Vol 3 (1) 2011 - UBM :: Departamentul de Chimie-Biologie

M. Ayaz Javed, M. Nadeem, Development of probiotics ice cream in Pakistan from buffalo milk by using
B. bifidum and L. acidophilus
mL). All these treatments were compared
with a control T 0 which did not contain any
probiotic culture. Each treatment was
replicated 3 times. The mix was heated to
80°C, homogenized in a clean and sanitized
double stage homogenizer (APV) at 100
kg/cm 2 and 35 kg/cm 2 pressure in the first
and second stage respectively. Probiotic Ice
cream was manufactured in the pilot plant of
the Research and Development Department,
Unilever Walls Ice Cream Factory Lahore,
Pakistan. One gm freeze dried culture for L.
acidophilus contained = 8.4 x 10 9 cfu and
one gm freeze dried culture for B.bifidum
contained = 7.5 x 10 9 cfu. From these
guidelines the quantities of probiotics
cultures were calculated as 1000 mg= 10 9,
100 mg= 10 8 , 10 mg= 107 and 1mg= 10 6 .
The composition of basic formulation
(control) comprised 8% milk fat, 10%
MSNF, 10% sucrose, 4% liquid glucose,
0.25% sodium alginate and 0.25% glycerol
monostearate. The experiments involved
making of 10 types of ice cream i.e., T 1 to T 9
along with control T 0 .
2.4. Inoculation of probiotic culture in ice
cream mix
Ice cream mix was divided into ten
equal parts. Each part was inoculated by
probiotic culture according to the
experimental design except for control. Each
sample was mixed well and kept for
fermentation for two hours at 41°C. Mix was
cooled at 4°C; vanilla flavor was added in it
and aged for 24 hours followed by freezing
and hardening. It was packed manually in 1
liter plastic tubs and was first frozen at -5°C
at -18 °C.
2.5. Analysis
Ice cream samples were stored at -18
°C ± 2°C and were analyzed for physical,
chemical, microbiological and sensory
evaluation. Melting resistance, overrun and
viscosity was determined by using the
procedures as described by Upadhyay et al.
[8]. pH, acidiy, fat, protein, total solids
content was determined following the
respective procedures as prescribed in
AOAC [9]. Reinforced Clostridial Agar
(RCA) (Oxoid) was used as a media to
enumerate L. acidophilus and B. bfidum
(Ming, 1989) in probiotic ice cream samples.
RCA media was prepared by following the
instructions of manufacturer (Oxoid). Frozen
fermented ice cream was thawed and then
diluted in autoclaved 0.85% saline. With the
help of micropipette ,one-tenth milliliter of
each dilution was placed on the centre of
solidify RCA plates and spread evenly over
the surface by means of a sterilized bentglass-rod
The plates were then incubated in
an anaerobic environment (Anaerogas Pack
Himedia ref LE 002A-5NO) at 37ºC for 48
hours. The total numbers of L. acidophilus
and B. bifidum were determined based on
their colony morphology when grown on
RCA. L. acidophilus produced pinpointsized
colonies, but B. bifidum produced
large colonies.
2.6. Sensory evaluation
Ice cream samples prepared from all
the treatments of probiotic cultures together
with regular (control) were subjected to
sensory evaluation by trained taste panel of
five judges. Evaluation was done by using
nine (9) point hedonic scale on a sensory
evaluation Performa for following
parameters; color, thickness, flavor, taste
and overall acceptability. All the samples
were coded with three digit random number
and all orders of serving were completely
randomized [10].
2.7. Statistical analysis
The data was obtained by using
Completely Randomized Design (CRD) and
outcome of the analysis was analyzed
through two way analysis of variance under
factorial arrangement to find out the effect of
treatment and effect of storage time on
probiotics ice cream by following the
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