic surfactant in the micellar-
enhanced ultrafiltration (MEUF). To confirm the ability of the surfactant to
solubilize the dye, the analysis of dye solubilization and micelle loading were
conducted.

4.1. The investigation of saponin content extracted from Sapindus rarak DC

To find the saponin content in the Sapindus rarak extract, the extraction
process in various condition was conducted. The extraction process was performed
by two different methods for various extraction condition. The first method was
conventional water maceration extraction (ME) and the green ultrasound-assisted
extraction (UAE) methods. The maceration extraction is a solid–liquid extraction
where the bioactive compound (solute) inside the plant material is extracted by
soaking the plant material in a specific solvent for a period of time (Takeuchi et al.,
2009). The efficacy of maceration process is determined by two main factors,
solubility and effective diffusion. The solubility is governed by basic rule of “like
dissolves like” which indicated that polar compounds dissolve in polar solvents,
and nonpolar compounds dissolve in nonpolar solvents (Reichardt and Welton,
2011). In the other hand, UAE methods utilize the ultrasonic wave to help the
extraction process. The phenomenon of ultrasound in creating cavitation bubbles in
the solvent by acting as a micro jet to denature the plant cell wall when the bubbles
collapse at rare fraction resulted in a greater extraction yield of bioactive
compounds. Few researchers have reviewed the applications of ultrasound-assisted
extraction on bioactive principles from herbs (Vinatoru, 2001), food industry and

45
processing (Vilkhu et al., 2008). Figure 4.1 present the yield of saponin at various
the extraction time for both UAE and ME methods.
20
UAE

Saponin Yield (mg/100mg feed)

ME
18

16

14

12

10
0 20 40 60 80 100 120 140

Extraction Time (minutes)

Figure 4.1 Effects of extraction time on the saponin content of the Sapindus rarak
extracts, at temperature of 30oC and solid-liquid ratio of 2 mL/gram

The UAE reaches the maximum saponin content at the time of 40 minutes at
total saponin content of 18.708 mg/100 mg feed. On the other hand, saponin content
of 15.186 mg/100 mg feed was obtained for ME at the extraction time of 120
minutes, which is lower than those of the UAE. According to Figure 4.1, the effect
of extraction time was different on the UAE and ME. On both process, it can be
seen that the saponin yield was increased by the extraction times until a specific
time. And then the yields were constant or slightly decreased by the extension of
processing time. The results indicates that with or without ultrasound irradiation,
the diffusion of the bioactive compounds from material to solvent tend to increase
by the time and the equilibrium for dissolution is established after a certain time.
The increase of saponin yield by the time extension indicates that the solubilization
of saponin compound to the solvent requires a certain time. The extraction with
assistance of ultrasonic waves reach its highest yield at a shorter time than the
maceration. The assistance of ultrasonic waves helps to release the saponin
compound into the solvent by destructs the plant tissues. This process allowing
more saponin to treanfer into the solvent in shorter time.

46
 However, it is possible that the components degrade after a long exposure of
ultrasonic waves. The decline of saponin content at the end of the UAE indicates
that an excess extraction time is not preferable. Ultrasonic wave has an ability to
disturb a plant material. At the beginning of the process this disruption is desired.
However, a longer exposure to ultrasonic wave can also disturb the desired
bioactive compound, saponin, which lowering the saponin yield. The similar results
were also reported on the previous study about UAE of anthocyanin and phenolic
from purple sweet potato (Zu et al., 2016), extraction of natural antioxidant from
Jatropha integerrima (Xu, et al., 2015) and Boldo leaves (Petigny et al., 2013).
Tarade et al. (2006) reports the degradation phenomena of saponin from soybean
flour in the food processing by slow cooking, and long heating. As much of 81-84%
of saponins were reduced in the soaked, dehulled and autoclaves sample of faba
beans (Sharma and Sehgal, 1992). The degradation of saponin also found in the
extraction of chickpeasaponin by microwave assisted extraction (Cheng et al.,
2017). The study by Cheng et al. (2017) also reports that the reduction of saponin
contents depends on the processing parameters such as solid/liquid ratio, extraction
temperature, microwave irradiation power, irradiation time, and pH.
30
Saponin Yield (mg/100 mg feed)

28
26
24
22
20
18
16
14
UAE
12 ME
10
0 20 40 60 80 100

Ratio of Solvent to Feed (mL/gr)

Figure 4.2 The effect of solvent to material ratio on the saponin content of the Sapindus
rarak extracts at 30oC for 120 minutes

Figure 4.2 presents the effect of various ratio of solvent to the dried-ground
Sapindus rarak to the yield of saponin extracted. The yield of saponin was

47
calculated on dry based. The saponin content (ppm) found from the spectroscopy
analysis was recalculated to obtain the content in milligrams of saponin per mL of
total solvent. The total extracted saponin in a batch of solvent (milligrams) was then
divided by the amount of total dry feed in the same batch of solvent (milligrams).
The study was conducted for both UAE and ME process. From the previous
investigation, since the maximum yield was obtained after 40 minutes of extraction,
then the extraction were conducted for 40 minutes, at the extraction temperature of
30 oC. Based on Figure 4.2, it can beSSS seen that the saponin content increase by
the addition of solvent and then slightly decrease or stay stable with a further
increase of solvent. The maximum saponin yields obtained from the UAE process
was 27.871 mg/100 mg feed at the addition of 10 mL of solvent per gram of solid
component. While in the ME process, the saponin content remain constant at the
addition of 50 mL of solvent per gram solid, with the maximum saponin yield
achieve was only 20.586 mg/100 mg feed.

The extraction process of saponin from Sapindus rarak was conducted based
on the solubility of saponin on to the solvent. Saponin is an amphiphilic molecule
having ability to solubilize well on both water or oil/fat soluble solvent. However,
there is a possibility of fatty component to be solubilize together with the saponin
if a fat soluble solvent is used (Hierro et al., 2018). The existence of fat in the
solution is going to disturb the saponin performance as a surfactant (Reichert et al.,
2017; Li et al., 2013). The separation of fat from the crude saponin solution difficult
and complicated, therefore, water was chosen as the solvent to extracted saponin.

With the used of water as the solvent, the saponin will be solubilized with the
addition of solvent. By increasing the volume of solvent, the dissolved materials is
higher and resulted in more extraction yield. In addition, the mass transfer
parameter is affected by the amount of solvent volume. More solvent results on
enlargement of mass transfer parameter and accelerate the diffusion of molecule
into the medium (solvent) (Xu et al., 2016). The solvent also have an effect to
increase the diffusion of the component to the solvent, resulting to the dependence
of extraction rate on the particle concentration gradient. Furthermore, the extraction
is influenced by how fast the component is dissolved and the equilibrium in the

48
liquid achieved (Cacace, 2003). However, at a point, the further addition of solvent
gives no difference due to the saturation of solvent by the extracted compound.
After saturation point, the solvent cannot contain any further component. The
extraction of other natural compound such as anthocyanin and phenolic components
of black currants also shows a similar phenomenon. The amount of extracted
anthocyanin increased as the solute-solvent ratio increases until specific points
(Cacace, 2002). Similar results were also found in anthocyanin extraction by UAE
from Mulberry (Zou et al., 2011), extraction of antioxidant from the flower of
Jatropha integerrima (Xu et al., 2016), extraction of phenolics from wine lees (Tao
et al., 2014) as well as ursolic acid extraction from Ocimum sanctum leaves (Vetal
et al., 2012).
30
UAE
Saponin yield (mg/100 mg feed)

25 ME

20

15

10

0
30 40 50 60
o
Extraction Temperature ( C)

Figure 4.3 The effect of extraction temperature on the saponin content of the Sapindus
rarak extracts at solvent to material ratio of 10 mL/g, for 40 min

The effect of temperature on to the saponin content in the extract solution is

presented in Figure 4.3. The Figure indicates that increase of temperature condition
has different effect for different extraction methods. The extraction by maceration
process shows a positive dependency of extracted saponin with the increase of
temperature. The saponin concentrations increase as the temperature rises. The
highest saponin yield of 19.888 mg/100mg feed was obtained at the extraction
temperature of 50 oC. And the increase of temperature to 60 oC reduced the saponin
yield to 18.842 mg/100mg feed, meaning that in 10 mL of solvent, a gram of
Sapindus rarak powder can produce 188.42 mg of saponin. Considering that the

49
maximum solubility of saponin in water is 50 mg/mL (Sigma-Aldrich, Singapore),
the maximum amount of soluble saponin in the 10 mL of water solvent is 500 mg.
Shows that the extraction of saponin in 10 mL of water was considered possible and
the reduction of saponin yield was not caused by the solvent saturation. A slight
decrease of saponin yield by the addition of heat was due to the heat degradation of
saponin. Bioactive compound in liquid extract form was known to be more
susceptible to heat (Peron et al., 2017). Bioactive compound such as anthocyanin
shows a degradation at the temperature of 50-60 0C (Aurelio et al., 2008).

While, the extraction process with the assistance of ultrasonic wave shows a
different phenomena the rise of temperature decrease the saponin content. The
highest saponin yield of 27.871 mg/100mg feed was obtained at the temperature of
30 oC. The addition of heat decreased the yield to 22.551 mg/100mg feed at
extraction temperature of 60oC. A significant decline of saponin yield was found in
the extraction with assistance of ultrasonic waves. This phenomena shows that a
long exposure of ultrasonic waves on higher temperature condition was able to
gaves destructive effect to the saponin compound, in which, decrease the saponin
yield significantly. In the UAE process, the exposure of ultrasonic waves producing
a lot of cavitating bubbles which creates local increase of temperature in the feed
surface called hotspot. The hotspot induced a destructive effect which helps the
extraction process under right condition (Alupului et al., 2009).

Comparison of UAE and ME reveals that the UAE gives a better result. Figure
4.1 shows that UAE process was faster and gaves higher saponin content compared
to ME. The maximum yield of the saponin obtained from the ME process not even
surpassed the saponin yield obtained from the first 5 minutes of UAE. Figure 4.2
shows that UAE requires less solvent to extract saponin compares to ME. By using
ultrasonic wave in the extraction process, about 80% of required solvent can be
reduced. While, Figure 4.3 showing that UAE need lower heat to extract saponin
compared to ME. Even the addition of temperature can increase the saponin content
in the ME process, the maximum yield cannot compared the yield achieved by UAE
process. The UAE process achieved its maximum yield on 30oC, while the ME

50
achieved its maximum yield on 50oC. The extraction mechanism of saponin through
the ME and UAE was illustrated in Figure 4.4.

(a)

(b) (c)

Figure 4.4 Schematic illustration of (a) Saponin in plant tissue, (b) ME

mechanism and (c) UAE mechanism

Considering the research result thoroughly, the saponin for the further used
in this study will be extracted using UAE process on its optimum condition where
the highest saponin yield obtained. The detail of maximum condition of each
process (UAE and ME) was presented in the Table 4.1.
Table 4.1 The optimized parameter of the saponin extraction by UAE and ME

Ratio of Maximum saponin

Extraction Time solvent to solid Temperature yield (mg/100mg
Methods (minutes) (ml/gr) (oC) feed)
UAE 40 10 30 27.87125
ME 120 50 50 23.7365

51
4.2. Characterization of saponin as surfactant

The extract with highest saponin yields used for the further analysis. The
concentration of saponin in the extract was also known, and used as the calculation
basis (CMC, molar). The saponin characterization was conducted by the
investigation of functional group using FTIR, the analysis of surfactant CMC,
foaming properties analysis and the value of hydrophilic and lipophilic balance
(HLB). All of the investigation was conducted with comparison to the
commercialized pure saponin.

4.2.1 Specific Functional Group

Figure 4.5 shows the FTIR spectra of extract saponin from Sapindus rarak
DC and commercialized pure saponin. In general the peak of sapoin extract was
similar with the peak of the pure saponin. Almost all of the relative peaks shows on
the pure saponin FTIR was also showed in the saponin extract. This confirms that
the extract of Sapindus rarak contain saponin. Both samples shows the infrared
absorbance characteristic of the hydroxyl group (OH) at 3393.90 cm-1 in the pure
saponin and at 3421.77 cm-1 in the saponin extract. Carbon – hydrogen (C-H)
absorption at 2932.16 in the pure saponin and at 2934.24 cm-1 in the saponin extract.
The C=C absorbance was observed at 1609.73 cm-1 and 1638.54 cm-1 in the pure
saponin and saponin extract respectively. The peak of oligosaccharide linkage
which indicating the existence of sapogenins (aglycone), C-O-C, were also present
at 1077.05 cm-1 in the pure saponin and at 1047.13 cm-1 in the saponin extract. The
peak at 1412.58 cm-1 in pure saponin and 1388.07 cm-1 in saponin exract indicating
the group of –CHO from the oleanane structure. A weak peak at 1243.9 cm-1 in pure
saponin and 1261.12 cm-1 in saponin extract indicating the aromatic =C-H and a
broad peak at 617.7 cm-1 and 656.48 cm-1 in pure saponin and saponin extract
presents the pyranose sugar at the glycone structure. The presence of both water
soluble glycone structure and the hydrophobic aglycone structure in the FTIR
spectrum shows that the sample has the saponin characteristic. The IR spectrum of
extract Sapindus rarak also shows a high similarity to the pure commercial saponin,
proving that the extracts contains saponin.

52
Figure 4.5 The FTIR spectra of pure saponin and extract of saponin from Sapindus rarak
DC.

The aforementioned characterization of saponin utilize the functional group

infrared absorptions has been conducted for soapnuts, quilaja saponaria (Almutairi
and Ali, 2014), basellasaponin (Toshiyuki et al., 2001) and steroidal saponin (Da
silva et al., 2002). The previous study reports that the existence of –OH, C-H, C=C,
and C-O-C were identified as oleanane triterpenoid saponins (Kareru et al., 2008).
The study also states that the C=O groups can be either exist or not. The present of
C=O groups indicates a bidesmosidic saponin (Kirmizigul et al., 2002). However,
in Figure 4.5 there is no infrared absorption to confirm the present of C=O groups.
Hence, the saponin was presumably a monodesmosidic oleanane triterpenoid
saponins. The peak of C=O group also cannot be found for the pure commercial
saponin. Indicating that the pure commercial saponin used in this study has a good
compatibility with the saponin from Sapindus rarak. The molecular structure of
saponin from sapindus rarak has also been provided by Morikawa et al. (2009),
which is presented in Figure 4.6. The observation of saponin molecule from various
plant has also been conduted by Kareru et al. (2008) and showing a similar results.

53
 Figure 4.6 Molecular structure of Sapindus rarak (monodesmosidic oleanane
triterpenoid saponin)

4.2.2 Critical Micelle Concentration

The saponin characterization as a surfactant was also conducted to find the

critical micelle concentration (CMC). A knowledge of the critical micelle
concentration (CMC) of the extract of saponin from the plant material is very
important in the context of its applications. The data was required in the surfactant-
integrated process such as foaming, emulsion stabilisation, as well as the
incorporation of lipophilic compounds into micelles, as in micellar-enhanced
ultrafiltration (MEUF). At the air-water or oil-water interface the hydrophilic side
chains of saponins are directed towards the aqueous phase. The hydrophobic
aglycone faces towards the nonpolar air or oil phase. This results in a decrease of
the water’s surface tension or the interfacial tension between water and oil,
respectively, which proceeds until the CMC is reached. At the CMC the interface
is saturated with surfactant molecules. Further increase of the saponin content leads
to self-aggregation of the surfactant molecules in the aqueous phase, resulting in
the formation of micelles. Thus, no further decrease in surface (or interfacial)

54
tension can be observed. The main reason for micelle formation is the attainment
of a minimum free energy state. The illustration of micelles formation in water
solution is presented in Figure 4.7

Figure 4.7 The micelle formation in water solution, (A) Below CMC, (B) Right at
CMC, and (C) Above CMC

The CMC was determined by the surface tension decline for both of pure
saponin and extract of saponin from Sapindus rarak. The surface tension of water
at various concentration of saponin is present in Figure 4.8.
75
Saponin Extract
Surface Tension (dyne.cm-1)

70 Pure Saponin
Surface Tension (dyne.cm-1)

70

65
60 60

55
CMC (%)
50 CMC (%)
50

45
40 40

35
0 1 2 3 4 5 6 7 8 9 10 0.00 0.05 0.10 0.15 0.20 0.25 0.30

Concentration of Saponin Extract (%) Concentration of Pure Saponin (%)

(a) (b)
Figure 4.8 Surface tension of (a) Saponin extract in aqueous solution, (b) Pure
saponin in aqueous solution, at various concentration

To determine the CMC of the saponin extract, the measurement of surface or

interfacial tension of aqueous saponin solutions in various concentrations was
conducted. The surface tension was declined with the addition of saponin for both

55
extract saponin and pure saponin solution as clearly presented in Figure 4.8. At a
certain point the surface tension stops decreasing with a further addition of saponin
and remain almost constant. The surfactant concentration at which the surface
tension reached its constant value considered as the CMC of saponin. The constant
value was taken at the point where three or more point after them is remain the
same. The critical micellar concentration was found to be 0.07 wt % saponin for
pure saponin and 7 wt % of saponin extract for aqueous saponin extract solution.
The CMC of pure saponin is corresponds with the CMC provides by the
manufacturer, which is 0.001-0.1% wt.

There is a huge difference between the CMC of pure saponin and saponin
extract. The value of 7% of weight was calculated based on the addition of aqueous
S. rarak extract to the solution. However, if the calculation was conducted based on
the saponin content in the solution, the CMC was found to be 0.19% wt. The value
was higher than the calculated CMC of the pure saponin. The aqueous rarak extract
was not purified completely like the commercial one. Even after simple purification
using centrifugation, a little amount of impurities such as starch was supposed to
remain in the solution. The starch in the solution can disturb the molecules of
surfactant to assembly in the solution surface. The solution of saponin extracts were
also thicker than the pure saponin solution, indicating a more viscous solution. In
viscous solution, the surface tension was tend to be a little bit higher. The addition
of surfactant to lowering the surface tension was then interrupted by this properties.
Resulting to a higher CMC of the extract solution compared to the pure saponin.
The similar results also found in the previous study on the surfactant production
from the pericarps of Sapindus muccorosi, where the CMC of the crude solution
was higher than the standard saponin (Samal et al., 2017). If the CMC expresses in
millimolar (mM), the CMC of saponin extract and the pure saponin are 3.075 mM
and 1.102 mM respectively.

4.2.3 Foam Ability of Saponin

Liquid films are the basic structural elements of any foaming system containing
surfactant micelles. Consequently, the stability of the foam is highly dependent on

56
stability of the intervening film between the bubbles (Lee et al., 2014; Wasan and
Nikolov, 2008). The amount of foam collapsed (or retained) during a certain
observation period is a measure of its stability. The foam ability is presented as the
initial total foam volume (V0) and the foam stability is the volume of foam after 6
hours (V6) (Behera et al., 2014). Figure 4.9 shows the foam properties of Sapindus
rarak saponin and pure saponin at various saponin concentration. The concentration
of saponin were taken based on the CMC (0.5; 1; 1.5; and 2 times of CMC) which
was under, at, and above CMC value. Please note that the critical micelle
concentration of saponin extract and pure saponin is 3.075 mM and 1.102 mM
respectively.

Total foam after 6 hours (mL)

100 100 100 100
Total foam after 6 h (mL)

(a) (b)

Total initial foam (mL)

Total initial foam (mL)

80 80 80 80

60 60 60 60

40 40 40 40

20 20 20 20

0 0 0 0
0.0 0.5 1.0 1.5 2.0 0.0 0.5 1.0 1.5 2.0
Amount of saponin extract ( -times of CMC) Amount of pure saponin ( -times of CMC)
Foam after 6 hours Foam after 6 hours
Initial Foam Initial Foam

Figure 4.9. The value of total initial foam (V0) and total foam after 6 hours (V6) of
(a) saponin extract and (b) pure saponin

Figure 4.9 shows that the addition of saponin, increased the initial foam
formation. This is expected because the increase in surfactant concentration
increased the concentration of surfactant molecules at the air−water interface,
which stabilized the foam film. The addition of saponin at its CMC and above its
CMC gives more prominent foam formation. The similar phenomena was found for
the foaming of ionic and nonionic surfactant (Behera et al., 2014).
The volume of foam slowly decreased with time. The total foam after 6 hours
showing the foam stability. The solution containing more saponin gives a better
foam stability for both pure saponin and extract saponin. A high concentration of
the surfactant in the bulk solution may increase the rate of transport of the surfactant

57
molecules toward the interface (Chang et al., 1995). The similar results of foaming
stability by the addition of anionic synthetic surfactant (Lukenheimer and Malysa,
2003). Figure 4.9 also presents that he addition of saponin above the CMC also
grant a better foam stability. The previous study initiate that the foam surface
elasticity was increased by the formation of surfactant micelle (Belhaij and Al-
Mahdy, 2015). High foam stability at surfactant concentrations above the CMC was
also reported elsewhere for various surfactant systems (Lukenheimer and Malysa,
2003; Folmer and Kronberg, 2000).

4.2.4 Hydrophile-Lipophile Balance

The investigation of the surfactant properties was also conducted to determine

the hydrophilic and lipophilic balance of the saponin solution. Empirical
hydrophile–lipophile balance, which is commonly used to express the relative
degrees of hydrophilic and lipophilic character possessed by respectively
hydrophilic and lipophilic parts of a surfactant molecule (Verdinelli et al., 2008).
HLB was determined empirically with the range of 0 to 20. A hydrophilic properties
were found for the higher HLB, and in contrast the hydrophobic properties found
for the lower HLB. The HLB values was only counted for nonionic surfactant such
as saponin. The common value of HLB and its application was presented in the
Table 4.2.
Table 4.2 Common HLB value and its application

Range of HLB Common Application

2-6 Water in oil emulsifier

7-9 Wetting agent

8-18 Oil in water emulsifier

10-15 Detergent

15-19 Solubilize

In this study, the HLB value was calculated for the saponin extract and the
pure saponin. It was calculated based on the acid value and saponification value

58
(Gaud and Gupta, 2009). From the calculated HLB value, the results shows that the
saponin extract from Sapindus rarak and pure saponin was confirmed to be a
hydrophilic surfactant. Both of the saponin (pure and extract) having a HLB number
between 15 to 19. This kind of surfactant was often used for solubilization.
Generaly, high solubility surfactant is often use in cleansing and detergency (Croda,
2016). As mention before, saponin has a high value of HLB showing that saponin
is rather to have hydrophilic characteristic. This characters match the solution
system where the dye wastes are water soluble and the solvent is water. The detail
of the HLB value and the other parameter of saponin characterization was
summarized on the Table 4.3.
Table 4.3 The surfactant characteristic and properties of saponin extract from Sapindus
rarak and commercialized pure saponin

Characteristic Extract Saponin Pure Saponin

Monodesmosidic Monodesmosidic
Saponin Types oleanane triterpenoid oleanane triterpenoid
saponins saponins
7% w/w of extract 0.07% w/w of saponin
CMC (in aqueous
saponin / 0.2% w/w of content
solution)
saponin content
Lowest Surface 39.9 dyne.cm-1 41.4 dyne.cm-1
Tension
HLB Value 16.8 18.7

4.3. Dye Solubilization

Solubilization is a key function of the micelle. It is the favorable transfer of a

solute from an aqueous region to a micelle inside, and results of intermolecular
interactions contrast among the surfactant, water, and the solute to be solubilized
(Date et al., 2016). In MEUF process, the surfactant containing hydrophobic and
hydrophilic groups is added into the solution containing organic matters or heavy
metal ions. The surfactant aggregates to form large amphiphilic micelles that bind

59
the solutes. The micelle is made up of the inner core (constituted by hydrophobic
groups), palisade layer (constituted by CH2 groups) and outer layer (constituted by
hydrophilic groups). The illustration of surfactant micelles part is presented in
Figure 4.10. The organic matters get embedded in the micelles via ‘‘like dissolves
like’’ principle while heavy metal ions are adsorbed on the opposite-charged
micelles via electrostatic interaction (Purkait et al., 2005 (a); Huang et al., 2009).
The micelles along with the solutes are rejected into the retentate stream by
ultrafiltration membrane. Due to different solubilization principles, organic and
inorganic contaminants could be retained simultaneously by MEUF (Lee et al.,
2005; Purkait et al., 2005 (b)). The solubilization system of organic matters or heavy
metal ions with surfactant is a dynamic balance system, namely, the location of
solutes in micelles changes with time. It spends only 106–1010s on solutes within
micelles (Zhao, 2006). Hence the study surfactant ability to solubilized pollutant
molecule was important.

Figure 4.10 Part of surfactant micelle in water

If the solubilization effect is seen as the distribution of solutes between

micelles and water, the partition constant (Km) and solubilizing power (SP) could
be estimated (Bielska et al., 2006; Samal et al., 2017 (b); Tehrani-Bagha et al.,
2013). The molar solubilization capacity or solubilization power (SP) of a
surfactant is defined as moles of solubilized dye per mole of micellized surfactant
(Tehrani-Bagha et al., 2013). The solubilization power of a specific surfactant can
thus be determined from the slope of its solubility curve at various surfactant

60
concentration. If the molecular weights of the surfactant and the solubilisate are
known, the SP can also be expressed as the weight of the compound solubilized per
unit mass of surfactant.
120
(A)
Solubilized Dyes (mM) 100

80

60

40

20 R.Red; Sapindus rarak

R.Red; Pure Saponin
0
0 2 4 6 8 10 12 14
Surfactant Concentration (mM)

80 (B)
Solubilized Dyes (mM)

60

40

20
R.Blue; Sapindus rarak
R.Blue; Pure Saponin
0
0 2 4 6 8 10 12 14
Surfactant Concentration (mM)
Figure 4.11 Effect of saponin concentration (in mM) on the solubility of (A) Remazol red
RB, and (B) Remazol Blue TQ

The solubilization capability of saponin micelle was studied at various

concentration of saponin for two different remazol dyes, red RB and blue TQ. The
saponin concentration was applied for below and above the CMC. From the
previous analysis, the critical micelle concentration of saponin extract and pure
saponin is known to be 3.075 mM and 1.102 mM respectively. The concentration
of saponin was kept under the maximum solubility of saponin in water which is 50
mg/mL (78.76 mM) (Sigma-Aldrich, Singapore).

61
 The amount of solubilized dye in mM with respect to mM of saponin in
solution is presented in Figure 4.11. It is shows that the application of surfactant in
aqueous system enhanced the solubility of dyes. It also shows a sharp increase of
solubilized dyes after the CMC and above the CMC of surfactant for both dyes.
However, the concentration of saponin below the CMC showing a very low dye
solubilization capability. This trend confirms that surfactant micelles were
responsible for the enhancement of dye solubilization. Simultaneously, the
agglomerates of micelles may have adsorbed some dye molecules which resulted
into higher solubilization. This was due to the incorporation of dyes into the
micelles of surfactant. At the concentration below CMC, some dyes can be
solubilize in the surfactant, because micelles can also form in low concentration
(Tehrani-Bagha et al., 2013). However, the location of solubilization of dyes in
nonionic surfactant micelle is not well understood. In many studies, it was observed
that the solubilization of dyes occurs within the inner core of surfactant micelle
(Purkait et al., 2004 (c)).

The increase of solubilization of remazol red and blue in saponin micelle were
due to the resultant of two phenomena, first, the micellar solubilization of dyes by
saponin micelles, and the sorption of dye molecules by agglomerates of saponin
micelles (Samal et al., 2017). In general the inorganic compound is trapped in the
hydrophobic core of the surfactant micelle, and the ionic component is adsorbed on
the outer surface of the surfactant micelle (Huang et al., 2012). The study conduct
by Date et al. (2016) proposed that it is likely for the dye to accumulate in the
innermost part of the micelles outer region. When the solute is polar, the solvation
free energy was similar between the micellar center and bulk water. Thus, there is
no strong polarity gradient in the shell, making the solute solubilization placement
more variable. Some polar molecule may solubilized near the micelle core and some
other may solubilized in the outer region of the micelle. When nonionic surfactant
such as saponin was used, the outer region of the micelles is considerably more
polar than core region (Tehrani-Bagha et al., 2013). Meaning that, the polar solute
is localized more strongly in the innermost part of the hydrophilic region. The

62
molecules with polar substituents, such as dye, also may incorporated in the outer
region of nonionic surfactant micelles to some extent.

Another study also state that the interface between the hydrophobic and
hydrophilic regions is thus the preferable location of the polar solute (Tehrani-
Bagha et al., 2013). In some case, the polar molecule also solubilized near the
micelle core, because the polarity of the outer region of nonionic surfactant was
weaker than ionic surfactant. The study conduct by Samal et al., (2017) state that
some molecules of the dye may also trapped in the core of the saponin micelle, and
some dye molecules may have adsorbed on the agglomerates surface.

Considering that the molecular structure of the dye have both of polar and
nonpolar region. Further, this micellar solubilization allows the hydrodynamic size
of methylene blue and eosin yellow to increase several fold. Also, the micelle size
of saponin, which is much greater than the other synthetic surfactants micelle,
makes its beneficial in surfactant based process such as MEUF. Figure 4.12 shows
the molecular structure of remazol red, remazol blue and the illustration of dye
solubilization in the nonionic saponin micelles.

(A) (B)

63
 (C)

(D) (E)

Figure 4.12 Solubilization of remazol dye in saponin micelle (a) molecular

structure of remazol blue, (b) molecular structure of remazol red, (c) saponin
micelle, (d) solubilization at innermost part of outer region, (e) solubilization at
outer part of hydrophobic side

The molar solubilization power (SP) is determined by the slope of the linear
regression plot of solubilized dyes in respect to surfactant concentration at
concentration above the CMC. The calculated SP of remazol red and blue were
summarized in Table 4.4. The SP of extract rarak is considered similar with the pure
saponin for both remazol dye. This confirms the performance of saponin from
Sapindus rarak is also able to perform as good as the commercial saponin. Higher
SP of extract saponin compared to pure saponin also shows that the addition of
saponin give more prominent effect for saponin extract. Both of saponin micelle
can accommodate more remazol red molecules as compare to remazol blue. It is
natural considering that remazol red was smaller in molecular structure than
remazol blue.

64
 Another important information is the knowledge of thermodynamic
parameters, such as, a change in Gibbs free energy (ΔG) is important. To understand
the feasibility of such solubilization process. Therefore, the Gibbs free energy of
solubilization (ΔG) was calculated for remazol red RB as well as remazol blue TQ.
The ΔG of Sapindus rarak saponin was found to be -12.283 and -13.388 KJ.mol-1
for remazol red and blue, respectively. As for the pure commercialized saponin the
ΔG were -11.821 and -12.634 kJ.mol-1 for remazol red and blue respectively. The
negative value of Gibbs free energy of solubilization (ΔG) suggests that, the
solubilization of the dyes in saponin micelle was a feasible and spontaneous
process. The ln Km, and Gibbs free energy of solubilization (ΔG) of both the dyes,
are summarized in Table 4.4.

Table 4.4 Molar solubilization power (SP), ln Km and ΔG of solubilization of pure

saponin and extract of S.rarak for remazol dye, at 27oC

Remazol Type of Solubilization power ln Km Gibbs free energy

Dye Surfactant (SP) (mM/mM) (ΔG) (KJ.mol-1)

Extract S.rarak 0.251 4.924 -12.283

Red RB
Pure Saponin 0.199 4.739 -11.821

Extract S.rarak 0.239 5.367 -13.388

Blue TQ
Pure Saponin 0.192 5.065 -12.634

4.4. Permeate flux profile of surfactant-enhanced ultrafiltration system for

dye removal

The next experiment is conduct to observe the effect of saponin addition and
the remazol dye variants the MEUF system performance based on the flux profile.
The ready to used saponin extract obtained from the extraction process was used as
the surfactant in the ultrafiltration process to treat the dye wastewater. The
wastewater was a model made of two kind of reactive dye, Remazol Red RB and
Remazol Blue G. In this study, the effect of saponin concentration on the flux
profile was investigated. The flux profile was compared for filtration and

65
ultrafiltration process with feed solution in various saponin concentration. The flux
profile of each run was presented in the Figure 4.13.

(a) 1.2
R.Red 0 CMC
R.Red 0.5 CMC
1.0 R.Red 1 CMC
R.Red 1.5 CMC
R.Red 2 CMC
Normalized Flux (J/J0)
0.8

0.6

0.4

0.2

0.0
0 20 40 60 80 100 120

Time (minutes)
(b)
R.Blue 0 CMC
0.8 R.Blue 0.5 CMC
R.Blue 1 CMC
R.Blue 1.5 CMC
R.Blue 2 CMC
Normalized Flux (J/J0)

0.6

0.4

0.2

0.0
0 20 40 60 80 100 120

Time (minutes)

Figure 4.13 Variation of the observed permeate flux of remazol red RB and Tq
Blue with time at room temperature and pressure of 1.5 bar

Figure 4.13 shows the observed normalized flux of remazol red RB and Tq
blue compared to time. The permeate flux of all variables were slightly decline with
the processing time and then remained almost constant for the rest of the
experiment. This flux (J) constant value is considered as the steady state permeate
flux. The phenomenon of flux decline is mainly due to the concentration

66
polarization effect and membrane fouling (Huang et al., 2014), although osmotic
pressure and precipitation could also contribute to flux decline (Zaghbani et al.,
2007). Concentration polarization appears as a result of a local increase of solutes
concentration close to the membrane surface. Membrane fouling was mainly due to
gel layer formation and pore blocking, occurs as a result of deposition and
accumulation of solutes on the membrane surface and within the membrane pores
(El-Abbasi et al., 2011). With the addition of saponin, the surfactant micelles
generate a deposited layer over the membrane surface (concentration polarization),
thus increasing the resistance against the solvent flux through the membrane. As a
result, in the initial stage of MEUF experiments, the micelles are accumulated near
the membrane surface, producing a sharp decline of permeate flux due to
concentration polarization and gradual formation of gel layer. As the filtration
proceeds, the gel layer becomes denser, leading to a slow flux decline. In the final
stage, the membrane fouling is fixed and the permeate flux reaches the steady state
value (Huang et al., 2014). Besides, the increment of the osmotic pressure
difference across the membrane (related to micelles concentration in permeate and
retentate streams) reduces the effective TMP and, as a result, the permeate flux
decreases with operation time (Zaghbani et al., 2007). The similar result was also
found in the previous study using ionic and non-ionic surfactant to remove
emerging contaminants (Acero et al., 2017), removal of cadmium ions (Li et al.,
2011) and recovery of phenolic compound (Victor-Ortega et al., 2017). Removal of
various indigosol reactive dye using cationic and anionic surfactant, SDS and CPC,
also shows a similar flux profile with this study (Aryanti et al., 2016).

In absence of saponin, the remazol red solution was showing more rapid flux
decline at the end, while the blue one showing a rapid decline at the beginning of
UF process. This phenomena associated with the molecular weight difference of
the two dyes. The dyes molecular weight were 668.999 g/mol and 1098.062 g/mol,
for remazol red rb and remazol tq blue, respectively. The molecule of remazol tq
blue was bigger in size. It will starts blocked the membrane pores as soon as the
process begin. This direct pore blocking causing the flux decline rapidly in the
beginning of the process. However, the flux was then only slightly declined after

67
40 minutes of UF. This was because of too much deposition of dye molecule in the
surface of the membrane. Where, there is a possibility of the excess dye molecule
to be carried out again by the feed current. Meanwhile, the remazol red rb molecules
were small and way smaller than the membrane pore, which is 10.000 Da. In the
UF process, the red dye molecule will just through the membrane pore easily, which
gives a rather high and stable flux profile. However, the deposit of dye molecule
inside the membrane pore or surface cannot be denied. After 60 minutes of filtration
process, the deposit of dye molecules start to affect the flux profile. The flux profile
decline rapidly in the end of the filtration process because of the continuously
blocked membrane pores.

Figure 4.13 also shows the effect of saponin addition to the flux profile. The
feed without any addition of saponin has the highest flux. In general, the flux value
is decreased with the increase of saponin concentration on the feed solution, both
for above or below its CMC. The addition of saponin at 2 times CMC having a
lowest normalized flux on both remazol red rb and blue tq. This is due to the
interaction of saponin molecule with the pollutant where the pollutant is entrapped
in the micelle saponin structure. An ionic reactive dyes, such as remazol red rb and
remazol blue tq, can be solubilized into the hydrophobic and hydrophilic medias in
micelles, or dissociate to ions which are adsorbed on the surfactant micelles
(Zaghbani et al., 2009; Bielska et al., 2009). Addition of saponin (surfactant)
molecule in the feed solution at a concentration above the CMC generates the
formation of surfactant micelles (Chang et al., 2011). In general, the micelle
structure is the hydrophobic region in the internal core, and hydrophilic region in
the external side. The hydrophobic core had the ability to solubilize hydrophobic or
less polar molecule. As for the polar or charged layer of the external side,
micelle has the more hydrophilic characteristic (Acero et al., 2017). Based on this
fact, the pollutant molecule will be trapped in the innermost part of the micelle outer
region, whit hydrophilic character. This hydrophilic side of surfactant micelle had
the tendency to attached one to each other in low aqueous solution. The micelles
forms a layer and blocking the water to get through the membrane, resulting to the
lower of flux. However, this layers are not permanent, proving by the rather stable

68
flux. The micelle surfactant molecule can goes on to the retentate again because of
the feed flow. On both type of dye, the addition of saponin higher than its CMC
showing a similar result. The solution with addition of saponin at 1.5 and 2 times
of CMC was showing a similar normalized flux. It is indicating that the addition of
excess saponin will not generates more blocking in the membrane. Yet its effect to
the rejection capability was definitely important.

4.5 Rejection of dye and saponin residue

The performance of membrane ultrafiltration is determined by its ability to
retain a particular component. This performance was measured by the value of
rejected compound divided by the initial compound concentration, known as %
rejection (Danis et al., 2009; Melo et al., 2017). It is an important parameter to
present the membrane selectivity which means the ability of the membrane to retain
or let pass a particular species. Membrane selectivity depends on the interfacial
interaction between the membrane surface to the species that pass through it, the
size of the species and the membrane pore size. Substances with molecular weight
bigger than the membrane pore size are retained on the membrane surface, whereas
the smaller-molecular-weighted species will pass through the membrane. In this
experiment, the permeate stream is expected to be water with relatively low
concentration of insolubilized solutes with free surfactants (Purkait et al., 2004).
Hence, the concentration of dye on permeate was analyzed. The saponin
concentration on the permeate solution was also analyzed to investigate the amount
of free saponin monomer pass through the membrane. Table 4.5 shows the
impurities concentration on permeate after filtration. A various concentration of
saponin extract was used (0; 0.5; 1; 1.5; and 2 times CMC), with CMC of Sapindus
rarak saponin extract is known to be 3.075 mM.

Based on table 4.5, on both remazol red and blue, the dye concentration in
permeate of UF system is higher than the MEUF. This corresponds to the
phenomena where, more dye impurities transfer into permeate on the ultrafiltration
system. Whether caused by direct pass through the membrane film or convective
transfer of solute particles. The addition of surfactant into the polluted aqueous

69
wastewater resulting in the lower of impurities concentration on permeate. As the
saponin added at 0.5 time of CMC, the dye concentration slightly decreased from
187.73 to 148.58 ppm and 170.75 to 100.03 ppm. This is showing that the addition
of saponin below the CMC also affect the dye concentration. It was unexpected that
the permeate dye concentrations decreased, since it was often considered that
surfactant monomers did not form micelles while surfactant concentration was
below one CMC value. Due to the concentration polarization effect, the gel layer
was formed at the membrane-bulk solution interface. In the gel layer, the
concentration of saponin might exceed the CMC value, then surfactant micelles
were formed (Misra et al., 2009; Karate and Marathe, 2008). In addition, even if the
surfactant concentration was lower than CMC, the surfactant molecules could form
small pre-micelles, which could result in the weak solubilization effect. At the same
time, high concentration of saponin in permeate was also found. This is showing
that most of the saponin present in the state of free monomer molecule. Considering
its size, the monomer molecule was expected to pass through the membrane pore
easily.

Table 4.5 Concentration of dye and saponin on the permeate after membrane
separation
Dye Concentration on Saponin Concentration on
Surfactant Permeate (ppm) Permeate (ppm)
Concentration Remazol Red Remazol Remazol Red Remazol Blue
RB Blue Tq RB Tq
0 times cmc 187.73 170.75 0 0
0.5 times cmc 148.58 100.03 2246.25 2160.63
1 times cmc 10.34 3.83 471.75 535.25
1.5 times cmc 9.52 2.02 573.38 619
2 times cmc 8.93 1.74 611.89 639

The addition of saponin right at CMC was drastically decreased the dye
concentration, from 148.58 to 10.34 ppm and 100.03 to 3.83 ppm for remazol red
and blue respectively. At CMC, the saponin start to aggregate and form a micelles,

70
resulting to the lowering of dye concentration. Moreover, the saponin concentration
on permeate was also decrease, showing that the saponin start to grow in size and
able to be retained by the membrane. The dye concentration in the permeate keep
decrease by the increase of saponin. As the feed saponin concentration increased
from 1 to 2 times CMC, the permeate dye concentration slowly decrease from 10.34
to 8.93 ppm and 3.83 to 1.74 ppm for remazol red and blue respectively. Obviously,
with the increment of saponin concentration, more micelles would be aggregated
and more dye molecules solubilized in the micelles. The surfactant was added at
concentration higher than CMC, where the surfactant molecule aggregates and
forming micelles. Saponin was acted as a non-ionic surfactant in this process. It
does not has any charge, hence the dye was solubilized in the micelles by the
hydrophobic and hydrophilic relation. The dye was trapped inside the surfactant
micelles and grow bigger than the membrane pore. As a result, the dye pollutant
can be retained by the high MWCO (molecular weight cut off) ultrafiltration
membrane. The previous study has also investigated MEUF process to treat
wastewater using ionic and non-ionic surfactant. The successful results are reported
by the previous study for removal of cadmium ions using SDS and mixed surfactant
(Li et al., 2017), chromium ions using anionic (SDS) and non-ionic (NPE) (Aoudia,
et al., 2003), boron ion (Tortora et al., 2018) and zinc ions (Zhang et al., 2009).

As seen in table 4.5, the remazol red RB permeate have lower concentration
of dye impurities compared with the remazol blue Tq. Based on the molecular
structure of each dye used in this experiment, the remazol blue has a bigger
molecular weight than remazol red. This more prominent structure of remazol blue
allows molecules to retain easily on the membrane than other smaller molecules.

The pollutant concentration on permeate also affects the membrane rejection.

Permeate with lower impurities concentration specify a better membrane rejection.
The membrane rejection of various remazol dye filtration under ultrafiltration and
MEUF system is exhibited in Figure 4.14 which have conformity with the trend of
impurities concentration on permeate. The increase of surfactant concentration will
increased the dye and saponin rejection. The MEUF system showing a superior
result than the UF system with the highest % rejection of dye of 97.02% and 99.42%

71
for remazol red and blue respectively, at the saponin concentration of 2 times CMC.
While, the UF system only able to achieve the % rejection of 37.42% and 42.08%.

(A) (B)

100 R.Red RB 100

R.Blue TQ

Rejection of Saponin (%)

Rejection of Dye (%)

80 80

60 60

40 40

20 20
R.Red RB
R.Blue TQ
0 0
0.0 0.5 1.0 1.5 2.0 0.5 1.0 1.5 2.0
Saponin Concentration (- times CMC) Saponin Concentration (- times CMC)

Figure 4.14 Rejection of remazol dye at various concentration (0; 0.5; 1; 1.5; and
2 times CMC), (A) rejection of dye, (B) rejection of saponin

4.6 Performance of Surfactant in MEUF system: Micelle Loading an

Equilibrium Distribution Constant

The basic idea for MEUF is that a surfactant forms large amphiphilic
aggregate micelles when it is added to aqueous streams at a concentration higher
than its critical micelle concentration (CMC). The solute in the solutions can be
mostly trapped by the micelles if they tend to be strongly attracted by the micelle
surface and will solubilize in the micelle interior, respectively. The solute type can
be ions, insoluble compound or dissolved organic compounds with various polarity.
After trapped in the surfactant micelle interior, the hydrodynamic size of the solutes
increases and the solutes can be retained. Whereas the untrapped species readily
pass through the ultrafiltration membranes (Juang et al., 2003; Huang et al., 2012).
In this study, most of the solute in dye-containing effluent (dye molecule) can be
solubilized in the micelles. Micelles containing solubilized solute are larger in size
which makes it easier to be filtered by an ultrafiltration membrane. For this reason,
the ability of surfactant to solubilize the pollutant molecule was very important.

The performance of surfactant can be predicted by calculating the micelle

loading (Lm) and the equilibrium distribution constant (Kd). Micelle loading is the

72
proportion of dye solubilized in the micelles by the concentration of surfactant that
formed micelles. As for equilibrium distribution constant is a ratio of dye in the
micelle and in the solution. The effect of saponin concentration to the Lm and Kd of
remazol dye is present in Table 4.6. The saponin concentration variation was
calculated based on the CMC of Sapindus rarak saponin, which, one CMC is
determined to be 3.075 mM.

Table 4.6. The effect of saponin concentration to the equilibrium distribution

constant and micelle loading for remazol red and blue

Micelle Loading (Lm) Equilibrium Distribution Constant

Saponin (mM/mM) (Kd) (mM/mM)
Concentration Remazol Red Remazol Blue Remazol Red Remazol Blue
RB TQ RB TQ

0.5 times CMC 0.0017 0.0056 2.079 4.187

1 times CMC 0.0678 0.0547 43.263 134.556
1.5 times CMC 0.0413 0.0474 47.068 256.336
2 times CMC 0.0344 0.0433 50.249 297.960

The system feed is a pseudophase solution which contain dye molecule, free
surfactant molecule, free surfactant micelles, and a surfactant micelles with
entrapped dye molecule. In this solution, micelles are a dynamic aggregates which
equilibrium with the individual surfactant molecules. The individual surfactant
molecule can pass through the pores of a membrane along with the free dye
molecule. Bielska et al. (2006) state that the distribution of the dye in the
pseudophases (micellar and aqueous), in permeate and retentate were in
equilibrium. As a result, the ultrafiltration could be used to estimate the distribution
coefficients of dye molecule in the retentate and permeate. It is defined as the ratio
of dye concentrations in the retentate and permeate. The study has also arrange an
equation presenting the loading of the micelles (Lm), defined by equation 3.7, which
also use in this study.
𝐷𝑚 𝑥 𝐷𝑤
𝐿𝑚 = (3.7)
𝑆𝑚 𝑥 𝑆𝑤

73
 A recent study conduct by Huang et al. (2012) has also shows a straight-line
relationships for this equation with the free term near zero were observed. The study
shows high (>0.9) R2 denotes the determination coefficient that confirms the
statistical validity of the equation.

In this study, a very low Lm for the solution with addition of saponin below
the CMC values, in both of remazol red and blue was found. This was
corresponding to the fact that under the CMC there is not much micelles formation.
Hence, very little amount of dye that solubilize in the saponin micelles. At the CMC
the micelle loading was increased significantly, as much of 0.0961 mM of remazol
red loaded in 1 mM of saponin, and 0.0674 mM of remazol Blue loaded in 1 mM
of saponin. Further addition of saponin into 1.5 and 2 times of CMC leads to
decreasing the Lm into 0.0352 mM/mM and 0.0428 mM/mM for remazol red and
blue respectively. This phenomena was unexpected considering that more
surfactant added into the dye solution, more dye will loaded into the micelle,
increasing the Lm (Tehrani et al., 2012; Tehrani et al., 2013). However, in this study,
the saponin was applied into the MEUF system where other parameter should be
taken into account, such as the maximum solute capacity of the membrane. The
membrane use in this study was a commercial standard with the suggested feed
solute capacity of 180-300 ppm. Thus the dye concentration in the feed was remain
constant at 300 ppm for all variables in this study. There is a possibility that this
much of dye concentration does not need too much saponin to solubilize it. Hence,
the Lm was decreased after the further addition of saponin. It is also confirmed by
the rejection value discussed in the previous section, which showing an
insignificant increased despite of addition of saponin above its CMC.

According to table 4.6 the equilibrium distribution constant was continuously

increased by the addition of surfactant regardless to the concentration below the
CMC. Kd was define as the ratio of dye concentration in the micelle and in the
surrounding water for a particular surfactant concentration (Tehrani-Bagha et al.,
2013). Thus, the increase of Kd was obvious, considering that by the addition of
saponin, the concentration of free dye in the solution was decreased, and more dye
molecule solubilized in the saponin micelles

74
4.7 Model of Fouling Mechanism

Mathematical model can be useful to accurately predict the fouling

phenomena on the membrane filtration process. Blocking mechanism of remazol
dye during ultrafiltration and MEUF was studied by applying Hermia’s
mathematical model. Hermia model provides a comprehensive fouling prediction
models, well equipped with four different fouling mechanisms (Chang et al., 2011).
The schematic of each membrane fouling mechanism was illustrated in Figure 4.15.

Figure 4.15 Schematic illustration of mechanisms of four blocking filtration laws,

adapted for cross-flow filtration. PT is the trans-membrane pressure acting as
driving force (Iritani et al., 2013)

The experimental filtration data is fit to the empirical fouling models by

Hermia to identify well suited fouling mechanisms. Previous study reported a well
fitted result of Hermia’s model with the experimental data for removal of
polysaccharides (Nataraj et al., 2008), organic pollutant (Zhang et al., 2015),
indigosol dye (Aryanti et al., 2018), and remazol dye (Aryanti et al., 2017) from
wastewater using synthetic ionic surfactant. The result of experimental data fitting
to Hermia’s model in UF and MEUF of remazol dye was presented in Table 4.7.

The fitting experimental data and the degree of model fitness (represent by
R2) based on Hermia’s model was presented in Table 4.7. The value of
corresponding correlation (R2) was simply used to determine the fitted blocking
mechanism rationally. The bold value in the table showing the best fitted fouling

75
 mechanism model with the experimental data. Based on the fitted model, the MEUF
of remazol red RB and Blue TQ showing a similar results.

Table 4.7 Mathematical model parameter of UF and MEUF blocking phenomena

on indigo sol dye removal

Complete Intermediate Standard Cake Formation

Remazol CMC Blocking (n=2) Blocking (n=1) Blocking (n=3/2) (n=0)
Dye R2 Kc R2 Ki R2 Ks R2 Kfc
0 (UF) 0.9232 -0.0038 0.9235 0.0001 0.9523 0.0001 0.9214 3.10-7
0.5 0.8783 -0.0027 0.8872 0.0001 0.8878 0.0002 0.9124 1.10-6
Red RB 1 0.8872 -0.0037 0.8551 0.0004 0.8682 0.0004 0.8699 1.10-5
1.5 0.8764 -0.0036 0.871 0.0003 0.8719 0.0003 0.8293 4.10-6
2 0.9009 -0.0113 0.8137 0.0035 0.8879 0.0022 0.6898 0.0007
0 (UF) 0.8691 -0.002 0.8695 9.10-5 0.8965 9.10-5 0.8631 3.10-7
0.5 0.9096 -0.0057 0.9177 0.0003 0.918 0.0003 0.9252 3.10-6
Blue TQ 1 0.8762 -0.0039 0.8494 0.0004 0.8565 0.0004 0.776 2.10-5
1.5 0.9138 -0.0035 0.8997 0.0003 0.9119 0.0003 0.8956 6.10-6
2 0.9746 -0.0032 0.9478 0.0002 0.9479 0.0002 0.9418 4.10-6

The UF process without addition of saponin fitted to the fouling mechanism

of standard blocking, with the R2 of 0.9523 and 0.8965 for remazol red and blue
respectively. The molecular size of remazol red and blue was way smaller than the
membrane MWCO. This fact supporting the phenomena of dye molecule to easily
goes through the membrane pore. Some of the dye molecule may entrapped inside
the membrane pore instead of escaping it. This was resulting to the blocking of
membrane pore which shows a standard blocking mechanism in this study. Figure
4.15 shows the illustration of membrane fouling mechanism. The previous study of
MEUF for synthesis of galacto-oligosaccharides also proposed the fact that a solid
molecule with molecular weight less than the membrane MWCO supposed to
caused standard blocking. The blocking process also follows with a weak ability of
membrane rejection (Cordove et al., 2016), which correspond with this study.

76
 The micellar-enhanced ultrafiltration of all remazol dye with saponin
concentration above it CMC shows a fitting to complete blocking mechanism, with
range of R2 from 0.8762 to 0.9746. Complete blocking is the blocking mechanism
resulting a reduction of open pores without deposition of foulant particles on the
membrane surface. This blocking occurs when the foulant particle size is similar or
bigger than the membrane pore size (Grzegorzek and Majewska-Nowak, 2018;
Sarkar et al., 2009). As explained before, that addition of saponin above the CMC
will generates a molecular grow because of the micelle formation. The dye
molecule will gro in sized and expected to be bigger than the membrane MWCO.
Resulting to a complete blocking of membrane pore. However, this molecule was
supposed to not develop a cake layer and there might be only a "flowing cake" layer
and a very small (or nil) "stagnant cake" layer on the membrane surface (Field et
al., 1995). The big molecule of dye only blocked the pore membrane due to the
pressure caused by the feed flow and not generates a cake layer.

Instead, the formation cake was taken a places when the saponin was added
below the CMC. Indicated by the fitted data to the cake formation fouling
mechanism (R2 of 0,9252). In this condition, the solution consist of free saponin
molecule, free dye molecule and a little amount of uncompleted surfactant micelle.
The free saponin molecule is an amphiphilic substance with both of hydrophobic
and hydrophilic part. While the PES membrane use in this study was tend to be
hydrophobic. Because of this characteristic, some of the dye was attached to the
saponin structure and the saponin structure can be attach in the membrane surface.
On the membrane surface, the concentration of solute (saponin and dye) was
increased inducing a concentration polarization (Cordove et al., 2016). Hence,
when the solution filtered by PES membrane the molecule will deposit on the
membrane surface, causing fouling over the time of filtration, and induce membrane
pore blocking. The illustration of blocking condition based on the micelle formation
is presented in Figure 4.16. Considering the blocking mechanism, the MEUF of dye
wastewater using saponin as the surfactant should be conducted with saponin
concentration above its CMC.

77
 (a) (b)

(c) (d)

Figure 4.16 Illustration of membrane blocking based on the formation of

surfactant micelle, (a) without saponin, (b) below CMC, (c) at CMC,
(d) above CMC

The determination of fouling mechanism also confirmed by the FTIR and

SEM analysis. Figure 4.17 shows the FTIR spectra of clean membrane and fouled
membrane. Generaly, the spectra of both fouled membrane shows a similar spectra
with the clean membrane. However, the fouled membrane without addition of
saponin shows a peak of C-N and O=S=O at 1010.69 and 918.12 respectively. The
peaks was reflecting the specific fungtional group of remazol blue as the foulant. A
weak peak at 3402.43 cm-1 also can be assumed to be the small amount of –OH
group from the remazol blue. The molecular structure of remazol blue can be seen
in Figure 4.12(A). As for fouled membrane with addition of saponin, a similar peaks
of C-N and O=S=O also showed at 1041.56 cm-1 and 910.4 cm-1 respectively. Look

78
after the no saponin fouled membrane, the peaks reflects the structure of remazol
blue dye. Another peak of –OH and –CH stretch vibration at 3302.13 and 2939.52
respectively. Those peak shows the functional group of saponin molecule. As
expected from the modeling calculation, either in present of saponin or not, the
remazol dye remained as a foulant on the membrane surface. The addition of
saponin in the ultrafiltration feed also showed an addition of saponin foulant. The
saponin foulant was identified as the saponin monomer as the micelle functional
group was not identified.

Figure 4.17. FTIR Spectra of clean PES membrane and fouled membrane.

The SEM analysis was also conduct to confirm the membrane fouling
mechanism. The SEM figure of clean and fouled membrane was showed in Figure
4.18. The figure shows that the used membrane was fouled compared to the clean
membrane. The thick layer of cake fouling was identified from the surface of
membrane use to filter the feed with saponin concentration under CMC. This result
was correspondence to the modeling calculation which claims that the process with
saponin addition under CMC concentration shows cake formation fouling
mechanism. While the other fouled membrane shows a thiner fouling. However,
different type of fouling was found from the SEM analysis. The fouled membrane

79
without addition of saponin shows an organic fouling, a similar appearance of
organic fouling was found from the previous research by Hu et al. (2013). It is
expected, as the dye wastewater model was made using remazol dye, which is an
organic dye. On the other hand, by the addition of saponin in the feed, a colloidal
fouling was shown. Colloidal fouling refers to a fuling of membrane surface caused
by the colloids or particles depositing on the membrane materials (Khayet, 2016).
The common colloidal fouling formed by organic macromolecules in the feed
solution, such as polysaccharides (Tang et al., 2011). Saponin is one of a
polysaccharides surfactant, therefore the colloidal deposition of saponin in the
membrane surface is possible. According to the modelling calculation, the addition
of saponin above the CMC will change the fouling mechanism from cake formation
to the complete blocking. The SEM analysis shows a compatible results, where, the
membrane use to filter wastewater with addition of saponin above the CMC shows
less fouling.
(a) (b)

(c) (d)

Figure 4.18 SEM figure of (a) clean membrane, and fouled membrane (b) without
saponin, (c) with saponin under CMC, (d) with saponin above saponin

80
 CHAPTER V
CONCLUSION AND SUGESTION

5.1 CONCLUSIONS
In this study, the extraction of saponin from the pericarps of Sapindus rarak
was extracted under various extraction condition. Maceration extraction (ME) and
ultrasound-asissted extraction (UAE) methods was used and compared. The
investigation of surfactant characteristics for the saponin extracts has conducted by
FTIR, measurement of CMC, HLB number analysis, and the surfactant ability to
solubilize dye. The MEUF system performance to remove dye pollutant was also
investigated using saponin as the natural surfactant. The research results were,

1.) The saponin extraction giving highest saponin yield was conducted under the
temperature of 30oC, at ratio of 10 mL/gr, running for 40 minutes using UAE
methods. The highest saponin yield obtained was 27.87125 mg of
saponin/100 mg dry-grounded Sapindus rarak feed. Compared to the ME, the
results of UAE showing a better performance of faster extraction time, lower
temperature, and less solvent requirement.
2.) The FTIR analysis shows that both of pure saponin and saponin extract has a
similar functional group and the structure depicts the structure of
monodesmosidic oleanane triterpenoid saponins. The CMC of pure saponin
and extracts saponin were 0.07% w/w of saponin content and 7% w/w of
extract saponin / 0.2% w/w of saponin content, respectivey. Increase of
saponin concentration above CMC helps to stabilize the foam. The HLB
number of 18.7 and 16.8 were found for pure saponin and extract saponin
respectively.
3.) From the dye solubilization study it is known that addition of more saponin
in dye solution increased the solubilized dye. The solubilization power (SP)
of extract rarak is similar with the pure saponin for both remazol dye. The ΔG
of Sapindus rarak saponin and pure commercialized saponin for remazol red
and blue showing a negative value, suggests that, the process was feasible and
spontaneous.

81
4.) The MEUF performance analysis presented that the highest flux profile was
achieved without any addition of saponin, but the %rejection of dye was very
low. In the other hand, the addition of surfactant decrease the flux value until
a certain point, but increasing the %rejection of dye molecule. Highest %
rejection of dye of 97.02% and 99.42% for remazol red and blue respectively,
at the saponin concentration of 2 times CMC. A very low Lm was found for
the solution with saponin below CMC, in both of remazol dye. At the CMC,
the micelle loading increased significantly. However, further addition of
saponin leads to decrease the Lm into 0.0352 mM/mM and 0.0428 mM/mM
for remazol red and blue respectively. The fouling mechanism of UF process
was standard blocking, UF process with addition of saponin below CMC
shows a cake formation blocking, and the addition of saponin above CMC
shows complete blocking mechanism.

5.2 SUGESTIONS
This research is expected to contribute for further research related to the
application natural-based saponin for MEUF system and can be applied to achieve
an efficient methods to treat dye wastewater. Further investigation is definitely
needed to obtain more comprehensive study, such as its application for more
variative pollutant. The investigation for different kind of pollutant with various
characteristic (ionic, hydrophobic, etc.) will help to understand the solubilization
nature of saponin as well. To accomplish more thorough knowledge of MEUF using
natural-based surfactant, further study using such as the combination of saponin
with ionic surfactant, and a study using real wastewater from textile industry will
also help. Another plant source of saponin also suggested to be studied more in the
future such as, Jatropha curcas L., Acacia collinsii.

82
 REFFERENCE

Abrahart EN, 1977. Dyes and their Intermediates. New York: Chemical Publishing,
pp. 1–12.
Acero, J. L., Benitez, F. J., Real, F. J., Teva, F., 2017, Removal of emerging
contaminants from secondary effluents by micellar-enhanced ultrafiltration,
Separation and Purification Technology 181, 123–131.
Ahmad, A.L., Puasa, S.W., Zulkali, M.M.D., 2006, Micellar-enhanced
ultrafiltration for removal of reactive dyes from an aqueous solution,
Desalination 191, 153–161.
Alighourchi, H. R., Barzegar, M., Sahari, M. A. and Abbasi, S., 2013, Effect of
sonication on anthocyanins, total phenolic content, and antioxidant capacity
of pomegranate juices, International Food Research Journal 20(4): 1703-
1709.
Allen R.L.M., 1971, Colour Chemistry. London: Thomas Nelson and Sons Ltd. pp.
11–13.
Almutairi, M. S., and Ali, M., 2014, Direct detection of saponins in crude extracts
of soapnuts by FTIR, Natural Product Research, 1-5.
Aoudia, M., Allal, N., Djennet, A., and Toumi, L., 2003. Dynamic micellar
enhanced ultrafiltration: use of anionic (SDS)–nonionic (NPE) system to
remove Cr3+ at low surfactant concentration. Journal of Membrane Science.
217:181–192.
Aryanti, N., DH Wardhani, S Supandi, 2016, Flux profiles and mathematical
modeling of fouling mechanism for ultrafiltration of konjac
glucomannan,Scientific Study & Research. Chemistry & Chemical
Engineering, Biotechnology, Food Industry, 17(2).
Aryanti, N., Sandria, F. K. I., Putriadi, R. H., & Wardhani, D. H. 2017 (b).
Evaluation of MicellarEnhanced Ultrafiltration (MEUF) Membrane for
Dye Removal of Synthetic Remazol Dye Wastewater. Engineering Journal,
21(3), 23-35.
Aryanti, N., A Saraswati, RP Putra, A Nafiunisa, DH Wardhani, 2018, Fouling
Mechanism Of Micelle Enhanced Ultrafiltration With Sds Surfactant For
Indigozol Dye Removal, Jurnal Teknologi 80 (3-2).
Azmir J., Zaidul, I.S.M., Rahman, M.M., Sharif, K.M., A. Mohamed, F. Sahena,
M.H.A. Jahurul, Ghafoor, N.A.N. Norulaini, A.K.M. Omar, 2013,
Techniques for extraction of bioactive compounds from plant materials: A
review, Journal of Food Engineering 117 426–436.

83
Bade, R., Lee, S, H., Jo, S.S, Lee, H.S. and Lee, S.E., 2008. Micellar enhanced
ultrafiltration and Activated Carbon Fibre hybrid processes for chromate
removal from wastewater. Desalination, 229 (1-3), 264-278.
Bade R., Lee S. H., 2011, A Review of Studies on Micellar Enhanced Ultrafiltration
for Heavy Metals Removal from Wastewater, Journal of Water Sustainability,
Volume 1, Issue 1, 85–102.
Baek K., Yang J.W., Lee H.H., 2003, Micellar-enhanced ultrafiltration for
simultaneous removal of ferricyanide and nitrate, Desalination 158(1):157-
166.
Bart, H. -J., 2011. Extraction of natural products from plants—An introduction.
Industrial scale natural products extraction (pp. 1–26) (1st ed.). Verlag:
Wiley-VCH.
Behera, M. R., Shailesh Ravi Varade, and Pallab Ghosh, 2014, Foaming in Micellar
Solutions: Effects of Surfactant, Salt, and Oil Concentrations, Ind. Eng.
Chem. Res., 53, 18497−18507.
Belhaij, A., and Osama Al-Mahdy, 2015, Foamability and Foam Stability of
Several Surfactants Solutions: The Role of Screening and Flooding, Journal
of Petroleum & Environmental Biotechnology, 6:4.
Bielska, M., Szymanowski, J., 2006, Removal of methylene blue from waste water
using micellar enhanced ultrafiltration, Water Research 40 1027– 1033.
Bielska, M., A. Sobczyn ´ska, K. Prochaska, 2009, Dye–surfactant interaction in
aqueous solutions, Dyes Pigm. 80, 201–205.
Bird, R. Byron; Stewart, Warren E.; Lightfoot, Edwin N. (1960). Transport
Phenomena. Wiley. ISBN 0-471-07392-X.
Boruah, B and Gogoi, M., 2013, Plant based natural surfactants, Asia Journal of
Home Science 8(2), 759-762.
Böttcher, S., Drusch, S., 2017, Saponins—Self-assembly and behavior at aqueous
interfaces, Advances in Colloid and Interface Science 243 (2017) 105–113.
Cacace, J.E., Mazza, G., 2003, Extraction of Anthocyanins and Other Phenolics
from Black Currants with Sulfured Water, J. Agric. Food
Chem.,50,5939−5946.
Cacace, J.E., Mazza, G., 2003, Mass transfer process during extraction of phenolic
compounds from milled berries, Journal of Food Engineering 59: 379–389.
Chang, E.-E., Yang, S.-Y., Huang, C.-P., Liang, C.-H., and Chiang, P.-C.,
2011.Separation and Purification Technology. 79:329– 336.
Chen, J., Zhang, S., Yang, X., 2013, Control of brown rot on nectarines by tea
polyphenol combined with tea saponin, Crop Protection 45, 29-35.

84
Chen, X.-W., Yang, D.-X., Zou, Y., Yang, X.-Q., 2017, Stabilization and
functionalization of aqueous foams by Quillaja saponin coated nanodroplets,
Food Research International 99: 679–687.
Cheng, K., Gao, H., Wang, R-R., Liu, Y., Hou, Y-X., Liu, X-H., Liu, K., and Wang,
W. (2017) Evaluation of Extraction and Degradation Methods to Obtain
Chickpeasaponin B1 from Chickpea (Cicer arietinum L.), Molecules, 22
(332): 1-13.
Cheok, Choon Yoong., Hanaa Abdel Karim Salman, Rabiha Sulaiman, 2014,
Extraction and quantification of saponins: A review, Food Research
International 59, 16–40.
Chung, Y. S., S.H. Yoo, C.K. Kim., 2009, Effects of membrane hydrophilicity on
the removal of a trihalomethane via micellar-enhanced ultrafiltration process,
Journal of Membrane Science 326, 714–720.
Colivet J., A.L. Oliveira, R.A. Carvalho, 2016, Influence of the bed height on the
kinetics of watermelon seed oil extraction with pressurized ethanol, Sep.
Purif. Technol. 169: 187–195.
Córdova, A., Astudillo, C., Guerrero, C., Vera, C., Illanes, A., 2016, Assessment of
the fouling mechanisms of an ultrafiltration membrane bioreactor during
synthesis of galacto-oligosaccharides: Effect of the operational variables,
Desalination 393: 79–89.
Corrales M., García, A.F., P. Butz, Tauscher, B., 2009, Extraction of anthocyanins
from grape skins assisted by high hydrostatic pressure, J. Food Eng. 90: 415–
421.
D’Alessandro, L.G., Dimitrov, K., Vauchel, P., Nikov, I., 2013, Kinetics of
ultrasound assisted extraction of anthocyanins from Aronia melanocarpa
(black chokeberry) wastes, Chemical Engineering Research and Design.
Da Silva BP, de Sousa AC, Silva GM, Mendes TP, Parente JP. 2002. A new
bioactive steroidal saponin from Agave attenuata. Z Naturforch. 57c:423–
428.
Dai, J., and Russell, J., Mumper, 2010, Plant Phenolics: Extraction, Analysis and
Their Antioxidant and Anticancer Properties, Molecules, 15, 7313-7352.
Danis, U., Aydiner, C., 2009, Investigation of process performance and fouling
mechanisms in micellar-enhanced ultrafiltration of nickel-contaminated
waters, Journal of Hazardous Materials 162: 577–587.
Das C, DasGupta S, De S., 2008, Prediction of permeate flux and counterion
binding during crossflow micellar-enhanced ultrafiltration. Colloids
Surfaces, A. Physicochem. Eng. Asp., 318: 125-33.
Date, A.,Ishizuka, R., and Matubayasi, N., 2016, Energetics of nonpolar and polar
compounds in cationic, anionic, and nonionic micelles studied by all-atom

85
 molecular dynamics simulation combined with a theory of solutions, Phys.
Chem. Chem. Phys.
De, Shirshendu and Mondal, Sourav, 2012, Micellar Enhanced Ultrafiltration
Fundamentals & Applications,CRC Press: Taylor & Francis Group, LLC.
Dini, I., Schettino, O., Simioli, T., & Dini, A., 2001. Studies on the constituents of
Chenopodium quinoaseeds: Isolation and characterization of new triterpene
saponins.Journal of Agricultural and Food Chemistry, 49,741–746.
El-Abbassi A., Khayet, M., Hafidi, A., 2011, Micellar enhanced ultrafiltration
process for the treatment of olive mill wastewater, water research 45, 4522-
4530.
Fajriaty, I, Hariyanto I.H., Saputra, I.R., Silitonga, M., 2017, Skrining fitokimia dan
analisis kromatografi Lapis tipis dari ekstrak etanol Buah lerak (Sapindus
rarak), Jurnal Pendidikan Informatika dan Sains, Vol.6, No. 2.
Falleh, H., Ksouri, R., Lucchessi, M.E., Abdelly, C., and Magné, C., 2012,
Ultrasound-Assisted Extraction: Effect of Extraction Time and Solvent
Power on the Levels of Polyphenols and Antioxidant Activity of
Mesembryanthemum edule L. Aizoaceae Shoots, Tropical Journal of
Pharmaceutical Research; 11 (2): 243-249.
Folmer, B.M., and B. Kronberg, 2000, Effect of Surfactant–Polymer Association
on the Stabilities of Foams and Thin Films: Sodium Dodecyl Sulfate and
Poly(vinyl pyrrolidone), Langmuir 16:5987.
Fuleki T., Ricardo-da-Silva, J.M., 2003, Effects of cultivar and processing method
on the contents of catechins and procyanidins in grape juice, J. Agric. Food
Chem. 51 (2003) 640–646.
Garcia-Salas, P., Morales-Soto, A., Segura-Carretero, A., Fernandez-Gutierrez, A.,
2010, Phenolic-compound-extraction systems for fruit and vegetable
samples. Molecules, 15, 8813–8826.
Gaud RS, Gupta GD. 2009, Practical Physical Pharmacy. CBS Publisher and
Distributors.71-72.
Özlem, G.-Ü. & Mazza, G., 2007, Saponins: Properties, Applications and
Processing, Critical Reviews in Food Science and Nutrition, 47:231–258.
Grzegorzek M., Majewska-Nowak K., 2018, The use of micellar-enhanced
ultrafiltration (MEUF) for fluoride removal from aqueous solutions,
Separation and Purification Technology 195, 1–11.
Guo, N, T. Tong, N. Ren, Y. Tu ,B. Li, 2018, Saponins from seeds of
GenusCamellia:Phytochemistry and bioactivity. Phytochemistry 149, 42-55.
He, A., Dong, B., Feng, X., Yao, S., 2018, Extraction of bioactive ginseng saponins
using aqueous two-phase systems of ionic liquids and salts, Separation and
Purification Technology 196, 270–280.

86
Heng, M. Y., Tan, S. N., Yong, J.W. H., & Ong, E. S., 2013, Emerging green
technologies for the chemical standardization of botanicals and herbal
preparations. Trends in Analytical Chemistry, 50, 1–10.
Hierro, J. N. d., Teresa Herrera, Mónica R. García-Risco, Tiziana
Fornari,Guillermo Reglero, Diana Martin, 2018, Ultrasound-assisted
extraction and bioaccessibility of saponins from edible seeds: quinoa, lentil,
fenugreek, soybean and lupin, Food Research International 109: 440–447.
Hostettmann, K., and Marston, A. 1995. Saponins.Cambridge University
Press,Cambridge, New York.
Huang, J-H., Zeng, G.M., Fang, Y. Y., Qu, Y.H., Li, X., 2009. Removal of cadmium
ions using micellar-enhanced ultrafiltration with mixed anionic-nonionic
surfactants, Journal of Membrane Science, vol. 326, pp. 303-309.
Huang J.H., Zhou, C.F., Zeng, G. M., Li, X., Huang, H.J., Niu, J., Li, F., Shi, L.J.,
He, S.B., 2012, Studies on the solubilization of aqueous methylene blue in
surfactant using MEUF, Separation and Purification Technology 98, 497–
502.
Huang J. H., Xiong, Y.L., Zeng, G.M., Guo, S.H., Xie, G.X., Zhang, D.M., Tang,
X.J Liu, Z.F., 2012, Separation of phenol from various micellar solutions
using MEUF, Separation and Purification Technology 98, 1–6.
Huang, J., Peng, L., Zeng, G., Li, X., Zhao, Y., Liu, L., Li, F., Chai, Q., 2014,
Evaluation of micellar enhanced ultrafiltration for removing methylene blue
and cadmium ion simultaneously with mixed surfactants, Separation and
Purification Technology 125: 83–89.
Huang J., Yuan, F., Zeng, G., Li, X., Gu, Y., Shi, L., Liu, W., Shi, Y., 2017,
Influence of pH on heavy metal speciation and removal from wastewater
using micellar-enhanced ultrafiltration, Chemosphere 173, 199-206.
Hwang, K.J., Lin, T., 2002, Effect of morphology of polymeric membrane on the
performance of cross-flow microfiltration, J. Membr. Sci. 199: 41–52.
Iqbal, J., Kim, H. J., Yang, J. S., Baek, K. and Yang, J. W., 2007. Removal of
arsenic from ground water by micellar-enhanced ultrafiltration (MEUF).
Chemosphere, 66 (5), 970-976.
Iritani, E., 2013, A review on modeling of pore-blocking behaviors of membranes
during pressurized membranefiltration, Dry. Technol. 31: 146–162.
Ismail, D. N., Suparmanto, N., Rahayu, D., Rusliyanto, 2012, Pengaruh Emulgator
Buah Lerak (Sapindus Rarak) Dengan Emulsi Air Dan Solar Sebagai Bahan
Bakar Motor Diesel Ramah Lingkungan, Pelita, Volume VII, Nomor 1, 29-
38.
Ji, S., Wang, Q., Qiao, X., Guo, H. -c., Yang, Y. -f., Bo, T., 2014. New triterpene
saponins from the roots ofGlycyrrhiza yunnanensisand their rapid screening

87
 by LC/MS/MS.Journal of Pharmaceutical and Biomedical Analysis, 90,15–
26.
Jonsson B, Lindman B, Holmberg K, Kronberg BJ. 1998. Surfactants and polymers
in aqueous solutions. Introdution to surfactants .
Jordens J., B. Gielen, L. Braeken, T.V. Gerven, 2004, Determination of the effect
of the ultrasonic frequency on the cooling crystallization of paracetamol,
Chem. Eng. Process. 84: 38–44.
Juang,R-S., Y-Y. Xu and C-L. Chen, 2003, Separation and removal of metal ions
from dilute solutions using micellar-enhanced ultrafiltration, J. Membr. Sci.,
218:257–267.
Karate,V.D., K.V. Marathe, 2008, Simultaneous removal of nickel and cobalt from
aqueous stream by cross flow micellar enhanced ultrafiltration, J. Hazard.
Mater. 157 : 464–471.
Kareru, P. G, Keriko, J. M., Gachanja, A. N., Kenji, G. M, 2008, Direct Detection
Of Triterpenoid Saponins In Medicinal Plants, Afr. J. Trad. CAM 5 (1): 56 –
60.
Katheresan V., Kansedo J., Lau S. Y., 2018, Efficiency of Various Recent
Wastewater Dye Removal Methods: A Review, Journal of Environmental
Chemical Engineering, https://doi.org/10.1016/j.jece.2018.06.060.
Khan, N. A., Bhadra, B.N., and Jhung. S., 2018. Heteropoly Acid-Loaded Ionic
Liquid Metal-Organic Frameworks: Effective And Reusable Adsorbents
For The Desulfurization Of A Liquid Model Fuel. Chemical Engineering
Journal 334: 2215-2221.
Khayet, M., 2016. Fouling and scaling in desalination. Desalination 393:1.
Khosa, M. A., Shah, S. S., & Feng, X., 2013. Micellar enhanced ultrafiltration of
organic dyes. Separation Science and Technology, 48(9), 1315-1323.
Kiełczyn´ski P., Szalewski, M., Balcerzak, A., K. Wieja, A. Malanowski, R.
Kos´ciesza, R. Tarakowski,, A.J. Rostocki, R.M. Siegoczyn´ski, 2014,
Determination of physicochemical properties of diacylglycerol oil at high
pressure by means of ultrasonic methods, Ultrasonics 54 : 2134–2140.
Kirmizigul S, Anil H. 2002. New triterpenic saponins fromCelphalaria
transsylvanica. Turk J Chem. 26:947–954.
Kobayashi, T., Kaminaga, H., Navarro, R.R., Iimura, Y., 2012, Application of
aqueous saponin on the remediation of polycyclic aromatic hydrocarbons-
contaminated soil, J. Environ. Sci. Health Part A 47: 1138–1145.
Kumar R., A.M. Isloor, A.F. Ismail, T. Matsuura, 2013, Performance improvement
of polysulfone ultrafiltration membrane using N succinyl chitosan as additive,
Desalination 318: 1-8.

88
Landaburu-Aguirre, J., García, V., Pongrácz, E., Keiski, R.L., 2009. The removal
of zinc from synthetic wastewaters by micellar-enhanced
ultrafiltration:statistical design of experiments, Desalination, vol. 240, pp.
262-269.
Lau, A. -J., Woo, S. -O., Koh, H. -L. 2003. A nalysis of saponins in raw and steamed
Panax notoginsengusing high-performance liquid chromatography with diode
array detection.Journal of Chromatography A, 1011,77–87.
Lee,J., J.S. Yang, H.J. Kim, K. Baek, J.W. Yang, 2005, Simultaneous removal of
organic and inorganic contaminants by micellar enhanced ultrafiltration with
mixed surfactant, Desalination 184:395–407.
Li, L., Zhang, J. -I., Sheng, Y. -x., Guo, D. -a., Wang, Q., & Guo, H.Z., 2005.
Simultaneous quantification of six major active saponins of Panax
notoginseng by highperformance liquid chromatography–UV method.Journal
of Pharmaceutical and Biomedical Analysis, 38,45–51.
Li, X., Zeng, G.M., Huang, J.H., Zhang, J. M., Shi, L. J., He, S. B., Min Ruan, 2011,
Simultaneous removal of cadmium ions and phenol with MEUF using SDS
and mixed surfactants, Desalination 276, 136–141.
Li, R., Wua, Z. L., Wang, Y.J., Li, L.L., 2013, Separation of total saponins from
the pericarp of Sapindus mukorossi Gaerten. by foam fractionation,
Industrial Crops and Products 51: 163– 170.
Liang Z., B. Wu, P. Fan, C. Yang, W. Duan, X. Zheng, C. Liu, S. Li, 2008,
Anthocyanin composition and content in grape berry skin inVitis germplasm,
Food Chem. 111: 837–844.
Liu, Z., Li, Z., Zhong, H., Zeng, G., Liang, Y., Chen, M., Wu, Z., Zhou, Y., Yu,
M., Hao, B., 2017, Recent advances in the environmental applications of
biosurfactant saponins: A review, Journal of Environmental Chemical
Engineering 5, 6030–6038.
Lukenheimer, K., and Malysa, K., 2003, Simple and Generally Applicable Method
of Determination and Evaluation of Foam Properties, Journal of Surfactants
and Detergents, Vol. 6, No. 1.
Luo, F., Zeng, G.-M., Huang, J.-H., Zhang, C., Fang, Y.-Y., Qu, Y.-H., Li, X., Lin,
D., Zhou, C.-F., 2010, Effect of groups difference in surfactant on
solubilization of aqueous phenol using MEUF, Journal of Hazardous
Materials 173, 455–461.
Majd, M.H.; Rajaei, A.; Bashi, D.S.; Mortazavi, S.A.; Bolourian, S. 2014,
Optimization of ultrasonic-assisted extraction of phenolic compounds from
bovine pennyroyal (Phlomidoschema parviflorum) leaves using response
surface methodology. Ind. Crop. Prod., 57, 195–202.

89
Marchese J., Ponce, M., Ochoa, N.A., Pradanos, P., Palacio, L., Hernandez, A.,
2003, Fouling behavior of polyethersulfone UF membranes made with
different PVP, J. Membr. Sci. 211 (2003) 1–11.
Materna, K., Goralska, E., Sobczynska, A. and Szymanowski, J., Recovery of
various phenols and phenylamines by micellar enhanced ultrafiltration and
cloud point separation, Green Chem., 2004, 176–182.
Melo R.P.F., Barros, E.L., Neto, Nunes, S.K.S., Castro, T.N., Dantas, A.A., Dantas
Neto, 2017, Removal of reactive blue 14 dye using micellar solubilization
followed by ionic flocculation of sufactants, Separation and Purification
Technology.
Misra, S. K., Mahatele, A.K., Tripathi, S.C., Dakshinamoorthy, A., 2009, Studies
on the simultaneous removal of dissolved DBP and TBP as well as uranyl
ions from aqueous solutions by using Micellar-Enhanced Ultrafiltration
Technique, Hydrometallurgy, vol. 96, pp. 47-51.
Morikawa, T., Xie, Y., Asao, Y., Okamoto, M., Yamashita, C., Muraoka, O.,
Matsuda, H., Pongpiriyadacha, Y., Yuan, D., Yoshikawa, M., 2009,
Oleanane-type triterpene oligoglycosides with pancreatic lipase inhibitory
activity from the pericarps of Sapindus rarak, Phytochemistry, 70, 1166–
1172.
Mostafa, A., Sudisha, J., El-Sayed, M., Ito, S. -I., Ikeda, T., Yamauchi, N., Shigyo,
M., 2013, Aginoside saponin, a potent antifungal compound, and secondary
metabolite analyses from Allium nigrum L. Phytochemistry Letters, 6,274–
280.
Mungray, A. A., Kulkarni, S. V. and Mungray, A. K., Removal of heavy metals
from wastewater using micellar enhanced ultrafiltration technique: a review,
Cent. Eur. J. Chem., 2011, 10,27–46.
Nataraj, S., Schomacker, R., Kraume, M., Mishra, I. M., and Drews, A., 2008.
Analyses of polysaccharide fouling mechanisms during crossflow membrane
filtration. Journal of Membrane Science. 308:152–161.
Ncube, B., Ngunge, V. N. P., Finnie, J. F., & Staden, J. V. 2011. A comparative
study of the antimicrobial and phytochemical properties between outdoor
grown and micropropagated Tulbaghia violacea Harv. plants. Journal of
Ethnopharmacology, 134, 775–780.
Noer S., Pratiwi, R.D., Gresinta, E., 2017, Penetapan Kadar Senyawa Fitokimia
(Tanin, Saponin Dan Flavonoid Sebagai Kuersetin) Pada Ekstrak Daun Inggu
(Ruta angustifolia L.), Eksakta: Jurnal Ilmu-ilmu MIPA.
Oakenful, D., 1986, Aggregation of saponins and bile acids in aqueous solution.
Aust. J. Chem., 39, 1671–1683.

90
Oancea, S., Grosu, C., Ketney O., and Stoia, M., 2013, Conventional and
Ultrasound-Assisted Extraction of Anthocyanins from Blackberry and Sweet
Cherry Cultivars, Acta Chim. Slov., 60, (2), 383–389.
Oberholster A., Francis, I.L., Iland, P.G., Waters, E.J., 2009, Mouthfeel of white
wines made with, without pomace contact, added anthocyanins, Aust. J.
Grape Wine Res. 15: 59–69.
Oleszek, W., and Hamed, A., 2009, Saponin-Based Surfactants, Surfactants from
Renewable Resources, Kjellin chapter 12.
Park Y.K., E. Park, J. Kim, Kang, M., 2003, Daily grape juice consumption reduces
oxidative DNA damage and plasma free radical levels in healthy Koreans,
Mut. Res. 529: 77–86.
Pelosi B. T., Lima, L. K. S. and Vieira, M. G. A., 2014, Removal of the Synthetic
Dye Remazol Brilliant Blue R from Textile Industry Wastewaters by
Biosorption on the Macrophyte Salvinia natans, Brazilian Journal of
Chemical Engineering, Vol. 31, No. 04, pp. 1035 – 1045.
Petigny, L., Périno-Issartier , S., Wajsman, J., and Chemat, F., 2013, Batch and
Continuous Ultrasound Assisted Extraction of Boldo Leaves (Peumus
boldusMol.), Int. J. Mol. Sci., 14, 5750-5764.
Puasa S.W, Ruzitah M.S and Sharifah A.S.A.K, 2011, An overview of Micellar –
Enhanced Ultrafiltration in Wastewater Treatment Process, IPCBEE vol.12
IACSIT Press, Singapore.
Purkait, M.K., DasGupta, S., and De, S. 2004. Separation of aromatic alcohols
using micellar enhanced ultrafiltration and recovery of surfactant. J. Membr.
Sci. 250: 47–59.
Purkait, M.K., S. DasGupta, S. De, 2005, Separation of aromatic alcohols using
micellar-enhanced ultrafiltration and recovery of surfactant, J. Membr. Sci.
250: 47–59. (a)
Purkait, M. K., DasGupta, S., and De, S. 2006, Micellar enhanced ultrafiltration
of eosin dye using hexadecyl pyridinium chloride, Journal of Hazardous
Materials, vol. 136, pp. 972-977, 2006.
Reichardt, C., & Welton, T. 2011.Solvents and solvent effects in organic chemistry
(4th ed.). Weinheim: Wiley-VCH (pp. 63- 457).
Reichert C.L., Salminen, H., Utz, J., Badolato B. G., Schäfer, C., Weiss, J., 2017,
Aging Behavior of Quillaja Saponin–Pea Protein Interfaces, Colloid and
Interface Science Communications 21, 15–18.
Rosello-Soto, E.; Galanakis, C.M.; Brncic, M.; Orlien, V.; Trujillo, F.J.; Mawson,
R.; Knoerzerf, K.; Tiwari, B.K.; Barba, F.J. 2015, Clean recovery of
antioxidant compounds from plant foods, by-products and algae assisted by

91
 ultrasounds processing. Modeling approaches to optimize processing
conditions. Trends Food Sci. Technol. 42, 134–149.
Roy D., Kommalapati, R. R., Mandava, S. S., Valsaraj, K. T., and Constant, W. D.,
1997, Soil Washing Potential of a Natural Surfactant, Environ. Sci. Technol.,
31,670-675.
Salleh-Mack S.Z., Roberts, J.S., 2007, Ultrasound pasteurization: the effects of
temperature, soluble solids, organic acids and pH on the inactivation of
Escherichia coliATCC 25922, Ultrason. Sonochem. 14: 323–329.
Samal K., Das, C., Mohanty, K., 2017a, Application of saponin biosurfactant and
its recovery in the MEUF process for removal of methyl violet from
wastewater, Journal of Environmental Management 203, 8-16.
Samal K., Das, C., Mohanty, K., 2017b, Eco-friendly biosurfactant saponin for the
solubilization of cationic and anionic dyes in aqueous system, Dyes and
Pigments 140, 100-108.
Samani B.H., Zareiforoush, H., Lorigooini, Z., Ghobadian, B., S. Rostami, E.
Fayyazi, 2016, Ultrasonic-assisted production of biodiesel fromPistacia
atlantica Desf.oil, Fuel 168: 22–26.
Saputra, E. H., 2018, Prospek Industri Tekstil masih Sangat Cerah,
http://mediaindonesia.com/read/detail/151724-prospek-industri-tekstil-
masih-sangat-cerah, accesed on 28 Mar 2018, 03:00 WIB.
Sarkar, B., DasGupta, S., De, S., 2009. Application of external electric field to
enhance the permeate flux during micellar enhanced ultrafiltration.
Separation and Purification Technology. 66:263–272.
Sarvin, B., Stekolshchikova, E., Rodin, I., Andrey Stavrianidi, Shpigun, O., 2018,
Optimization and comparison of different techniques for complete extraction
of saponins fromT. Terrestris, Journal of Applied Research on Medicinal and
Aromatic Plants, 8, 75–82.
Schmitt C., Grassl, B., G. Lespes, J. Desbriè res, V. Pellerin, and Reynaud, S., 2014,
Saponins: A Renewable and Biodegradable Surfactant From Its Microwave-
Assisted Extraction to the Synthesis of Monodisperse Lattices,
Biomacromolecules, 15, 856−862.
Schwarzea, M., Groß, M., Moritz, M., Buchner, G., L. Kapitzki, L. Chiappisi,
Gradzielski, M., 2015, Micellar enhanced ultrafiltration (MEUF) of metal
cations with oleylethoxycarboxylate, Journal of Membrane Science 478,
140–147.
Schwarze M., 2017, Micellar-enhanced ultrafiltration (MEUF) – state of the art,
Environmental Science: Water Research & Technology.

92
Shanei, A., Shanei, M.M., 2017, Effect of gold nanoparticle size on acoustic
cavitation using chemical dosimetry method, Ultrason. Sonochem. 34: 45–
50.
Sharma, A., & Sehgal, S. (1992). Effect of processing and cooking on the
antinutritional factors of faba beans (Vicia faba). Food Chemistry, 43, 383–
385.
Shockravi A., Vatanpour, V., Najjar, Z., Bahadori, S., Javadi, A., 2017, A new high
performance polyamide as an effective additive for modification of
antifouling properties and morphology of asymmetric PES blend
ultrafiltration membranes, Microporous and Mesoporous Materials 246,24-
36.
Silviani, Y., 2017, Short Communication: Inhibitory effect of Sapindus rarakethyl
acetate extract on Staphylococcus aureus, BIOTEKNOLOGI, Volume 14,
Number 1, Pages: 16-18.
Takeuchi, T. M., Pereira, C. G., Braga, M. E. M., Maróstica, M. R., Leal, P. F., &
Meireles, M. A.. 2009, Low-pressure solvent extraction (solid–liquid
extraction, microwave assisted, and ultrasound assisted) from
condimentary plants. In M. A. A. Meireles (Ed.), Extracting bioactive
compounds for food products—Theory and applications (pp. 140–144).
Boca Raton: CRC Press, 151-158.
Tang, C.Y., Chong, T.H., Fane, A.G., 2011. Colloidal interactions and fouling of
NF and RO membranes: a review. Adv. Colloid Interf. Sci. 164:126–143.
Tarade, K.M., Singhal, R.S., Jayram, R.V., Pandit, A.B., (2006). Kinetics of
degradation of saponins in soybean flour (Glycine max.) during food
processing, Journal of Food Engineering, 76: 440–445.
Tehrani-Bagha, A.R.; Singh, R.G.; Holmberg, K., 2012, Solubilization of two
organic dyes by cationic ester-containing gemini surfactants. J. Colloid Interf.
Sci., 376, 112–118.
Tehrani-Bagha, A.R.; Singh, R.G.; Holmberg, K., 2013, Solubilization of two
organic water-insoluble dyes by anionic, cationic and nonionic surfactants.
Colloid Surf. A, 417, 133–139.
Tippei J., Lehmann, M., Klitzing, R.V., Drusch, S., 2016, Interfacial properties of
Quillaja saponins and its use for micellisation of lutein esters, Food Chemistry
212, 35–42.
Tiwari B.K., Patras, A., N. Brunton, P.J. Cullen, O’Donnell, C.P., 2015, Effect of
ultrasound processing on anthocyanins and color of red grape juice,
Ultrasonics Sonochemistry 17, 598–604.

93
Tmáková L., Sekretár, S., Schmidt, S., 2015, Plant-derived surfactants as an
alternative to synthetic surfactants: surface and antioxidant activities,
Chemical Papers.
Tortora, F., Innocenzi, V., Celso, G. M-d., Vegliò, F., Capocelli, M., Piemonte, V.,
Prisciandaro M., 2018. Application of micellar-enhanced ultrafiltration in the
pre-treatment of seawater for boron removal. Desalination 428:21–28.
Toshiyuki M, Kazuhiro H, Masayuki Y. 2001. Medicinal Foodstuffs: XXIII
structures of new oleanane-type triterpene oligoglycosides, basellasaponins
A, B, C, and D, from the fresh aerial parts ofBasella rubraL. Chem Pharm
Bull (Tokyo). 49:776–779.
Udarno, L. 2009. Lerak (Sapindus rarak) Tanaman Industri Pengganti
Sabun. Warta Penelitian dan Pengembangan Tanaman Industri 2 (15). Bogor
: Badan Penelitian dan Pengembangan Perkebunan.
Urtiaga, A.M., G. P erez, R. Ib a~ nez, I. Ortiz, 2013, Removal of pharmaceuticals
from a WWTP secondary effluent by ultrafiltration/reverse osmosis followed
by electrochemical oxidation of the RO concentrate, Desalination 331: 26-34.
Urum K, Pekdemir T., 2004, Evaluation of biosurfactants for crude oil
contaminated soil washing. Chemosphere; 57: 1139-1150.
Verdinelli, V., Messina, P.V., Schulz, P.C., Vuano, B., 2008, Hydrophile–lipophile
balance (HLB) ofn-alkane phosphonic acids and theirs salts, Colloids and
Surfaces A: Physicochem. Eng. Aspects 316: 131–135.
Verhaagen,B., D.F. Rivas, Measuring cavitation and its cleaning effect, Ultrason.
Sonochem. 29 (2016) 619–628.
Vetal, M. D., Vikesh G. Lade, Virendra K. Rathod, 2012, Extraction of ursolic
acid from Ocimum sanctum leaves: Kinetics and modeling, food and
bioproducts processing 90: 793–798.
Víctor-Ortega, M.D., Martins, R. C., Gando-Ferreira, L.M., Quinta-Ferreira, R.M.,
2017, Recovery of phenolic compounds from wastewaters through micellar
enhanced ultrafiltration, Colloids and Surfaces A 531, 18–24.
Vilkhu, K., Mawson, R., Simons, L., Bates, D., 2008, Applications and
opportunities for ultrasound assisted extraction in the food industry—A
review. Innov. Food Sci. Emerg. Technol. 9, 161–169.
Vinatoru, M. (2001). An overview of the ultrasonically assisted extraction of
bioactive principles from herbs.Ultrasonics Sonochemistry, 8,303–313.
Vincent-Vela, M.C., S. Álvarez-Blanco, J. Lora-García, E. Bergantiños-Rodríguez,
2009, Analysis of membrane pore blocking models adapted to crossflow
ultrafiltration in the ultrafiltration of PEG, Chem. Eng. J. 149: 232–241.
Wina E., Muetzel, S., Hoffmann, E., Makkar, H.P.S., Becker, K., 2005, Saponins
containing methanol extract of Sapindus rarak affect microbial fermentation,

94
 microbial activity and microbial community structure in vitro, Animal Feed
Science and Technology 121, 159–174.
Wu, J., Lin, L., & Chau, F., 2001. Ultrasound-assisted extraction of ginseng
saponins from ginseng roots and cultured ginseng cells. Ultrasonics
Sonochemistry, 8(4), 347–352.
Xu, K., Zeng, G.-M., Huang, J.H., Wu, J.-Y., Fang, Y.Y., Huang, G., Li, J., Xi, B.
and Iu, H., 2007. Removal of Cd2+ from synthetic wastewaterusing
micellarenhanced ultrafiltraion with hollow fiber membrane. Colloids and
Surfaces A: Physicocemical and Engineering Aspects, 294 (1-3), 140-146.
Xu, D.P., Zhou, Y., Zheng, J., Li, S., Li, A.-N and Li, H.-B., 2015, Optimization of
Ultrasound-Assisted Extraction of Natural Antioxidants from the Flower of
Jatropha integerrima by Response Surface Methodology, Molecules, 21, 18.
Yohana O., 2009, Isolasi dan identifikasi aglikon saponin kecambah kacang hijau
(Phaseolus radiayusL.), Universitas Sanata Dharma, Yogyakarta.
Yolmeh, M., Najafi, M.B.H., Farhoosh, R., 2014, Optimisation of ultrasound-
assisted extraction of natural pigment from annatto seeds by response surface
methodology (RSM). Food chem., 155, 319–324.
Zaghbani,N., A. Hafiane, M. Dhahbi, 2007, Separation of methylene blue from
aqueous solution by micellar enhanced ultrafiltration, Sep. Purif. Technol.
55:117–124.
Zaghbani, N., Amor, H., Dhahbi, M., 2009. Removal of Direct Blue 71 from
wastewater using micellar enhanced ultrafiltration, Desalination and Water
Treatment, vol. 6, pp. 204-210.
Zaghbani, N., Amor, H, Dhahbi, M., 2008, Removal of Safranin T from wastewater
using micellar enhanced ultrafiltration, Desalination 222, 348–356.
Zahro L. dan Agustini R., 2013, Antibacterial effectivity test of saponins crude
extract from white oyster mushroom (Pleurotus ostreatus) against
Staphylococcus aureus and Escherichia coli, UNESA Journal of Chemistry
Vol. 2 No. 3.
Zeng, G.M., Li, X., Huang, J.-H., Zhang, C., Zhou, C-F., Niu, J., Shi, L.-J., He, S.-
B., Li, F., 2011, Micellar-enhanced ultrafiltration of cadmium and methylene
blue in synthetic wastewater using SDS, Journal of Hazardous Materials185,
1304–1310.
Zhang, Z., Zeng, G. M., Huang, J. H., Fang, Y. Y., Xu, K., Qu, Y. H., Yang, C.,
and Li, J. B., 2009. Removal of zinc ions from aqueous solution using
micellar-enhanced ultrafiltration at low surfactant concentrations. Water S.
A. 33(1):129–136.

95
Zhang, W., Liang, W., Huang, G., Wei, J., Ding, L., Jaffrin, M. Y., 2015. Studies
of membrane fouling mechanisms involved in the micellar-enhanced
ultrafiltration using blocking models. RSC Advance. 5(60):48484-48491.
Zhao, Z.G., 2006, Micellar Catalysis and Microemulsion Catalysis, Chemical
Industry Publishing Company, Beijing.
Zou, T.B., Min Wang, Ren-You Gan and Wen-Hua Ling, 2011., Optimization of
Ultrasound-Assisted Extraction of Anthocyanins from Mulberry, Using
Response Surface Methodology, Int. J. Mol. Sci., 12, 3006-3017.
Zhu, Z.,Guan, Q., Guo, Y., He, J., Liu, G., Li, S., Barba, F.J., and Jaffrin, M.Y.,
2016, Green ultrasound-assisted extraction of anthocyanin and phenolic
compounds from purple sweet potato using response surface methodology,
Int. Agrophys., 30, 113-122.

96