LWT - Food Science and Technology 50 (2013) 17e24

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LWT - Food Science and Technology

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Optimization of microencapsulation of probiotics in raspberry juice by spray

drying
Kartheek Anekella a, Valérie Orsat b, *
a
Department of Food, Bioprocessing and Nutrition Sciences, Schaub Hall, North Carolina State University, Raleigh, NC 27695, USA
b
Bioresource Engineering Department, McGill University, 21111 Lakeshore Rd., Ste-Anne de Bellevue, QC, Canada H9X 3V9

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

Article history: Aims: Probiotics were microencapsulated in raspberry juice through spray drying.
Received 27 April 2012 Methods & Results: A combination of probiotics (Lactobacillus acidophilus NRRL B-4495 and Lactobacillus
Received in revised form rhamnosus NRRL B-442) was chosen to offer high viability. Maltodextrin’s role as a carbon source was
31 July 2012
also assessed for its prebiotic potential. Spray drying inlet temperature (
C), total solids: maltodextrin
Accepted 2 August 2012
ratio, and inlet feed rate (mL/min) were fixed as independent variables while % recovery, % survival and
color were the dependent outputs.
Keywords:
Conclusions & Significance: High temperatures during spray drying are detrimental to probiotics and can
Sub-lethal thermal effect
Probiotic/prebiotic
be circumvented by sub-lethal thermal shock (50
C for L. acidophilus and 52.5
C for L. rhamnosus).
Lactobacillus Increasing the microencapsulating material concentration increased the survival rate of the probiotics.
Non-dairy probiotic foods are becoming popular as they do not pose problems of lactose intolerance
while they offer an alternative.
Ó 2012 Elsevier Ltd. All rights reserved.

1. Introduction without losing its basic properties of providing nutrition and

Probiotics are considered as “good buddies” to human health.
Although historically, probiotics were products of the pharmaceu-
tical industry, the current trend is moving toward the health food
sector, making Hippocrates’ statement “Let food be your medicine”
true once again.
Probiotics are usually sold as capsules, powders and combina-
tions of different species which may have multiple advantages
(Timmerman, Koning, Mulder, Rombouts, & Beynen, 2004). It was
proven in various animal studies that the use of one or more
strains/species of probiotics can have beneficial effect and it is
logical to assume that a mosaic of probiotics could help in exerting
multiple benefits they possess (Famularo, de Simone, Matteuzzi,
& Pirovano, 1999; Sanders, 1999). In many cases however, the
functionality of probiotics is an issue due to the poor quality in the
standards of preparation of probiotics foods and lack of sound and
thorough clinical evidence (Azcarate-Peril, Tallon, & Klaenhammer,
2009; Hamilton-Miller & Shah, 2002; Klaenhammer, 2000;
Timmerman et al., 2004). The main goal of any food industry is to
increase the versatility in consumption of different forms of food
* Corresponding author.
E-mail addresses: kanekel@ncsu.edu (K. Anekella), valerie.orsat@mcgill.ca
(V. Orsat).
ensuring health.
Formulating and enriching foods with probiotics would not
only improve public health but also the diversity in food choices.
Most probiotic foods in the current market are refrigerated dairy-
based products (Burgain, Gaiani, Linder, & Scher, 2011) while
preparations of non-dairy foods will attract a broader range of
consumers with different preferences. The food matrix encapsu-
lation must act as a buffer during storage as well as in the
stomach transit until the probiotic is delivered to the intestinal
tract along with offering a protection during thermal processing
(Ranadheera, Baines, & Adams, 2010). The probiotic microorgan-
isms present in food should survive in significant numbers of
at least 106e108 CFU/g, although the numbers vary from strain
to strain (Boylston, Vinderola, Ghoddusi, & Reinheimer, 2004;
Chávez & Ledeboer, 2007; Ishibashi & Shimamura, 1993;
Kailasapathy & Rybka, 1997). It is possible to induce a sub-lethal
effect on microorganisms which adapts them to adverse condi-
tions during drying, storage and other processes (Broadbent & Lin,
1999). Usually a temperature rise of 10
C above the optimal
growth temperature leads to shock (Teixeira, Castro, & Kirby,
1994). Thermal sub-lethal treatment can increase the survival
rate of Lactobacilli remarkably (between 16 and 18 folds
depending on the adaptation media) during and following spray
drying (Desmond, Stanton, Fitzgerald, Collins, & Paul Ross, 2001;
Gardiner et al., 2002). The stress resistance proteins are produced

0023-6438/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.lwt.2012.08.003
18 K. Anekella, V. Orsat / LWT - Food Science and Technology 50 (2013) 17e24
mainly during the sub-lethal exposure prior to drying (Dabbah,
Moats, & Mattick, 1969; Teixeira et al., 1994).
Although spray drying of Lactobacillus cultures was first done in
1914 by Rogers, it was not adopted due to very low survival rate,
difficulty in storage as well as poor rehydration capacity (Porubcan
& Sellers, 1975; Teixeira, Castro, Malcata, & Kirby, 1995). Use of
berries with probiotics has been tested for uropathogenic and
urogenital disorders. The proanthocyanins present in berries, such
as cranberries, can modulate the immune system in conjunction
with probiotics (Reid, 2002). Anthocyanin rich raspberries also
have a high amount of dietary fibers (6.5 g/100 g) with good
absorption characteristics which could potentially serve as a carrier
and microencapsulating agent as well as a prebiotic (Chiou &
Langrish, 2007). This supports our premise of studying the
combination of probiotics organisms with raspberry juice for spray
drying. Additionally, thermoplasticity and hygroscopicity of fruit
juice might pose problems during the spray drying causing them to
adhere to the chamber wall due to their stickiness, clogging and
caking (Chegini & Ghobadian, 2005). Adding suitable wall materials
such as maltodextrin can reduce the caking and stickiness to the
walls and increase the free flowing nature of the spray dried
powder.
2. Materials and methods
Following a review of the literature on the health benefits of
probiotics, two species widely used in commercial probiotic foods
were chosen. Lactobacillus rhamnosus NRRL B-4495 and Lactoba-
cillus acidophilus NRRL B-442 were obtained from the USDA’s
Agricultural Research Services Culture Collection. Dried pellets
were reconstituted in 50 mL MRS broth (deMan, Rogosa and Sharpe
medium, Difco) and grown overnight (14e16 h) in an incubator
shaker (Model G24, New Brunswick Scientific, USA) at 110 rpm and
37
C. This was subcultured and grown overnight again. The culture
thus obtained after the second sub-culture was used for further
experiments. A small part of the culture was stored in sterile 30mL/
100 mL of glycerol at 80
C (86C ULT Freezer Model 5698,
Thermo Forma, USA) for later usage. A similar recipe was adapted to
prepare MD-MRS medium where the dextrose was replaced by
maltodextrin (MD, referred as MD-MRS) with Dextrose Equivalent
value 5e8 (Oxoid, USA).
2.1. Growth curve and dry biomass estimation
The increase in biomass over the growth cycle period provides
information on the kinetics of the growth in relation to substrate
utilization. A 5 mL sample of this culture was taken every 3 h and
centrifuged in a pre-weighed centrifuge at 1130 g for 6e7 min.
The supernatant was decanted and stored for substrate estima-
tion. The weight of dry biomass was measured as the weight
difference between the tube and the pellet after drying at 103  2
C
in hot air oven (Thermoelectron, USA) for over 12 h (Eq. (1)).
K ¼ ½logðXt1 Þ  logðXt2 Þ= 0:301*ðt2  t1Þ
K ¼ number of populations doubling in an hour (growth rate
constant)
Xt1 ¼ number of cells/mL at time t1; Xt2 ¼ number of cells/mL at
time t2
During any phase, the doubling time (Td) or generation time is
the time required for the cell biomass to double in number as
represented in Eq. (2).
Td ¼ 1=K
2.2. Determination of substrate concentration (anthrone method)
Understanding the substrate utilization pattern of a microor-
ganism is also helpful to assess the growth behavior and survival in
the gut which is the final destination (Verdenelli et al., 2009). A
procedure similar to the one described by Sadasivam and
Manickam (2005) was followed. Anthrone reagent (Sigma
Aldrich, USA) was prepared by dissolving 200 mg of anthrone in
100 mL of 95 g/100 g sulfuric acid. A 1 mL volume of the super-
natant obtained from the biomass estimation was diluted 10 times
and 4 mL anthrone reagent was added. The mixture was heated in
a boiling water bath (100  2
C) for 6e7 min until a stable green
color was obtained after which the mixture was cooled immedi-
ately in an ice bath for 2 min. The maximum absorbance at OD620
was recorded. The concentration of pure dextrose/maltodextrin
was estimated from the measured OD620 using the standard curve
obtained using a known concentration (0.1 g/L) of dextrose/mal-
todextrin. OD620 was plotted on the X-axis and concentration of
substrate on the Y-axis.
2.3. Raspberry juice
The extracted raspberry pulp had a solid content of 13e14

Brix measured by a portable refractometer (Fischer Scientific,
USA). The seeds and skin were sieved out using a fine metal sieve
filter. The
Brix unit was adjusted to 11 (total solid concentration
0.1 g/L) as otherwise the pure extract was too viscous to be spray
dried.
2.4. Sub-lethal temperature (Tsl) treatment
A sub-lethal temperature treatment was selected to subject the
microbes to thermal stress before spray drying. Since the sub-
lethal temperature of any given microorganism is strain-specific,
each strain was tested individually. Five mL of late log phase
Lactobacillus cultures (incubated at 37
C, on a rotary shaker at
110 rpm for 12e14 h) was subjected to different sub-lethal
temperatures (45
C, 50
C, 52
C, 55
C) in a hot water bath.
Test tubes containing either MRS or raspberry juice (with inserted
thermocouples) were used to regulate the temperature. One mL of
sample was collected every 5 min and transferred into 9 ml of
sterile MRD (maximum recovery diluents; 1 g/L- peptone, 8.5 g/L
NaCl) up to 15 min. CFU analysis was performed from each of these
samples at regular intervals (0, 5, 10, 15 min). Tsl was determined
from the curves in the graph where the temperature was retaining
the highest number of cells after 15 min of thermal stress expo-
sure. Prepared graphs represent the average of triplicate Petri
plates for each of three trials.
2.5. Spray drying
Spray drying of raspberry juice with maltodextrin as an additive
(wall material) at different ratios and mixture of lactobacilli was
(1) performed using a Buchi B-290 mini spray dryer. A five mL volume
of each probiotic cultures (at late exponential phase, grown over-
night) were subjected to sub-lethal treatment and centrifuged at
1130 g for 1e2 min in order to extract the pure cells. These pellets
were diluted in 1 mL of sterile distilled water each and mixed with
50 mL of raspberry pulp and maltodextrin thoroughly by magnetic
stirring just before spray drying. Total number of cells just before
spray drying was approximately 9.5 log CFU/mL. The spray dryer
was allowed to reach uniform process temperature for 15e20 min
prior to the spray drying. The aspiration was maintained at 100%
and cyclone air flow rate at 30 m3/h. The temperatures used to
(2) optimize the model were 100
C, 115
C and 130
C. Feed rates used
 K. Anekella, V. Orsat / LWT - Food Science and Technology 50 (2013) 17e24 19

Table 1
Spray drying responses (generated by JMP-SAS) with the outputs for each dependent variables.

Response Pattern Inlet temp Maltodextrin Inlet feed rate Outlet % recovery % survival Color (DE)
(
C) ratio (mL/min) temperaturea (
C)
R1 0,0,0 115 1.5 50 82e86 25.4 71.14 54.252
R2 0,0,0 115 1.5 50 80e85 32.9 84.44 54.094
R3 0,0,1 115 1.5 40 81e86 36 68.24 52.766
R4 þ1,1,1 130 1 40 92e97 55 58.80 56.719
R5 1,þ1,þ1 100 2 60 69e74 25.6 78.97 52.187
R6 0,0,0 115 1.5 50 80e85 35.2 69.10 52.149
R7 1,1,þ1 100 1 60 67e72 28 80.26 56.623
R8 0,þ1,0 115 2 50 81e85 31.1 64.38 52.689
R9 0,0,0 115 1.5 50 81e86 35.2 71.46 52.062
R10 1,þ1,1 100 2 40 71e76 32.1 82.62 55.443
R11 1,1,1 100 1 40 68e74 47.1 84.33 56.394
R12 1,0,0 100 1.5 50 71e76 30.75 79.08 53.313
R13 þ1,0,0 130 1.5 50 91e95 36.8 53.65 53.413
R14 0,0,0 115 1.5 50 83e88 27.6 67.70 54.337
R15 þ1,þ1,1 130 2 40 91e96 38 53.65 51.558
R16 0,1,0 115 1 50 79e83 41.5 69.74 57.082
R17 þ1,þ1,þ1 130 2 60 88e92 32.5 68.03 53.238
R18 þ1,1,þ1 130 1 50 90e94 35.5 56.87 58.018
R19 0,0,þ1 115 1.5 60 77e83 24.65 70.39 53.165
R20 0,0,0 115 1.5 50 80e85 27.6 67.70 54.337
a
Outlet Temp is neither of the variables. It was only included in the table to compare how it changed with respect to the other variables.

were 10, 15, 20 mL/min and the total solids: maltodextrin ratios
(referred as maltodextrin ratio from here on) used were 1:1, 1:1.5
and 1:2.
Following spray drying, the raspberry powder thus obtained
was stored in small glass jars with screw caps to evaluate their end-
product color.
2.6. Color
Color of the end products was measured by Chromameter
(Model- CR300, Konica-Minolta, USA). The measurement was done
in small glass tube filled with the spray dried powder and recon-
stituted liquid, fully covering the base diameter of the instrument.
From the obtained L*, a* and b* values, the total change in color (DE)
was calculated using Eq. (3).
qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
DE ¼ ðLo  L*Þ2 þðao  a*Þ2 þðbo  b*Þ2
Where the measurements made with the subscript o are made with
the reference white calibration. Thus the most desirable bright red
color of our raspberry spray dried powder would be indicated in
terms of the largest DE color difference.
3. Results
3.1. Growth curves
The organisms’ growth curves and their characteristics are
presented in Fig. 1 and Table 2 for L. acidophilus and in Fig. 2 and
Table 3 for L. rhamnosus. The relation between OD600 and
log CFU/mL for L. acidophilus in MRS is given by the empirically
derived equation y ¼ 0.74x þ 7.81. The relation between OD600 and
log CFU/mL for L. rhamnosus is given by the empirically derived
equation y ¼ 1.91x þ 7.23. These relations were used to estimate the
concentration of live cells in MD-MRS from OD600 values obtained
under the same growth conditions.
Although the growth was not very strong with maltodextrin
(MD-MRS), as with dextrose (MRS), maltodextrin could still
potentially act as a prebiotic as well as a microencapsulating agent
during spray drying. From the data in Tables and Figs. 1 and 2, it is
(3) concluded that dextrose was utilized better as a carbon source than
maltodextrin at similar concentrations. Under similar culture
conditions for the two growth media, the residual concentration of
dextrose was 6.8 g/L for L. acidophilus and 5.5 g/L for L. rhamnosus
(Figs. 3 and 4). The residual maltodextrin was 11.1 g/L for
L. acidophilus and 13.8 g/L for L. rhamnosus (Figs. 3 and 4).

2.7. Response surface design

Process optimization for the spray drying of probiotics in rasp-

berry juice was performed using response surface methodology
(RSM). Initial trials were performed by varying the different
conditions for the spray drying of probiotics in raspberry juice such
as inlet temperature, maltodextrin ratio, and the inlet feed rate as
found in the literature (preliminary results not shown). Inlet
temperature (
C), maltodextrin ratio, feed rate (mL/min) were the
independent variables and % recovery, % survival and color (DE)
were the dependent variables. After analyzing the effects of
different variables on the probiotics viability, powder color and
powder recovery, the maximum and minimum values of the each
factor were adjusted for heat shock treated probiotic mixture in the
raspberry juice. A central composite design- uniform precision
model by JMP-8 (SAS Institute, NC, USA) generated the 20 Fig. 1. OD600 and log CFU of L. acidophilus over time in MRS and MD-MRS (37
C,
responses shown in Table 1. 110 rpm).
20 K. Anekella, V. Orsat / LWT - Food Science and Technology 50 (2013) 17e24

Table 2 Table 3
Growth curve characteristics of L. acidophilus in MRS and MD-MRS. Growth curve characteristics of L. rhamnosus in MRS and MD-MRS.

MRS MD-MRS MRS MD-MRS

Growth Curve Exponential phase Exponential phase till Growth Curve Lag phase until 5 h and Lag phase until 5th h
Features 0e13th h followed 6th h followed by Features exponential till 16th h and followed by steady
by stationary phase stationary phase stationary phase from 17th h stationary phase
Increase in log 1.75 0.84 Increase in log 3.16 0.69
CFU/mL CFU/mL
Kmax 0.697 0.313 Kmax 1.65 0.202
Td 1.43 min 3.2 min Td 0.6 min 4.95 min

3.2. Sub-lethal temperature treatment
Exposing the probiotics to sub-lethal thermal shock increases
the subsequent tolerance to near lethal thermal stresses. In the
current study, two different heating media were used in the
assessment: MRS broth and raspberry juice on the two species
separately. A fruit juice was selected, in this case raspberry juice,
because it obviates the extra steps of centrifugation and purifica-
tion of the cultures before spray drying which was done in the case
of MRS. Spray drying trials with fruit juice with probiotic cells
without heat shock treatment had lower viability than heat shock
treated cells under similar conditions.
Figs. 5 and 6 present the organisms’ survival during sub-lethal
treatment. It can be seen that the viability, in MRS broth, of the
probiotics was maintained up to 50
C for L. acidophilus and 52.5
C
for L. rhamnosus for̴12 min and hence these temperatures have
been considered as their sub-lethal temperatures. Raspberry was
not as effective as MRS in survival of L. acidophilus as the cell count
decreased to zero within 5 min at 45
C and L. rhamnosus at 50
C.
The heating medium in which the cells are being adapted has
a significant effect on the survival, where a complex media offers
a better protection than a simple medium (here raspberry pulp)
(Corcoran, Ross, Fitzgerald, & Stanton, 2004; Desmond et al., 2001;
O’Riordan, Andrews, Buckle, & Conway, 2001; Teixeira, Castro, &
Kirby, 1995).
a maltodextrin ratio of 1:1 and an inlet feed rate of 40 mL/min. The
R2 value of the master model was 0.86 whereas the predictive
model had an R2 value of 0.71. The optimized coded equation (1,1)
of the predictive model is presented in equation (4).
% recovery ¼ 33:331  3:593*maltodextrin ratio  7:236*feed
rate þ 4:63*maltodextrin ratio*feed rate
(4)
3.4. % survival
The response plot for the % survival (in terms of log CFU/mL) for
the combined probiotics is presented in Fig. 8. As expected, the
processing temperature had a major effect on the probiotics’
survival. The survival dropped down to almost 55% (9.5e
5 log CFU/mL) when the inlet temperature was raised to 130
C.
The grid on top indicates the optimized conditions with respect to
maximum % survival (81.17%) at the input conditions of 100
C inlet
temperature, a maltodextrin ratio of 1:1 and an inlet feed rate of
40 mL/min.
The R2 value of the master model was 0.96 whereas the opti-
mized predictive model had an R2 value of 0.91. The optimized
coded (1, 1) equation of the predictive model for the probiotic
survival is presented in equation (5).

3.3. Spray drying process optimization using RSM % survival ¼ 67:933  8:98*inlet temp
 2:681*maltodextrin ratio þ 3:124*feed rate
The spray dried powder recovery varied between 25% and 55%
depending on the maltodextrin ratio and feed rate. The response þ 4:863*inlet temp*maltodextrin ratio
plot for % recovery with respect to total solids: maltodextrin ratio þ 5:840*maltodextrin ratio*feed rate
and feed flow rate is presented in Fig. 7. The grid on the top indi-
(5)
cates the overall optimum condition with respect to maximum
% recovery (48.79%) at input conditions of 100
C inlet temperature,

Fig. 2. OD600nm and Log CFU of L. rhamnosus over time in MRS and MD-MRS (37
C,
110 rpm). Fig. 3. Changes in dry biomass and substrate over time for L. acidophilus.
 K. Anekella, V. Orsat / LWT - Food Science and Technology 50 (2013) 17e24 21

color difference (DE 57.210) at conditions of 100
C inlet temperature,

maltodextrin ratio of 1:1 and inlet feed rate of 40 mL/min. The R2
value of the master model was 0.91 whereas the optimized predic-
tive model had an R2 value of 0.87. The optimized predictive model
(1, 1 coded values) for powder color is presented in equation (6).

Color ¼ 53:283  0:005*inlet temp

 2:068*maltodextrin ratio þ 0:114*feed rate
 0:689*inlet temp*maltodextrin ratio
þ 0:849*inlet temp*feed rate
þ 1:808*maltodextrin ratio*maltodextrin ratio (6)

Overall optimized conditions were determined at 100
C inlet

temperature, maltodextrin ratio of 1:1 and feed flow rate of
40 mL/min, where the maximum output dependent variables were
obtained with 48.79% recovery, 87.17% survival and 57.21 color
Fig. 4. Changes in dry biomass and substrate over time for L. rhamnosus.
change (DE). The overall desirability of the model was 91.15%.

3.5. Color 4. Discussions

The grid on top of the response plot for color, as presented in In the substrate utilization analysis (Figs. 3 and 4) it was made
Fig. 9, indicates the optimized conditions with respect to maximum evident that dextrose is better metabolized by the bacteria than

Fig. 5. Sub-lethal temperature-time assessment of L. acidophilus.

Fig. 6. Sub-lethal temperature-time assessment of L. rhamnosus.

22 K. Anekella, V. Orsat / LWT - Food Science and Technology 50 (2013) 17e24
Fig. 7. Response plot of % recovery with respect to maltodextrin ratio and feed rate
(mL/min).
maltodextrin. However the dry biomass at the end of the growth
cycle of L. acidophilus in MD-MRS (3.6 g/L) was higher than that of
MRS (2.4 g/L). Indeed, after the centrifugation and before drying,
there was a certain amount of insoluble maltodextrin present as
a residue in the pellet as a white mass (which was not seen in MRS)
in L. acidophilus cultures (hence the dry biomass curve of MD-MRS
in Fig. 3 did not start from 0). So the true weight might have been
influenced by the presence of insoluble maltodextrin.
Although the organism growth was not very strong with mal-
todextrin it could potentially act as a prebiotic as well as a micro-
encapsulating agent during spray drying. MRS acted as a better
heating medium, for thermal shock pre-treatments, than raspberry
because of the more complex nutrients present in MRS medium
than the plain raspberry juice which has mostly sugars and fibers.
It is assumed that the proteins present in the complex media may
contribute to the stability of the intracellular proteins of the
Lactobacilli during thermal shock. The sub-lethal stress induces
protective mechanisms such as an alteration or reprogramming of
the metabolic pathways to adjust to the new environment
(Teixeira et al., 1994), thus increasing their survival during subse-
quent harsh treatment and improved viability during storage
(O’Riordan et al., 2001; Selmer-Olsen, Birkeland, & Sørhaug, 1999;
Shah & Ravula, 2000).
Fig. 8. Response plot for % survival with inlet temperature (
C) and feed rate (mL/min).
Fig. 9. Response plot of powder color with maltodextrin ratio and inlet temperature
(
C) as independent variables.
Previous studies indicate that cells in the exponential growth
phase are more easily adapted than in the stationary phase proving
that age of cells also has a pronounced effect on the induction of
thermo-tolerance (Corcoran et al., 2004; Corcoran, Ross, Fitzgerald,
Dockery, & Stanton, 2006). Therefore the probiotic cultures experi-
ence both active growth and carbon depletion during the same phase
of the growth cycle making them more resistant subsequently.
The walls of the spray drying chamber was layered completely
with the raspberry solids by the end of spray drying due to the
stickiness of the solutions which lead to major losses in product
recovery. An interesting observation from the obtained data
(Table 1) is that the temperature had a minimal role in powder
recovery. From the response plot (Fig. 7), maltodextrin ratio and
feed rate had significant effects on % recovery of produced powder
from raspberry juice. As the maltodextrin ratio increased to 2, the
powder recovery dropped to 33%. Reducing the maltodextrin ratio
can increase the powder recovery but it will affect the cell survival
of the probiotic due to reduced encapsulation efficiency. Similarly
when the feed rate increased upto 60 mL/min the recovery dropped
to as low as 25%. Faster feed rate does not give enough time for
complete drying of the pulp thus reducing the %recovery.
On the other hand, outlet temperatures greater than 85
-90
C
are lethal for probiotics (Corcoran et al., 2004; Gardiner et al., 2000;
Zamora, Carretero, & Parés, 2006) but a sub-lethal temperature
pretreatment enabled cells to survive in that range with cell death
occurring only after 92
C outlet temperature in our current study.
It is seen in Table 1 that under same inlet temperature conditions,
a higher inlet feed rate had a lower outlet temperature and an
increased survival rate. This indicates that the cell survival is mostly
dependent on outlet temperature as expected from previous
studies (Boza, Barbin, & Scamparini, 2004). Cellular membrane heat
damage is one of the most susceptible target damage during spray
drying. High temperatures during spray drying cause the cellular
pores to leak the intracellular substances (Corcoran et al., 2004;
Gardiner et al., 2000). The molecular nature of heat damage (above
90
C) is not clearly known but denaturation and loss of metabolic
activity of critical proteins, DNA and ribosomes are few vital events
(Meng, Stanton, Fitzgerald, Daly, & Ross, 2008; Teixeira, Castro,
Mohácsi-Farkas, & Kirby, 1997). Heat shock proteins produced
during sub-lethal stress aid in protecting the probiotics during
subsequent stress.
Probiotics’ survival during spray drying can also attributed to
the strong adherence to the carrier, which protects cells from high
 K. Anekella, V. Orsat / LWT - Food Science and Technology 50 (2013) 17e24 23

Table 4
Color characteristics of the raspberry powder samples and their rehydrated liquid samples (1 g/9 mL of water) from all the responses.

Response Color characteristics of powder DE Color characteristics of rehydrated liquid DE

L a b L a b
R1 58.270  0.509 42.646  0.769 11.496  0.312 54.251  0.808 24.736  0.568 17.506  0.260 8.953  0.354 20.055  0.354
R2 59.268  0.195 43.108  0.482 11.758  0.196 54.094  0.446 25.360  0.392 18.095  0.333 9.465  0.325 20.718  0.436
R3 60.242  0.549 42.315  0.534 11.018  0.403 52.766  1.646 24.913  0.259 17.883  0.660 9.427  0.641 20.496  0.919
R4 57.410  0.127 44.795  0.474 12.855  0.021 56.719  0.444 24.890  0.099 19.490  0.552 10.045  0.247 22.268  0.612
R5 60.535  0.054 41.870  0.156 10.372  0.172 52.187  0.188 24.710  0.189 16.763  0.443 9.047  0.236 19.42  0.471
R6 59.850  0.786 41.288  2.132 10.913  0.709 52.149  2.270 24.493  0.238 17.970  0.206 9.438  0.230 20.698  0.23
R7 56.298  0.053 43.935  0.021 12.513  0.286 56.623  0.106 25.238  0.180 19.453  0.173 9.998  0.173 22.180  0.212
R8 60.454  0.101 42.599  0.087 10.981  0.217 52.689  0.281 24.821  0.762 17.163  1.425 9.259  0.900 19.876  1.537
R9 60.213  0.456 41.460  1.866 10.813  0.590 52.062  1.8 24.485  0.306 17.335  0.306 9.218  0.098 20.037  0.287
R10 58.199  0.225 43.976  0.394 11.996  0.164 55.443  0.475 23.463  0.293 14.674  0.281 7.735  0.304 17.262  0.314
R11 56.650  0.118 44.032  0.172 12.035  0.068 56.394  0.218 24.557  0.094 18.145  0.021 9.167  0.019 20.741  0.013
R12 59.612  0.276 42.608  0.483 10.783  0.165 53.313  0.575 23.788  0.205 16.712  0.016 8.687  0.099 19.381  0.017
R13 59.520  0.382 42.260  1.068 10.977  0.236 53.413  0.851 24.750  0.370 17.130  0.122 8.953  0.127 19.708  0.204
R14 58.840  0.255 43.120  0.320 11.707  0.075 54.337  0357 25.207  0.193 18.457  0.206 9.827  0.104 21.208  0.238
R15 61.310  0.297 41.615  0.714 10.785  0.021 51.558  0.747 25.125  0.163 15.750  0.990 8.675  0.276 18.297  0.986
R16 56.036  0.430 44.234  0.613 12.519  0.210 57.082  0.775 25.030  0.120 19.783  0.138 10.050  0.138 22.521  0.121
R17 60.053  0.100 42.753  0.514 11.093  0.229 53.238  0.506 24.063  0.240 16.040  0.115 8.480  0.173 18.642  0.203
R18 55.903  0.818 45.167  1.000 13.480  0.696 58.018  1.407 25.273  0.542 19.727  0.152 10.140  0.125 22.485  0.215
R19 59.413  0.141 42.332  0.394 10.545  0.191 53.165  0.268 24.212  0.021 16.775  0.016 8.783  0.094 19.4  0.024
R20 56.750  0.173 44.860  0.349 12.928  0.278 54.337  0.357 23.098  0.209 15.830  0.116 8.143  0.168 21.208  0.238

acidic and bile conditions (Crittenden et al., 2001). Overall, malto-
dextrin is confirmed to serve as a good encapsulating matrix as well
as a moderate prebiotic for high survival of probiotics (Cortés-
Arminio, López-Malo, & Palou, 2010). The sugars present in the
raspberry may also have contributed to the survival during drying
since sugars act as thermoprotectants during spray drying
(Carvalho et al., 2003).
The response plot of powder color (Fig. 8) shows that the mal-
todextrin ratio had a major effect on color (DE) as it reduced
(reaching toward white or increase in whiteness) with an
increasing concentration of maltodextrin. On the other hand, the
rehydrated liquid (Table 4), produced from the powder for all the
three MD ratios had similar bright red color. So, in the case where
consumption of the raspberry powder is via rehydration, then
maltodextrin ratio can be increased further as the rehydrated liquid
is more or less bright red in color irrespective of the MD concen-
tration (Table 4).
5. Conclusions
Dextrose was a better carbon source than maltodextrin in
support of the growth of the Lactobacilli (L. acidophilus and
L. rhamnosus) chosen in the current study. MRS medium acted as
a better heating medium than raspberry juice during the sub-lethal
heat shock pre-treatment. Microorganisms were able to withstand
up to 50
C (for L. acidophilus) and 52.5
C (for L. rhamnosus) in MRS
as heating medium whereas both the microorganisms were killed
at 45
C in raspberry juice as the heating medium.
Maltodextrin ratio and inlet feed rate have a major effect on
recovery and the optimization equation followed a linear two factor
model. Inlet temperature had a profound effect on % survival during
spray drying. Although in the two factor linear optimization
equation, the other independent variables inlet feed rate and
maltodextrin ratio and also their products were significant. Mal-
todextrin ratio had a major effect on color of the raspberry powder.
To the best of our knowledge there are no previous studies
which optimized the microencapsulation of probiotics in fruit
powder by spray drying. The overall desirability of the optimized
model was 91.15% which is quite high for a biological system on
which there is minimal process control. In conclusion, the choice of
the microencapsulating agent and its interaction between the
protein-carbohydrate-probiotics formulation should be studied
further to ensure minimal toxicity and better bioavailability. Health
benefits must be validated in the presence of food matrix dosage
rather than simply with isolated pure culture.
Acknowledgments
The authors would like to acknowledge the financial support of
the Natural Sciences and Engineering Research Council of Canada
(NSERC).
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