Food Chemistry 153 (2014) 35–43

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Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Physicochemical and biochemical properties of honeys from arid regions

Hosam M. Habib ⇑, Fatima T. Al Meqbali, Hina Kamal, Usama D. Souka, Wissam H. Ibrahim
Department of Nutrition and Health, College of Food and Agriculture, United Arab Emirates University, P.O. Box 15551, Al Ain, United Arab Emirates

a r t i c l e i n f o a b s t r a c t

Article history: This study was conducted to evaluate the quality of 11 honeys from arid regions for first time, and compare
Received 14 July 2013 it with 5 different honeys from non-arid regions. Mean values obtained for physicochemical parameters
Received in revised form 3 December 2013 were: pH 4.76 ± 0.55; 17.32 ± 1.8% moisture; 80.95 ± 1.60 °Brix sugar; 69.05 ± 4.41% total sugar;
Accepted 9 December 2013
413.81 ± 178.48 lS cm 1 electrical conductivity; 17.58 ± 7.68 meq/kg free acidity; 11.05 ± 3.18 meq/kg lac-
Available online 14 December 2013
tonic acidity; 28.63 ± 9.6 meq/kg total acidity; 12.66 ± 20.39 mg/kg HMF; 0.58 ± 0.03 water activity; and
0.98 ± 0.62 colour intensity. Potassium was the major mineral (1760.54 ± 685.24 mg/kg). All the samples
Keywords:
showed considerable significant variations with reference to their physicochemical and biochemical prop-
Honey
Colour characteristics
erties, moreover, the total free amino acids and total carotenoids were 61.13 ± 63.16 mg/100 g and
Physicochemical 4.07 ± 10.05 lg/100 g respectively. Acrylamide was detected only in one sample at 2.39 ± 0.22 lg/kg.
Biochemical properties Ó 2013 Elsevier Ltd. All rights reserved.
Arid regions

1. Introduction their essential physical, chemical and biological properties such as,
mineral, free amino acids, sugar and carotenoid profiles in different
Honey is the natural sweet product produced by Apis mellifera varieties of honey produced in arid regions. Therefore, the current
bees from nectar of plants (nectar honey), from secretions of study was conducted to assess the physicochemical properties
livings parts of plants or excretions of plant-sucking insects of composition and biochemical properties of honey samples from
the living part of plants (honeydew honey) (Silva, Videira, Monte- arid regions for the first time, as well as comparing it with different
iro, Valentão, & Andrade, 2009). This natural complex foodstuff is non-arid regions honey samples.
produced in almost every country and largely used as food source.
Honey cannot be considered a complete food by human nutritional
2. Materials and methods
standards, but it offers potential as a dietary supplement (Silva
et al., 2009). Honey mainly contains simple sugars or monosaccha-
2.1. Materials
rides [of which fructose and glucose are the main components
(65%)] and approximately 18% water, (Silva et al., 2009). Proteins,
All of the chemicals and reagents used were of analytical grade,
flavour and aroma, phenolic compounds (phenolic acids and flavo-
sucrose, fructose, glucose, maltose, formic acid, acrylamide, meth-
noids), free amino acids, organics acids, vitamins and minerals con-
anol, phosphoric acid, acetonitrile, bovine serum albumin, hexane,
stitute minor components of honeys (Silva et al., 2009). Honey
acetone, lutein, cryptoxanthine, zeaxanthine, lycopene, a-carotene,
commercially available varies greatly in quality all over the world.
b-carotene, c-carotene and ethyl acetate were purchased from Sig-
This is largely assessed on the basis of colour, flavour and density.
ma (St. Louis, MO, USA). Folin–Ciocalteu’s phenol reagent, HCl,
Honey composition is influenced by the plant species, climate,
FeSO4_7H2O, HMF, and standard solutions (1000 mg/l): [aluminum
environmental conditions and the contribution of the beekeeper.
(Al), arsenic (As), sulphur (S), cadmium (Cd), cobalt (Co), chromium
In general, honey is either monofloral or multifloral depending
(Cr), copper (Cu), iron (Fe), manganese (Mn), molybdenum (Mo),
on the source of the plant (Andrade et al., 1999; Anklam, 1998;
nickel (Ni), phosphorus (P), lead (Pb), vanadium (V), zinc (Zn), cal-
Azeredo, Azeredo, Souza, & Dutra, 2003; Gonzalez-Miret, Terrab,
cium (Ca), potassium (K), sodium (Na), magnesium (Mg), and
Hernanz, Fernandez-Recamales, & Heredia, 2005).
strontium (Sr)], were obtained from Merck (Darmstadt, Germany).
Several types of honeys are produced in arid regions. However,
AccQ_Tag kit was purchased from (Waters, Miliford, MA, USA).
the information available on their chemical and physical properties
is limited. Also there has been no particular research to determine
2.1.1. Honey samples
The present study was performed on eleven reputed commer-
⇑ Corresponding author. Tel.: +971 3 7136578; fax: +971 3 7675336. cial honey brands from arid regions (8 monofloral and 3 heterofl-
E-mail address: hosamh@uaeu.ac.ae (H.M. Habib). oral) and five from non-arid regions (3 monofloral and 2

0308-8146/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.foodchem.2013.12.048
36 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43

heterofloral) (Table 1). Fresh honey samples weighing 250–1 kg,
packed and sealed in glass bottles, were purchased from a local
market, and some samples provided directly by local UAE beekeep-
ers, and stored at 4 °C. The samples were diluted 10 time using
deionized water and were kept at 80 °C and analysed at the ear-
liest in such a way that none of the samples exceeded the storage
period beyond 6 months. The honey samples were thawed at ambi-
ent temperature before the analyses were performed.
2.2. Methods
2.2.1.5. Redox potential (Eh). Millivolt measurements were
performed at 20 °C using a pH-meter cyber scan pH 6000 (Eutech
instruments, Nijkerk, Netherlands). Honey samples were diluted
with freshly deionized water, ranging from 10% to 100% (w/v)
(Dimin ß s, Kuka, Kuka, & Cakste, 2006).
2.2.1.6. Ash. Ash was indirectly determined using the measured
electrical conductivity and applying the following equation:
X1 = (X2 0.143)/1.743, were: X1 = ash value; X2 = electrical con-
ductivity in lS/cm at 20 °C (Piazza, Accorti, & Persano Oddo, 1991).

2.2.1. Physicochemical analysis
2.2.1.1. Water content, RI, Brix. Water content was determined by
refractometry, measuring the refractive index (RI) according to
AOAC Methods (AOAC 969.38B, 2003), using a standard model
Abbe type refractometer at 20 °C. Water content (%) was then ob-
tained from the Chataway table.
2.2.1.2. Water activity. Water activity of liquid honey samples was
measured using an Rotronic Hygrolab (Rotronic Instrument Corp.
Hauppauge NY, USA) at 20 °C. (Acquarone, Buera, & Elizalde, 2007).
2.2.1.3. Acidity (free, lactone, and total). Free, lactone, and total acid-
ity were determined as follows by the titrimetric method (AOAC
962.19, 2003): 10 g honey samples were dissolved in 75 ml,
CO2-free water in a 250 ml beaker. The electrode of the pH meter
(Mettler Toledo Delta 320) was immersed in the solution, stirred
with a magnetic stirrer and titrated with 0.05 N NaOH to pH 8.5
(free acidity). Then the addition was stopped; immediately 10 ml
of 0.05 N NaOH were added and without delay back-titrated with
0.05 N HCl to pH 8.30 (lactone acidity). Total acidity resulted from
adding free plus lactone acidities. The results were expressed as
milliequivalents/kg (meq/kg).
2.2.1.4. pH determination. pH measurements were performed
potentiometricaly a 20 °C using a pH-meter Sartorius Professional
Meter PP-15 (Sartorius Ag, Goettingen, Germany) in Honey sam-
ples diluted with freshly deionized water, ranging from 10% to
100% (w/v) (Silva et al., 2009).
2.2.1.7. Electrical conductivity. Electrical conductivity was mea-
sured at 20 °C in solutions of honey samples in deionized water
with specific electrical conductivity lS/cm 1 using a conductivity
meter WTW 1970i (Werkstätten, GmbH, Germany), (Silva et al.,
2009).
2.2.1.8. Colour intensity. The net absorbance of the honey samples
was determined by the method of Beretta, Granata, Ferrero, Orioli,
& Facino, 2005. The honey samples were diluted to 50% (w/v) with
warm (45–50 °C) milli Q water and the solution was filtered
through a 0.45 lm filter. There was a complete absence of coarse
particles in the honey solutions as all the commercial samples
were non-crystalline liquid honeys. The absorbance was measured
using a spectrophotometer at 450 and 720 nm and the difference in
absorbance was expressed as mAU.
2.2.1.9. Colour. Visual colour was measured using Hunter colorim-
eter model ColorFlex (Hunter Associates Laboratory, Reston, VA,
USA) in terms of L (lightness), a (redness and greenness) and b
(yellowness and blueness). The instrument (45°/0° geometry,
10°observer) was calibrated with a standard black and white tile
followed by measurement of each honey samples (Beretta et al.,
2005).
2.2.2. Biochemical analysis
2.2.2.1. Sugar profile analysis. Honey (1 g) was dissolved in 10 ml
acetonitrile: water (1:1) solution. The mixture was homogenised
with constant shaking for at least 30 min. Then the samples were

Table 1
Classification of honey types and regional sources.

Sample Type of Family Botanical name Common Local name Region Sensory characteristics (colour,
honey name consistency)
Arid regions
H1 Monofloral Fabaceae Prosopis juliflora Ghaf Ghaf honey UAE Light amber, less viscous
H2 Monofloral Rhamnaccae Ziziphus spina-csisti Wild jujube Alain sider UAE Slightly dark amber, very less viscous
H3 Monofloral Fabaceae Acacia tortilis Wild mountain Ras ul Khaima UAE Light amber, less viscous
Samar
H4 Monofloral Rhamnaccae Ziziphus spina-csisti Wild jujube Oman sider Oman Light amber, less viscous
H5 Monofloral Fabaceae Acacia tortilis Wild mountain Oman samer Oman Slightly dark amber, viscous
H6 Monofloral Rhamnaccae Ziziphus spina-csisti Wild jujube Garden sider Yemen Dark amber, very viscous
H7 Monofloral Fabaceae Acacia tortilis Wild mountain Doany samer Yemen Light amber, viscous
H8 Monofloral Fabaceae Acacia tortilis Marya herbal Ashab marya Yemen Dark amber, viscous
samer
H9 Heterofloral – – Wild mountain Ashab gablaya UAE Light amber, viscous
H10 Heterofloral – – Herbs Ashab Omani Slightly dark amber, less viscous
H11 Heterofloral – – Mountain Ashab gbalya Yemen Light amber, viscous
herbal

Non arid regions

H12 Monofloral Rhamnaccae Ziziphus spina-csisti Wild jujube Pakistan sider Pakistan Light amber, viscous
H13 Monofloral Rhamnaccae Ziziphus spina-csisti Wild jujube Kashmir sider Kashmir Light amber, less viscous
H14 Monofloral Myrtaceae Leptospermum Manuka Manuka New Zealand Light amber, very fine granulated, medium
scoparium solid
H15 Heterofloral – – Black forest El ghabat el sawda KSA(Imported) Dark amber, less viscous
H16 Heterofloral – – Black forest El ghaba el sawda Germany Dark amber, less viscous
 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43
filtered through a 0.45 lm syringe filter. 1 ml of the concentrated
filtrate was then diluted with acetonitrile: water (1:1) solution to
a final volume of 25 ml. This mixture was then filtered again
through the 0.45 lm syringe filter into vials and set for injection
in the Waters ACQUITY UPLC with ELSD. The injection volume
was 10 ll, and separation was carried out on an ACQUITY UPLCÒ
BEH Amide Column C18 (2.1  150 mm, 1.7 lm) maintained at
35 °C with a run time of 20 min. The mobile phase A was 80/20
methanol/H2O with 0.2% triethylamine (TEA), and mobile phase B
was 30/70 methanol/H2O with 0.2% triethylamine (TEA) with a
flow rate of 0.17 ml/min, according to (Waters, ACQUITY UPLC
BEH Amide, Application Notebook, 2009).
2.2.2.2. Acrylamide. The procedure of sample preparation involved
an automated extraction method using accelerated solvent extrac-
tion unit ASEÒ 350 (DIONEX, CA, USA). Honey samples (5 g) were
extracted for 20 min using 10 mM formic acid in 34 ml cells of
the ASE 350. The conditions for the ASE extraction unit were set
at 70 °C, 1500 psi, with heat up time of 5 min and static time of
4 min, followed by 3 static cycles, flush volume 60%, and a purge
time (N2) of 120 s (DIONEX, CA, USA, application note 409). Quan-
tification of acrylamide in the above extracted honey samples was
performed on a UPLC–MS/MS with the electrospray positive ionisa-
tion (ESI+). In detail, an ACQUITY UPLC quaternary pump system
equipped with the micro vacuum degasser, thermostated autosam-
pler and thermostated column compartment (Waters, Milford, MA,
USA) was coupled with a Micromass Quattro Ultima triple-quadru-
pole mass spectrometer from Micromass Company Inc (Manches-
ter, UK). The analyte elution (injection volume 10 ll) was carried
out on a UPLC BEH C18 column (50 mm length, 2.1 mm i.d.,
1.7 lm particle size) (Waters, Milford, MA, USA) maintained at
25 °C with a run time of 3 min. The mobile phase was 10% metha-
nol/0.1% formic acid in water with a flow rate of 0.2 ml/min. The
conditions used for electrospray source were as follows: capillary
voltage, 3.5 kV; cone voltage, 50 V; source temperature, 100 °C;
desolvation gas temperature, 350 °C; desolvation gas flow, 400 L/
h nitrogen; cone gas flow, 45 L/h nitrogen; argon collision gas pres-
sure to 3  10 3 mbar for MS/MS, which gave a highest acrylamide
response in this study. The collision energy (CE) was optimised for
each multiple reaction monitored (MRM) transition (Zhang, Jiao,
Cai, Zhang, & Ren, 2007).
2.2.2.3. Hydroxymethylfurfural (HMF). Honey (1 g) was dissolved in
10 ml acetonitrile: water (1:1) solution. The mixture was homoge-
nised with constant shaking, for 10 min. This mixture was then fil-
tered again through the 0.45 lm syringe filter into vials and set for
injection in the Waters ACQUITY UPLC with TUV. The injection vol-
ume was 10 ll, and separation was carried out on an ACQUITY
UPLCÒ BEH Amide Column C18 (2.1  100 mm, 1.7 lm) main-
tained at 22 °C with a run time of 5 min. The mobile phase was
0.01 N Phosphoric acid (86%) and acetonitrile (14%) with a flow
rate of 0.2 ml/min (Chinnici, Masino, & Antonelli, 2003).
2.2.2.4. Minerals. Honey samples (1 g) were submitted to sequen-
tial digestion using CEM Mars 5 microwave digestion system. After
the digestion was completed, ultra-pure water was added up to a
final volume of 50 mL. The mineral components selected to be
quantified in the digested honey samples were: Al, As, Cd, Co, Cr,
Cu, Fe, Mn, Mo, Ni, P, Pb, Zn, Ca, K, Na, Mg, S and Sr. The concentra-
tion of these elements in the digested samples was determined
using inductively coupled plasma (Varian ICP-OES model 710-ES),
(Lachman et al., 2007).
2.2.2.5. Free amino acids. Free amino acids, histidine (His.), serine
(Ser.), arginine (Arg.), glycine (Gly.), aspartic acid (Asp.), glutamic
acid (Glu.), theronine (Thr.), alanine (Ala.), proline (Pro.), lysine
37
(lys.), tyrosine (Tyr.), methionine (Met.), valine (Val.), isoleucine
(LIc.), leucine (Leu.), phenylalanine (Phe.) and systine (Sys.), were
determined as described by Liming, Jinhui, Xiaofeng, Yi, and Jing
(2009).
2.2.2.6. Total protein. The total protein content was determined by
Lowry’s method of protein estimation which is based on the forma-
tion of a copper–protein complex and the reduction of phospho-
molybdate and phosphotungstate present in Folin–Ciocalteau
reagent to hetero polymolybdenum blue and tungsten blue,
respectively. Bovine serum albumin (BSA), (0–100 lg/ml) was used
as a standard for preparing the calibration curve (Saxena, Gautam,
& Sharma, 2010).
2.2.2.7. Carotenoids. Honey (5 g) was dissolved in 45 ml hexane:
acetone (60:40) solution. The mixture was homogenised with con-
stant shaking, for at least 10 min with addition of 5 ml water. The
mixture was then filtered through a Whatman 150 mm, 42 filter
paper and the filtrate was collected and dried under N2. After com-
plete drying the residue was re-dissolved in 2 ml of the mobile
phase containing acetonitrile: methanol: ethyl acetate
(730:200:70 ml) then filtered through 0.45 lm syringe filter into
vials and set for injection in the Waters ACQUITY UPLC with TUV.
The injection volume was 10 ll, and separation was carried out
on an ACQUITY UPLCÒ BEH Amide Column C18 (2.1  50 mm,
1.7 lm) maintained at 30 °C with a run time of 10 min. The mobile
phase was ran at a flow rate of 0.2 ml/min. ACQUITY UPLC, PDA
detector (450 nm) was used. lutein, cryptoxanthine, zeaxanthine,
lycopene, a-carotene, b-carotene, c-carotene were used as a
standards for preparing the calibration curve (Chauveau-Duriot,
Doreau, Noziere, & Graulet, 2010).
2.2.3. Statistical analysis
All analytical determinations were performed in triplicate.
Statistical analysis was performed using SPSS for windows (version
19; SPSS Inc., Chicago, IL, USA). The differences of mean ± values
among samples varieties was determined using one-way analysis
of variance (ANOVA) followed by Tukey.
3. Results and discussion
3.1. Physicochemical analysis
3.1.1. Water content, RI, Brix
Honey moisture content depends on the environmental condi-
tion and manipulation by beekeepers at the harvest period, and
it can vary from season to season and from year to year (Acquarone
et al., 2007). The moisture content (%) in the investigated samples
ranged from 13.63% to 20.60% (Table 2), which are within the al-
lowed parameters (620%) according to the international regula-
tions of quality (Codex Alimentarius, 2001). In our samples, the
values were similar to those previously reported for different kinds
of honey whose corresponding values ranged from 18.7% to 21.8%
(Manresa, 2005). Higher moisture content could lead to undesir-
able honey fermentation during storage caused by the action of
osmotolerant yeasts resulting in the formation of ethyl alcohol
and carbon dioxide; the alcohol can be further oxidised to acetic
acid and water resulting in a sour taste (Chirife, Zamora, & Motto,
2006). °Brix (directly related with sugar content) may be a reliable
index of adulteration. The analysed samples presented °Brix with-
out significant differences, ranging from 79.0 to 84.10 (aver-
age = 80.95 ± 1.60) (Table 2), which are similar to those from
others geographical locations (Silva et al., 2009. The Brix results
obtained in the current study suggest that the honey samples used
in the present study are most likely unadulterated. Similarly, RI
38 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43

Table 2
Physicochemical and biochemical properties of honeys.

pH mV Free acidity Lactonic acidity Total acidity HMF (mg/kg) Acrylamide Ash (%) Total protein (mg/
(meq/kg) (meq/kg) (meq/kg) (lg/kg) 100 g)

H1 4.61 ± 0.02ef 142.73 ± 0.40i 23.82 ± 0.47def 13.65 ± 0.10dcd 37.48 ± 0.50 g 4.45 ± 0.05d 2.39 ± 0.22 0.248 ± 0.000 h 345.20 ± 1.15b
H2 5.27 ± 0.00 k 102.97 ± 0.15b 15.23 ± 0.25bc 7.15 ± 0.25a 22.39 ± 0.11c 1.08 ± 0.05ab ND 0.077 ± 0.000b 402.33 ± 1.93d
H3 5.08 ± 0.00j 114.47 ± 0.42c 17.03 ± 0.13c 7.97 ± 0.15a 25.00 ± 0.15de 0.26 ± 0.05ab ND 0.114 ± 0.001e 453.74 ± 7.08 g
H4 4.24 ± 0.01b 160.67 ± 0.59k 16.33 ± 0.25bc 6.15 ± 0.51a 22.48 ± 0.71 cd 0.17 ± 0.01a ND 0.096 ± 0.000c 355.55 ± 0.87c
H5 4.44 ± 0.01d 150.07 ± 0.12j 25.55 ± 0.08f 15.01 ± 0.12d 40.57 ± 0.18 h 79.26 ± 0.87 h ND 0.116 ± 0.000e 578.87 ± 1.25i
H6 6.33 ± 0.02 m -16.00 ± 0.00a 3.86 ± 0.10a 14.79 ± 0.65d 18.66 ± 0.55b 1.16 ± 0.05b ND 0.316 ± 0.000 l 456.06 ± 4.17 g
H7 4.65 ± 0.03 fg 139.93 ± 0.25 h 4.28 ± 0.30a 6.60 ± 0.30a 10.88 ± 0.58a 15.33 ± 0.27e ND 0.077 ± 0.003b 204.84 ± 3.02a
H8 4.68 ± 0.02 g 135.40 ± 0.40f 15.98 ± 2.10bc 10.66 ± 0.25b 26.64 ± 2.33e 5.36 ± 0.46d ND 0.104 ± 0.001d 350.49 ± 2.84bc
H9 4.74 ± 0.01 h 133.40 ± 0.30e 23.31 ± 0.93de 12.51 ± 2.16bc 35.82 ± 1.24 g 0.75 ± 0.02ab ND 0.244 ± 0.000 g 399.79 ± 1.31d
H10 4.65 ± 0.01efg 137.70 ± 0.26 g 23.29 ± 0.08dc 14.15 ± 0.25 cd 37.44 ± 0.23 g 4.81 ± 0.13d ND 0.268 ± 0.000j 420.39 ± 2.81e
H11 4.61 ± 0.01e 140.03 ± 0.15 h 14.86 ± 0.62b 17.31 ± 1.01e 32.17 ± 1.28f 28.70 ± 0.30f ND 0.115 ± 0.000e 348.52 ± 0.68bc
H12 4.95 ± 0.01i 121.33 ± 0.21d 15.75 ± 0.08bc 10.56 ± 0.13b 26.31 ± 0.09e 2.47 ± 0.03c ND 0.285 ± 0.000 k 405.54 ± 1.32d
H13 5.32 ± 0.01 l 298.47 ± 0.25o 16.99 ± 0.15c 13.86 ± 0.13 cd 30.85 ± 0.03f 0.98 ± 0.07ab ND 0.262 ± 0.002i 435.81 ± 4.01f
H14 4.22 ± 0.00b 162.20 ± 0.17 m 28.45 ± 0.89 g 12.24 ± 0.30bc 40.69 ± 0.62 h 15.58 ± 0.52e ND 0.075 ± 0.000b 347.79 ± 1.11bc
H15 4.35 ± 0.01c 154.93 ± 0.15 l 25.34 ± 0.62ef 10.89 ± 0.26b 36.22 ± 0.88 g 4.95 ± 0.06d ND 0.196 ± 0.000f 475.14 ± 4.52 h
H16 3.99 ± 0.02a 176.53 ± 0.42n 21.81 ± 0.05d 13.99 ± 0.08 cd 35.80 ± 0.13 g 37.22 ± 0.22 g ND 0.066 ± 0.000a 427.70 ± 1.23ef
L a b Colour intensity RI Brix (%) Moisture (%) Water activity Electrical conductivity
(mAU) (wa) (lS cm 1)

H1 9.96 ± 0.12i 0.69 ± 0.10bc 11.80 ± 0.06gh 273.90 ± 2.40a 1.4879 ± 0.00a 79.00 ± 0.00a 19.40 ± 0.00 l 0.63 ± 0.00 k 570.67 ± 0.58 h
H2 7.60 ± 0.33 fg 2.19 ± 0.42a 7.50 ± 0.21de 1700.20 ± 4.60j 1.4901 ± 0.00a 79.80 ± 0.00a 18.60 ± 0.00j 0.60 ± 0.00 h 274.67 ± 0.58b
H3 7.85 ± 0.09 g 0.23 ± 0.63b 7.37 ± 0.14d 1238.65 ± 1.05 h 1.4849 ± 0.00a 77.90 ± 0.00a 20.60 ± 0.00 m 0.62 ± 0.00j 338.67 ± 1.15e
H4 7.27 ± 0.10ef 0.25 ± 0.16b 8.00 ± 0.30e 689.30 ± 0.00cde 1.4889 ± 0.00a 79.50 ± 0.00a 19.00 ± 0.00 k 0.56 ± 0.00d 307.33 ± 0.58c
H5 6.19 ± 0.09bc 1.91 ± 0.37 cd 6.64 ± 0.16c 679.85 ± 3.35cde 1.4980 ± 0.00a 82.80 ± 0.00a 15.40 ± 0.00c 0.53 ± 0.00b 341.67 ± 0.58e
H6 13.77 ± 0 .24j 3.11 ± 0.95d 16.02 ± 0.13i 653.35 ± 38.27 cd 1.5026 ± 0.00a 84.10 ± 0.00a 13.63 ± 0.29a 0.52 ± 0.00a 690.67 ± 0.58 l
H7 5.03 ± 0.04a 0.21 ± 0.17b 4.93 ± 0.04a 638.30 ± 2.00c 1.4915 ± 0.00a 80.50 ± 0.00a 18.00 ± 0.00i 0.64 ± 0.00 l 273.37 ± 5.74b
H8 6.83 ± 0.04de 0.42 ± 0.16b 6.05 ± 0.13b 976.90 ± 47.86 g 1.4980 ± 0.00a 83.00 ± 0.00a 14.80 ± 0.00b 0.55 ± 0.00c 320.67 ± 1.15d
H9 9.10 ± 0.32 h 1.73 ± 0.53a 9.50 ± 0.20f 746.80 ± 16.40def 1.4949 ± 0.00a 81.70 ± 0.00a 16.60 ± 0.00ef 0.57 ± 0.00 g 564.67 ± 0.58 g
H10 7.22 ± 0.23ef 1.99 ± 1.00 cd 7.51 ± 0.33de 835.30 ± 25.80f 1.4962 ± 0.00a 82.00 ± 0.00a 16.20 ± 0.00d 0.56 ± 0.00ef 605.67 ± 0.58j
H11 6.40 ± 0.07 cd 2.54 ± 0.35d 5.02 ± 0.20a 2874.80 ± 100.20 k 1.4884 ± 0.00a 79.20 ± 0.00a 19.20 ± 0.00kl 0.62 ± 0.00j 340.67 ± 0.58e
H12 8.84 ± 0.03 h 0.67 ± 0.28bc 10.26 ± 0.19 g 976.55 ± 3.65 g 1.4937 ± 0.00a 81.20 ± 0.00a 17.00 ± 0.00 g 0.57 ± 0.00f 636.67 ± 0.58 k
H13 10.17 ± 0.06i 0.66 ± 0.26bc 10.39 ± 0.03 g 617.10 ± 11.70bc 1.4955 ± 0.00a 81.80 ± 0.00a 16.40 ± 0.00de 0.57 ± 0.00 g 596.33 ± 3.21i
H14 8.88 ± 0.08 h 0.21 ± 0.36b 7.57 ± 0.17de 756.60 ± 2.90ef 1.4927 ± 0.00a 80.90 ± 0.00a 17.50 ± 0.00 h 0.55 ± 0.00c 270.33 ± 0.58b
H15 5.74 ± 0.17b 0.55 ± 0.33bc 5.93 ± 0.06b 1562.75 ± 30.35i 1.4945 ± 0.00a 81.40 ± 0.00a 16.80 ± 0.00 fg 0.56 ± 0.00e 480.33 ± 0.58f
H16 35.49 ± 0.17 k 14.50 ± 0.31e 36.54 ± 0.17 h 489.10 ± 1.50b 1.4919 ± 0.00a 80.40 ± 0.00a 18.00 ± 0.00i 0.60 ± 0.00i 254.67 ± 0.58a

Honey ± SD. Different letters in a column denote significant differences, P < 0.05.

values did not show any significant differences among the tested
samples, as presented in Table 2.
3.1.2. Water activity
The water activity of the honey samples varied from 0.52 to
0.64, with an average of 0.58 ± 0.03 as presented in Table 2. Our re-
sults are quite similar to those of Greek honeys for which the wa
values ranged from 0.53 to 0.67 (Lazaridou, Biliaderis, Bacandrit-
sos, & Sabatini, 2004). The water activity is an important factor,
which governs the food stability by preventing or limiting micro-
bial growth. The osmotolerant yeasts are able to grow at a minimal
water activity of 0.6 (Chirife et al., 2006). The honey sample H6,
which had the lowest moisture content was 13.63 ± 0.29%, had
the lowest water activity as well.
3.1.3. Acidity (free, lactone, and total)
Honey acidity is due to the presence of organic acids, mainly
gluconic acid, in equilibrium with their corresponding lactones or
internal esters, and to inorganic ions, such as phosphate, sulphate
and chloride (Terrab, Recalames, Hernanz, & Heredia, 2004). Table.
2 illustrate the free acidity, and lactonic acidity is considered as the
acidity reserve when the honey becomes alkaline, while the total
acidity is the sum of free and lactonic acidities. Table 2 illustrates
that the total acidity observed in the current study for different
honey samples were acceptable (below 50 meq/kg) (Codex
Alimentarius, 2001), indicating the absence of undesirable fermen-
tation. Lactonic acidity ranged from 6.15 ± 0.51 to 17.31 ±
1.01 meq/kg (average = 11.72 ± 3.31 meq/kg). Total acidity varied
between 10.88 ± 0.58 and 40.69 meq/kg, with a mean value of
29.96 ± 8.44 meq/kg. The results obtained for acidity were in
agreement with data reported from other geographical locations
(Terrab, Díez, & Heredia, 2002; Terrab et al., 2004). The variation
of total acidity has been attributed to harvest season.
3.1.4. pH
The pH is a parameter that is correlated with honey storage and
with microorganism growth that could change the texture and
honey stability (Feas, Pires, Iglesias, & Estevinho, 2010). The pH
limit is not described in EU Directive 2001/101/EC, however, honey
pH should be low to avoid microbiological contamination. The pH
values for studied honey samples averaged at 4.76 ± 0.55, and the
range was between 3.99 ± 0.02 and 6.33 ± 0.02 (Table 2), which
were acceptable values and comparable with those obtained in
other works (Corbella & Cozzolino, 2006; Feas, Pires, Estevinho,
Iglesias, & Araujo, 2010; Gomes, Dias, Moreira, Rodrigues, &
Estevinho, 2010).
3.1.5. Redox potential (Eh)
Table 2 shows the Eh values of honey samples. The range was
between 16.00 ± 0.00 and 298.47 ± 0.25 mV, with average of
141.93 ± 56.31 mV. H6 had the lowest redox potential, while H13
had the highest. Several substances found in honey participate in
 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43
oxidation–reduction processes. The oxidation–reduction potential
of honey partially indicates the characteristics of honey and
physicochemical processes in it during storage and treatment.
According to oxidation–reduction potential, honey can be divided
into different kinds. Every such kind of honey has its characteristic
oxidation–reduction potential (Dimin ß s et al., 2006). The redox
potential may also be an interesting indicator of the antioxidant
efficiency of food.
3.1.6. Ash
Ash content is a parameter used for the determination of the
botanical origin (floral, mix or honeydew). The results found
(0.066% to 0.316%) (Table 2) are within the limit allowed for hon-
eys (0.6%), indicating clearness of honey samples and possibly lack
of adulterations with molasses.
3.1.7. Electrical conductivity
The electrical conductivity of honey is closely related to the
concentration of mineral salts, organic acids and proteins. This
parameter shows great variability according to the floral origin
and it is important for the differentiation of honeys of different flo-
ral origins. The results obtained for the honey samples under study
varied between 154.67 ± 0.58 and 690.67 ± 0.58 lS cm 1 (aver-
age = 429.19 ± 152.78 lS cm 1). These values are below the maxi-
mum limit indicated by the international regulations of quality
(Codex Alimentarius, 2001) for nectar honey (800 lS cm 1), as
shown in Table 2.
3.1.8. The colour intensity
The colour intensity (ABS450) is supposed to be related to pig-
ments (carotenoids, flavonoids, etc.), which are also known to have
antioxidant properties (Frankel, Robinson, & Berenbaum, 1998).
The ABS450 values for the samples ranged from 524 to 1678 mAU
(Table 2). The reported ABS450 values for some Italian and Slove-
nian honeys are in the range of 25–3413 mAU and 70–495 mAU,
respectively (Beretta et al., 2005; Bertoncelj, Dobersek, Jamnik, &
Golob, 2007).
3.1.9. Colour
The colour characteristics are presented in Table 2, which sum-
marizes the means, standard deviations and ranges of the parame-
ters L⁄, a⁄ and b⁄, obtained with the Hunter colorimeter for honey
samples. The colour of the honeys was light to dark amber, with
reddish or green tinge. H16 and H6 had the highest average values
of parameter L⁄ that indicates lightness, 35.49 ± 0.17 and
13.77 ± 0.24, respectively, while H7 and H15 had the lowest aver-
age values of parameter L⁄, 5.03 ± 0.04 and 5.74 ± 0.17 respectively.
Honey samples analysed had red, yellow and green components:
green components (negative a⁄ values) for H9, H7, H2, H3, H14.
On the other hand, H16 and H6 also had the highest average values
of parameter b⁄ value at 36.54 ± 0.17 and 16.02 ± 0.13, respec-
tively. In general the parameters L⁄, a⁄ and b⁄ of the honey samples
analysed had significant variations among the colour parameter.
3.2. Biochemical analysis
3.2.1. HMF
HMF was investigated among the honey samples to determine
their quality. All samples analysed showed HMF values within
the parameters allowed according to the international regulations
of quality (Codex Alimentarius, 2001) for honeys of declared origin
from regions with a tropical or arid climate (lower than 80 mg/kg).
HMF content is widely recognised as a parameter of honey sample
freshness, because it is absent in fresh honeys and tends to increase
during processing and/or aging of the product. Several factors
influence HMF levels, such as temperature and time of heating,
39
storage conditions, pH and floral source, so it provides an indica-
tion of overheating and storage in poor conditions (Fallico, Arena,
Verzera, & Zappala, 2006). The data in Table 2 showed that H5
had the highest HMF values (79.26 ± 0.87 mg/kg) while H4 had
the lowest values (0.17 ± 0.01 mg/kg).
3.2.2. Acrylamide
Acrylamide is formed from reducing sugars and asparagine in
the Maillard reaction in a complex mechanism. Table 2 presents
the acrylamide in honey samples. All samples did not had acrylam-
ide except sample H1 which had very low amount of acrylamide
2.39 ± 0.22 lg/kg honey.
3.2.3. Minerals
Mean contents of each mineral found in the 16 honeys ex-
pressed in mg/kg fresh weight are shown in Table 3. Some minerals
were found in considerable amounts, and others in very small
amounts or not detected at all. The mineral content is an important
index of a possible environmental contamination when heavy
metals are detected. On the other hand, the major mineral content
contribution [potassium (K), calcium (Ca), sodium (Na), magne-
sium (Mg) and iron (Fe)] can provide also the nutritional value of
honey products and can be considered a potential indicator of
geographical origin of honey (Silva et al., 2009). The range of potas-
sium content was 86.00 ± 1.26–2690.29 ± 49.96 mg/kg, indepen-
dent of geographical origins; however, the lowest potassium
content was found in H7. On the other hand, H2, showed the
highest potassium contents. The mineral composition of so-
dium (6.76 ± 0.06–531.77 ± 1.37 mg/kg), calcium (7.87 ± 0.09–
183.90 ± 1.28 mg/kg), magnesium (2.28 ± 0.12–92.99 ± 1.21 mg/
kg), phosphorus (8.99 ± 0.10–264.18 ± 1.48), sulphur (6.97 ± 0.10–
333.73 ± 2.11), iron (1.15 ± 0.04–110.79 ± 1.32 mg/kg), manganese
(below <0.01 ± 0.00–10.31 ± 0.24), zinc (0.30 ± 0.02–6.73 ± 0.15)
and copper (0.26 ± 0.01–1.91 ± 0.01) were minority macro- and
micro-elements compared with potassium concentration in honey
samples. Nevertheless, the highest sodium and calcium contents
were obtained for H1, while H7 had the lowest value. The determi-
nation of potassium, sodium, magnesium, calcium, phosphorus,
sulphur and iron contents in honey is useful to determine the
nutritional value of some characteristic honey samples and to
establish a botanical origin differentiation. Nevertheless, the min-
eral content is not yet a quality parameter of the EU Directive (Co-
dex Alimentarius, 2001).
The total mineral contents of heavy metals in the honey sam-
ples ranged from 1.24 ± 0.03 to 13.93 ± 0.08, <0.010 to
3.96 ± 0.44, 0.06 ± 0.00 to 0.74 ± 0.01, <0.01 ± 0.00 to 13.31,
<0.02 ± 0.00 to 1.93 ± 0.05, and 0.00 to 7.78 ± 0.02 mg/kg for Al,
Sr, Cd, Cr, Mo, and Ni, respectively. While the readings for Pb, Co,
and As, were all below 0.01 mg/kg. These values can be compared
to the amount of heavy metals reported in New Zealand honey
(Vanhanen, Emmertz, & Savage, 2011). Overall, the amount of hea-
vy metals was comparable with other honey types analysed else-
where in the world.
3.2.4. Total protein content
Total protein contents are shown in Table 2. The highest values
of total protein was found in H5 (578.87 ± 1.25 mg/100 g of honey)
while the lowest values were found for H7(204.84 ± 3.02 mg/100 g
of honey). Honey protein level is dependent on the type of flora:
since it is variable, protein content in honeys can be attributed to
the presence of enzymes introduced by bees themselves, and oth-
ers derived from the nectar. Similar protein values in honey were
previously reported by Azeredo et al. (2003) and Escuredo, Miguez,
Fernandez-Gonzalez, and Seijo (2013).
40 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43

Table 3
Mineral contents of honeys samples are expressed as mg/kg honey.

K Na Ca Mg P Fe S Mn Zn Cu

H1 1415.53 ± 4.83c 531.77 ± 1.37 m 183.90 ± 1.28 h 70.37 ± 0.23 h 100.20 ± 0.35f 6.45 ± 0.05f 115.11 ± 0.97f 0.33 ± 0.01b 1.05 ± 0.02b 0.34 ± 0.02ab
H2 2690.29 ± 49.96j 99.27 ± 1.75f 71.93 ± 4.38 cd 28.01 ± 1.75b 38.28 ± 1.07b 3.06 ± 0.10bcd 52.75 ± 2.13b 0.01 ± 0.00a 1.33 ± 0.09bc 0.40 ± 0.02bc
H3 2476.46 ± 525.51ij 88.59 ± 0.75e 72.56 ± 3.79 cd 32.76 ± 2.08c 46.63 ± 0.68 cd 1.97 ± 0.10ab 57.55 ± 0.39bc 0.22 ± 0.01b 1.50 ± 0.03c 0.28 ± 0.05a
H4 2044.14 ± 9.82fgh 144.94 ± 2.50i 157.05 ± 6.46 g 75.30 ± 1.84i 110.74 ± 3.28 g 2.82 ± 0.27bc 170.92 ± 4.26 h 0.23 ± 0.04b 3.70 ± 0.18 g 0.49 ± 0.07de
H5 914.03 ± 10.39b 112.92 ± 1.37 h 118.60 ± 0.31f 57.25 ± 0.28g 113.73 ± 1.33 g 6.60 ± 0.06f 202.45 ± 2.44j 0.40 ± 0.00b 6.65 ± 0.25i 0.58 ± 0.02f
H6 2502.40 ± 19.86ij 56.79 ± 0.05d 77.68 ± 0.54 cd 39.69 ± 0.46e 39.95 ± 0.23b 2.81 ± 0.05bc 53.18 ± 0.50b 0.30 ± 0.00b 1.55 ± 0.02c 0.44 ± 0.00 cd
H7 86.00 ± 1.26a 6.76 ± 0.06a 7.87 ± 0.09a 2.28 ± 0.12a 8.99 ± 0.10a 1.15 ± 0.04a 6.97 ± 0.10a <0.01 ± 0.00a 0.30 ± 0.02a 0.26 ± 0.01a
H8 1715.36 ± 7.96cdef 146.57 ± 0.97i 110.69 ± 0.58ef 90.45 ± 0.33j 264.18 ± 1.48 k 3.02 ± 0.02bcd 333.73 ± 2.11 l 0.39 ± 0.00b 4.16 ± 0.07 h 0.70 ± 0.02 g
H9 2026.24 ± 6.10efgh 107.43 ± 0.39 g 76.67 ± 0.25 cd 37.86 ± 0.41de 49.35 ± 0.77d 3.58 ± 0.02cde 72.57 ± 0.99d 0.34 ± 0.01b 6.73 ± 0.15i 0.39 ± 0.01bc
H10 2205.65 ± 38.81ghi 20.54 ± 0.24b 66.78 ± 0.84c 92.99 ± 1.21j 192.97 ± 1.64j 4.86 ± 0.04e 159.34 ± 2.15 g 8.46 ± 0.10e 3.66 ± 0.02 g 1.91 ± 0.01j
H11 1800.19 ± 18.28cdfg 171.68 ± 2.03 k 248.20 ± 7.14i 160.65 ± 0.46 l 92.74 ± 0.21e 110.79 ± 1.32 h 321.68 ± 0.90 k 1.41 ± 0.01c 6.64 ± 0.02i 0.56 ± 0.01ef
H12 2333.63 ± 2.87hij 58.62 ± 0.26d 76.74 ± 1.32 cd 35.18 ± 0.76 cd 44.17 ± 0.75c 3.19 ± 0.08bcd 59.45 ± 2.18c 0.37 ± 0.01b 1.49 ± 0.04c 0.39 ± 0.02bc
H13 1923.97 ± 21.91defg 158.31 ± 1.88j 81.60 ± 0.36d 43.65 ± 0.32f 47.06 ± 1.13 cd 2.92 ± 0.07bc 78.05 ± 0.35d 0.25 ± 0.00b 3.58 ± 0.02 g 0.49 ± 0.03de
H14 1547.21 ± 71.97 cd 195.99 ± 2.72 l 161.99 ± 12.26 g 58.90 ± 3.28 g 121.28 ± 0.11 h 2.42 ± 0.07abc 180.73 ± 0.63i 0.27 ± 0.01b 2.81 ± 0.05e 0.44 ± 0.04 cd
H15 1637.47 ± 23.82cde 34.10 ± 0.61c 100.96 ± 1.88e 110.05 ± 0.50 k 162.45 ± 0.33i 4.27 ± 1.05de 107.38 ± 2.45e 10.31 ± 0.24f 3.29 ± 0.05f 1.58 ± 0.03i
H16 850.02 ± 2.12b 32.50 ± 0.40c 49.25 ± 0.47b 26.93 ± 0.15b 50.13 ± 0.51d 63.26 ± 0.06 g 52.09 ± 0.74b 4.74 ± 0.06d 2.04 ± 0.02d 1.15 ± 0.01 h
Al Sr Cd Cr Mo Ni Pb Co As

H1 3.07 ± 0.08e 2.19 ± 0.01 g 0.55 ± 0.09gh 1.09 ± 0.01c <0.02 ± 0.00a 0.10 ± 0.02b <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H2 3.73 ± 0.27 g 0.96 ± 0.07bc 0.21 ± 0.02bc <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H3 2.16 ± 0.04b 1.20 ± 0.07 cd 0.22 ± 0.00bc <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H4 1.96 ± 0.14b 3.96 ± 0.44 h 0.27 ± 0.02 cd <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H5 2.72 ± 0.04 cd 1.68 ± 0.00ef 0.38 ± 0.01ef 0.55 ± 0.01b <0.02 ± 0.00a 0.41 ± 0.04c <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H6 2.17 ± 0.05b 0.76 ± 0.01b 0.41 ± 0.04ef <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H7 1.24 ± 0.03a <0.00 ± 0.00a 0.14 ± 0.00ab <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H8 2.11 ± 0.03b 1.70 ± 0.02ef 0.06 ± 0.00a <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H9 2.46 ± 0.06c 1.12 ± 0.01bcd 0.46 ± 0.02 fg 0.08 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H10 13.93 ± 0.08 k 0.18 ± 0.00a 0.74 ± 0.01i <0.01 ± 0.00a <0.02 ± 0.00a 0.49 ± 0.04d <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H11 5.62 ± 0.12i 2.36 ± 0.05 g 0.17 ± 0.00bc <0.01 ± 0.00a <0.02 ± 0.00a 0.13 ± 0.02b <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H12 3.09 ± 0.06e 0.86 ± 0.05bc 0.64 ± 0.05 hi <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H13 3.42 ± 0.06f 1.43 ± 0.01de 0.65 ± 0.02hi <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H14 4.83 ± 0.05h 1.81 ± 0.16f 0.34 ± 0.01de <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H15 2.84 ± 0.04de 0.26 ± 0.00a 0.27 ± 0.06 cd <0.01 ± 0.00a <0.02 ± 0.00a 0.00 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a
H16 7.77 ± 0.06j 0.19 ± 0.00a 0.46 ± 0.00 fg 13.31 ± 0.25d 1.93 ± 0.05b 7.78 ± 0.02e <0.01 ± 0.00a <0.01 ± 0.00a <0.01 ± 0.00a

Honey ± SD. Different letters in a column denote significant differences, P < 0.05.

3.2.5. Sugar
The monosaccharaides glucose and fructose are the major con-
stituents of honey. Fructose is always the most important sugar
quantitatively followed by glucose. In the current study, fructose
was higher than glucose in all the honey samples analysed. Fruc-
tose and glucose contents of honey samples ranged from
32.26 ± 0.07 g/100 g to 42.42 ± 0.10 g/100 g and 27.78 ± 0.10 g/
100 g to 32.35 ± 0.04 g/100 g respectively. Sucrose, maltose and
raffinose were the minor sugars in honey as shown in Table 4;
the ranges were 0.56 ± 0.43–1.66 ± 0.01, 0.31 ± 0.02–2.09 ± 0.09
and 0.0010 ± 0.001–0.0079 ± 0.0032, respectively. Overall the total
sugar ranged from 61.21 ± 1.09 to 77.49 ± 0.63. All the honeys

Table 4
Sugar of honeys samples are expressed as percentage.

Fructose Glucose Sucrose Maltose Raffinose Total sugar F/G ratios

Sugar (%)
H1 37.16 ± 0.16f 27.78 ± 0.10a 0.98 ± 0.02b 0.48 ± 0.03ab 0.0065 ± 0.0005ab 66.41 ± 0.22bcd 1.34 ± 0.00e
H2 33.26 ± 0.05bc 31.72 ± 0.03c ND 0.31 ± 0.02a 0.0035 ± 0.0009ab 65.29 ± 0.09bc 1.05 ± 0.00a
H3 32.84 ± 0.09b 31.41 ± 0.09c ND 0.50 ± 0.02ab 0.0047 ± 0.0005ab 64.75 ± 0.09b 1.05 ± 0.01a
H4 33.14 ± 0.14bc 31.61 ± 0.05c 0.88 ± 0.01ab 0.52 ± 0.03ab 0.0064 ± 0.0005ab 66.15 ± 0.12bc 1.05 ± 0.00a
H5 42.42 ± 0.10j 31.64 ± 0.04c 1.60 ± 0.02c 0.59 ± 0.04ab 0.0050 ± 0.0020ab 76.25 ± 0.11 h 1.34 ± 0.00e
H6 36.44 ± 0.05de 28.59 ± 1.16ab 0.72 ± 0.00ab 0.98 ± 0.14abc 0.0038 ± 0.0013ab 66.74 ± 1.04bcd 1.28 ± 0.05d
H7 33.45 ± 0.05c 32.01 ± 0.10c 0.92 ± 0.02b 0.38 ± 0.05a 0.0053 ± 0.0023ab 66.77 ± 0.16 cd 1.04 ± 0.00a
H8 39.28 ± 0.27 h 32.25 ± 0.72c 1.66 ± 0.01c 4.29 ± 0.02d 0.0020 ± 0.0022ab 77.49 ± 0.63 h 1.22 ± 0.03c
H9 36.82 ± 0.07ef 29.28 ± 0.06b 0.56 ± 0.43a 1.72 ± 1.46bc 0.0070 ± 0.0076ab 68.39 ± 1.72de 1.26 ± 0.01cd
H10 39.87 ± 0.09i 32.35 ± 0.04c ND 0.44 ± 0.02ab 0.0010 ± 0.0018ab 72.65 ± 0.10 g 1.23 ± 0.00 cd
H11 32.26 ± 0.07a 27.93 ± 0.07a ND 1.02 ± 0.96abc 0.0079 ± 0.0032b 61.21 ± 1.09a 1.15 ± 0.00b
H12 38.32 ± 0.03 g 31.75 ± 0.43c 0.91 ± 0.01b 2.09 ± 0.06c ND 73.06 ± 0.51 g 1.21 ± 0.02bc
H13 38.95 ± 0.06 h 31.65 ± 0.05c 0.87 ± 0.01ab 2.09 ± 0.09c ND 73.56 ± 0.11 g 1.23 ± 0.00 cd
H14 39.21 ± 0.58 h 28.61 ± 0.52ab ND 1.01 ± 0.04abc ND 68.82 ± 0.92ef 1.37 ± 0.02e
H15 39.28 ± 0.06 h 29.37 ± 0.06b 1.40 ± 0.01c 0.48 ± 0.02ab 0.0047 ± 0.0005ab 70.54 ± 0.08f 1.34 ± 0.00e
H16 35.95 ± 0.06d 29.45 ± 0.05b ND 1.40 ± 0.04abc ND 66.80 ± 0.14 cd 1.22 ± 0.00c

Honey ± SD. Different letters in a column denote significant differences, P < 0.05.
ND, not detected.
 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43 41

Table 5
Carotenoids contents of honeys samples are expressed as lg/100 g honey.

Lutein Cryptoxanthine Zeaxanthine b-Carotene c-Carotene Total carotenoids

Carotenoids (lg/100 honey)
H1 0.0707 ± 0.0026ab 0.1558 ± 0.0041c 38.6644 ± 1.4230c ND ND 38.89 ± 1.42c
H2 0.0740 ± 0.0105b 0.0781 ± 0.0071ab 0.3082 ± 0.0224a ND ND 0.46 ± 0.04a
H3 0.3186 ± 0.0128f 0.0519 ± 0.0063a 0.3216 ± 0.0254a 0.0165 ± 0.0019c ND 0.71 ± 0.05a
H4 0.1842 ± 0.0064 cd 0.0615 ± 0.0038a 0.2777 ± 0.0208a 0.0151 ± 0.0005bc ND 0.54 ± 0.02a
H5 0.0136 ± 0.0029a ND 0.3644 ± 0.0212a 0.0820 ± 0.0087e ND 0.46 ± 0.02a
H6 ND 0.2263 ± 0.0269d 0.7667 ± 0.0654a ND ND 0.99 ± 0.09a
H7 0.3025 ± 0.0182ef 0.0917 ± 0.0020b 0.1020 ± 0.0191a 0.0324 ± 0.0056d ND 0.53 ± 0.04a
H8 0.4980 ± 0.0471 g 0.2752 ± 0.0180e 0.1463 ± 0.0216a 0.0964 ± 0.0049f ND 1.02 ± 0.04a
H9 0.0410 ± 0.0010ab ND 0.2688 ± 0.0379a ND ND 0.31 ± 0.04a
H10 0.4271 ± 0.0252 g 0.2321 ± 0.0032d 17.3199 ± 0.3182b 0.0173 ± 0.0022c 0.0787 ± 0.0092d 18.08 ± 0.32b
H11 0.4959 ± 0.0225 g 0.2209 ± 0.0137d 0.0796 ± 0.0098a 0.0820 ± 0.0012e ND 0.88 ± 0.03a
H12 0.1545 ± 0.0438c ND 0.1987 ± 0.0205a ND 0.0066 ± 0.0003ab 0.36 ± 0.05a
H13 0.0768 ± 0.0061b ND 0.3501 ± 0.0288a ND 0.0148 ± 0.0025c 0.44 ± 0.03a
H14 0.2813 ± 0.0055ef 0.0727 ± 0.0095 ab 0.5330 ± 0.0445a 0.0177 ± 0.0017c ND 0.91 ± 0.06a
H15 0.1593 ± 0.0234c ND 0.1572 ± 0.0285a 0.0067 ± 0.0012ab ND 0.32 ± 0.05a
H16 0.2473 ± 0.0475de ND ND 0.0113 ± 0.0008bc 0.0100 ± 0.0016bc 0.27 ± 0.05a

Honey ± SD. Different letters in a column denote significant differences, P < 0.05.
ND, not detected.

Table 6
Free amino acids of honeys samples are expressed as lg/100 g honey.

His. Ser. Arg. Gly. Asp. Glu. Thr. Ala. Pro.

Free amino acids (lg/100 g honey)

H1 419.11 ± 1.94c ND 168.13 ± 6.28 g 135.29 ± 5.15c ND 31.55 ± 5.27bc ND 45.00 ± 1.25e 2031.92 ± 7.51a
H2 ND ND 33.55 ± 0.33b 2194.42 ± 6.56 k 650.67 ± 1.28i 62.00 ± 0.78e 52.92 ± 0.72e 310.83 ± 1.91 k 7918.75 ± 6.96n
H3 ND 500.94 ± 1.24d 52.50 ± 2.17c 1415.42 ± 3.82j 11.58 ± 0.80b 105.00 ± 1.25 g 93.75 ± 1.25 g 244.58 ± 7.32i 7247.08 ± 6.17 m
H4 ND ND 297.79 ± 3.78 h 919.88 ± 57.52 g 62.34 ± 1.58f 0.46 ± 0.07a 0.62 ± 0.01a 11.92 ± 0.90bc 5224.17 ± 26.94j
H5 ND 359.92 ± 1.13c 2.34 ± 0.19a 735.42 ± 15.02e ND ND 2.46 ± 0.07a 6.21 ± 0.19a 5629.17 ± 19.09 k
H6 ND 360.37 ± 0.76c 81.25 ± 1.25d 1118.83 ± 9.12 h 30.00 ± 1.25c 5.00 ± 0.13a 11.96 ± 1.56bc 21.58 ± 0.80 cd 4943.33 ± 64.11i
H7 ND ND 132.54 ± 4.45e 1368.04 ± 6.50j 285.42 ± 1.91 h 37.17 ± 1.39c 48.71 ± 2.39e 294.25 ± 4.98j 2606.75 ± 20.07c
H8 1762.50 ± 8.50 g 410.83 ± 1.08c 720.58 ± 6.03j 361.13 ± 2.74d ND 269.79 ± 5.61i 261.25 ± 1.25 h 476.75 ± 6.31 m 2490.96 ± 15.99b
H9 632.30 ± 1.13d ND 437.09 ± 1.49i ND ND ND ND ND 2681.58 ± 5.65c
H10 ND ND 75.38 ± 0.90d 785.83 ± 6.41f 38.42 ± 0.31d 29.92 ± 0.19b 10.46 ± 0.19b 62.46 ± 1.31f 3021.67 ± 18.55e
H11 ND 756.83 ± 3.56e 147.38 ± 3.24f 2196.58 ± 10.22 k 57.50 ± 1.25e 50.00 ± 1.25d 18.33 ± 0.72c 120.83 ± 4.39 h 4070.00 ± 65.53 h
H12 3.49 ± 0.16a ND 29.13 ± 0.25b 3.70 ± 0.19a 1.77 ± 0.10a 29.92 ± 0.19b 3.67 ± 0.19a 11.29 ± 1.19b 3595.83 ± 26.02f
H13 792.50 ± 2.29e ND 1084.25 ± 10.82 l ND ND 81.09 ± 0.32f 0.55 ± 0.07a 24.21 ± 0.81d 6821.79 ± 3.94 l
H14 26.08 ± 0.15b 218.67 ± 1.26b 1.96 ± 0.64a 1268.75 ± 7.60i 111.29 ± 1.32 g 121.25 ± 1.25 h 80.42 ± 1.91f 85.00 ± 2.50 g 5609.58 ± 15.63 k
H15 ND ND ND ND ND ND 0.95 ± 0.01a 22.96 ± 0.14d 3695.42 ± 64.84g
H16 1739.29 ± 2.15f 103.08 ± 89.27a 943.96 ± 7.77 k 85.63 ± 2.84b ND 61.67 ± 0.85e 26.54 ± 7.63d 346.21 ± 3.81 l 2892.42 ± 6.57d
Lys. Tyr. Met. Val. Llc. Leu. Total FAA (mg/100 g)

Free amino acids (lg/100 g honey)

H1 ND ND ND ND ND ND 2.83 ± 0.02a
H2 1734.92 ± 14.54j ND 188.84 ± 1.44j 55.46 ± 0.71b 46.13 ± 0.13 h 1890.83 ± 14.81j 15.14 ± 0.04n
H3 1285.83 ± 1.91i ND 85.33 ± 0.80 g ND 3.71 ± 0.07bc 335.42 ± 1.91i 11.38 ± 0.02 m
H4 126.33 ± 1.26a ND 3.63 ± 0.13a ND 4.25 ± 0.13c 24.29 ± 0.69b 6.67 ± 0.09 g
H5 658.75 ± 3.31d 1.21 ± 0.07a 3.75 ± 0.13a ND ND ND 7.40 ± 0.03i
H6 813.25 ± 2.05e ND 10.38 ± 0.76c ND 1.21 ± 0.07a 36.63 ± 0.76c 7.44 ± 0.08i
H7 558.50 ± 3.77c ND 95.00 ± 1.25 h ND 15.50 ± 0.13f 49.17 ± 0.81d 5.49 ± 0.04f
H8 1009.21 ± 3.09 g 272.29 ± 2.49c 175.92 ± 0.50i ND 149.67 ± 0.80 k 253.71 ± 0.19 h 8.62 ± 0.02j
H9 546.50 ± 5.85c ND ND ND ND ND 4.30 ± 0.01d
H10 800.46 ± 1.02e 1.51 ± 0.02a 9.42 ± 0.19bc 65.42 ± 1.91c 25.88 ± 0.38 g 111.71 ± 1.48f 5.04 ± 0.02e
H11 937.50 ± 12.50f ND 23.33 ± 1.91d ND 11.63 ± 0.76e 97.92 ± 1.91e 8.49 ± 0.06j
H12 ND ND ND ND 59.50 ± 3.28j 3.75 ± 0.13a 3.74 ± 0.03b
H13 2227.79 ± 19.76 k 26.00 ± 0.38b 46.67 ± 2.93f 102.00 ± 0.76d ND ND 11.21 ± 0.04 l
H14 1162.08 ± 1.91 h ND 35.83 ± 1.91e ND 7.54 ± 0.19d 209.58 ± 2.60 g 8.94 ± 0.03 k
H15 360.00 ± 2.50b ND 5.96 ± 0.31ab ND 55.96 ± 0.89i ND 4.14 ± 0.07c
H16 655.59 ± 6.12d ND ND 45.00 ± 1.25a 16.50 ± 0.13f 17.46 ± 0.15b 6.93 ± 0.10 h

Honey ± SD. Different letters in a column denote significant differences, P < 0.05.
ND, not detected.

presented a value of glucose plus fructose higher than 60 g/100 g, evaluate honey granulation, because glucose is less water soluble
which is the value required for all the kinds of honey according than fructose. The proportion of fructose to glucose depends
to the international regulations of quality (Codex Alimentarius, largely on the nectar source (Anklam, 1998). Most investigators
2001). Fructose/Glucose (F/G) ratio has been recommended to reported an F/G average ratio around 1.2, which coincides with
42 H.M. Habib et al. / Food Chemistry 153 (2014) 35–43
our data for honeys according to the international regulations of
quality (Codex Alimentarius, 2001).
3.2.6. Carotenoids
The levels of different carotenoids are shown in Table 5. Lutein,
cryptoxanthine, zeaxanthine, b-carotene and c-carotene were
quantified in all honey samples, and results ranged from ND to
0.4980 ± 0.0471, ND to 0.2752 ± 0.0180, ND to 38.6644 ± 1.4230,
ND to 0.0964 ± 0.0049, ND to 0.0787 ± 0.0092 lg/100 g honey,
respectively. In general, the highest amount of carotenoids were
found in the honey from arid regions, and some honey values were
not within the quantification limit of the method. In Portuguese
honeys, Ferreira, Aires, Barreira, & Estevinho, 2009 reported carot-
enoid content in the range from 8.64 ± 0.06 mg/kg to
9.49 ± 0.15 mg/kg. These values are clearly much higher than the
values found in the present study. In this case it must be taken into
account that honey composition is rather variable and depends not
only on its floral source, but also on the geographical zone, as well
as on seasonal and environmental factors, which may be responsi-
ble for the detected differences. H1 from arid regions mono floral
had the highest amount of total carotenoids 38.89 ± 1.42 followed
by H10 from arid regions heater floral 18.08 ± 0.32.
3.2.7. Free amino acids
Free amino acid contents are shown in Table 6. H8 mono floral
honey from arid regions had the highest level of His.
(1762.50 ± 8.5 lg/100 g), while H11 hetero floral honey from arid
regions had the highest level of Ser. (756.83 ± 3.56 lg/100 g).
H13 mono floral from non-arid regions had the highest level of
Arg. (1084.25 ± 10.82 lg/100 g). H11 and H2 from arid regions
had the highest levels of Gly., 2196.58 ± 10.22, 2194.42 ± 6.56 lg/
100 g, respectively. H2, also from arid regions, had the highest lev-
els of Asp. (650.67 ± 1.28 lg/100 g). H8, a mono floral honey from
arid regions, exhibited the highest level of Glu. (269.79 ± 5.61 lg/
100 g). H3, a mono floral honey from arid regions had the highest
level of Thr. (93.75 ± 1.25 lg/100 g), while H16, a hetero floral hon-
ey from non-arid regions, had the highest level of Ala.
(346.21 ± 3.81 lg/100 g). In addition, Pro. was found in all honey
samples in quite large amount, ranging from 2031.92 ± 7.51 to
7918.75 ± 6.96 lg/100 g. H2, a mono floral honey from arid regions
had the highest level of Proline. Lys. results showed that the high-
est level was found in H2, a mono floral honey from arid regions
(1734.92 ± 14.54 lg/100 g). On the other hand, H8, a mono floral
honey from arid regions, showed the highest level of Tyr.
(272.29 ± 2.49 lg/100 g), while H2, another mono floral from arid
regions, had the highest values of Met. (188.84 ± 1.44 lg/100 g).
Val. was detected only in some honey samples, and H13 had the
highest value (102.00 ± 0.76 lg/100 g). LIc. showed the highest va-
lue in H8 mono floral from arid regions (149.67 ± 0.80 lg/100 g),
moreover, Leu. had the highest value in H2 mono floral from arid
regions (890.83 ± 14.81 lg/100 g). In general, total free amino
acids were higher in mono floral honey samples from arid regions.
H2 had the highest value (15.14 ± 0.02 mg/100 g). In addition, Sys.
and Phe. were not detected in all honey samples under investiga-
tion. To the best of our knowledge, this is the first time free amino
acids have ever been analysed in honey.
4. Conclusion
Since characterisation of honey from arid regions is not yet
available, this study provides a preliminary but comprehensive
and detailed evaluation of physicochemical and biochemical prop-
erties composition of some of these honeys. In conclusion, our ap-
proach, using all the parameters previously mentioned, provides
an entrance to further research with a larger number of samples
in order to improve the interest and knowledge about this honey
and its quality profiles.
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