MANIPAL INTERNATIONAL UNIVERSITY

ANALYZING THE COMPONENT OF RICE BRAN & RICE

STRAW TO PRODUCE THE ANIMAL FEEDS
Present by : AMIN RAHMAN BIN SABEER MOHAMED
Matrix no : 1000113
Industrial Supervisor : EN. MOHD FIRKHRY FADHLY BIN ABDUL
GHANI
Academic Supervisor : PROF. DR. AROKIARAJ PAPPUSAMY
Department : INDUSTRIAL BIOTECHNOLOGY RESEARCH CENTRE
(IBRC)

COMPANY PROFILE

Sirim Berhad
Standards and Industrial Research Institute of Malaysia
(SIRIM).
A corporate organization wholly owned by the Malaysian
Government.
Main headquarters Shah Alam, Selangor

OBJECTIVES
The objectives of SIRIM are:
1)

To innovate and develop processes, product

and technologies for industry.

2)

To promote standardization and quality.

3)

To provide technical services for industry and

the public.

INTRODUCTION
Rice Bran

A by product of rice milling industry.

Research by Mohammed et al. (2014) revealed that

rice bran is an incredible source of the:
Vitamins
Minerals
Amino acids
Essential fatty acids
Dietary fibre and
more than 100 antioxidant nutrients
that helps to fight against disease and promote good
health

CONT
Rice Straw
The vegetative part of the rice plant.
Cut at grain harvest or after.
It may be burned and left on the field before the next
ploughing.
Good source of energy, but is poor in protein (Heuze & Tran,
2013).
It has low digestibility.
Low in energy with a crude protein content of only 3
5 percent.

OBJECTIVE

-To analyze the content of rice bran

and rice straw.
-To gain the working experience.

TASKS
Moisture

Content test
Carbohydrate test
Lignin test
Cellulose test
Enzyme Activity
Enzyme Kinetic

METHODOLOGY
Moisture Content test- Oven- Dry method.
Carbohydrate test- Phenol Sulphuric Acid method.
Lignin test- Tappi 222 test method.

Cellulose test- Tappi 203 test method.

Enzyme activity- FP Assay, CMC Assay, Glucose
Enzyme Kinetic- Method of CMC assay (different
concentration of CMC solution)

Moisture Content test

1. The Petri dish was weighed before place the
sample.

2. 1 gram of sample was added and incubated at

90C for 16 hours.

3. The sample was weighed and minus the weight

of original Petri dish.

PHENOL SULPHURIC ACID METHOD (TOTAL

CARBOHYDRATE)
100 mg of sample weighed into boiling tube.
The boiling tube was kept in boiling water bath for 3
hours with 5 mL of 2.5 NHCL and cooled to room
temperature.
Solid sodium carbonate was added to neutralized the
mixture.
The volume was made up to 100 mL and
centrifuged.
0.2, 0.4, 0.6, 0.8 and 1 mL of working standard were
pipette out into a series of test tube.
0.1 and 0.2 mL of the sample solution were pipette
out in two separate test tubes. Made up the volume
in each tube to 1 mL with distilled water.

CONT
The blank was set with 1 mL of distilled water.
1 mL of phenol solution was added to each tube.
5 mL of 96% sulphuric acid was added to each tube
and shake well.
After 10 minutes the content in the test tubes were
shake and placed in a water bath at 25- 30 C for 20
min.
The color was read at 490 nm.

LIGNIN TEST
1 gram of
sample was
weighed

And dried in an
oven for 2 days at
90C

The sample placed

in vesicator for 1 day

Placed in a
beaker and 15
ml of 72% of
sulfuric acid
was added

The filtrate
washed using
hot water (3
times)

The dried sample

weighed.

The sample
was stirred in
acid until
fully
dissolved.

The solution
was filtered

Transfered into
a schott bottle
and distilled
water was
added until the
total volume
reached 575 ml.

The solution was

autoclaved and
allowed it to
cool

ALPHA- CELLULOSE
25 mL of the filtrate and 10.0 mL of 0.5N
potassium dichromate solution were pipette into
a 250-mL flask.
Add cautiously, while swirling the flask, 50 mL of
concentrated HSO.
Allowed the solution to remain hot for 15 min,
then add 50 mL of water and cool to room
temperature.
2 to 4 drops of Ferroin indicator were added and
titrated with 0.1N ferrous ammonium sulfate
solution to a purple color.
Make a blank titration substituting the pulp
filtrate with 12.5 mL of 17.5% NaOH and 12.5
mL of water.

BETA- AND GAMMA- CELLULOSE

50 mL of the pulp filtrate was pipette into a 100-mL graduated

cylinder having a ground glass stopper.
50 mL of 3N HSO was added and mix thoroughly by inverting.
The cylinder was heat submerged in a hot water bath at about
70-90C for a few minutes to coagulate the beta-cellulose.
Allowed the precipitate to settle for several hours, preferably
overnight, then decant or filter, if necessary, to obtain a clear
solution.
50 mL of the clear solution and 10.0 mL of 0.5N KCrO was
pipette into a 300-mL flask.
90 mL of concentrated H2SO4 was cautiously added.
The solution was allowed to remain hot for 15 min, then proceed
with titration as outlined in alpha cellulose method.
Made a blank titration substituting the solution with 12.5 mL of
17.5% NaOH, 12.5 mL of water and 25 mL of 3N HSO.

ENZYME ACTIVITY TEST FOR CMC-ASE

Stock dilute
solution

1 ml dilute
sample was
added in test
tube

Add 1 ml of
citrate buffer pH
4.8 and add 1 ml
of CMC

Incubate at 50C
for 1 hour

Add 3 ml of
DNS

Incubate at
boiling
temperature for
15 minutes

Add 1 ml of
40% potassium
tartarate

Add 20 ml of
Distilled Water

Cool down the

sample in room
temperature

Absorbance at 540 nm using

spectrophotometer.

Enzyme activity test for FP- ase method

1.

2.

3.
4.
5.

6.
7.

0.5 mL of diluted enzyme solution was taken into the

test tube.
1 mL 0.1 mM citrate buffer pH 4.8 and 50 mg (1x6
cm strip) of Whatman No. 1 filter paper that has
been curled round a glass rod was added.
Test tube was incubated for 1 hour at 50 C.
After incubation, 3 mL DNS reagent was added.
Then boiled for 15 minutes and 1 mL of 40% sodium
potassium tartarate was added.
20 mL of distilled water was added.
After cooling to room temperature, absorbance was
measured at 550 nm.

ENZYME ACTIVITY TEST FOR GLUCOSE

METHOD
10 mM glucose solution were taken and diluted
to final volume of 1 mL.
In 1 mL standard solution, 2 mL 0.l M citrate
buffer, pH 4.8 was added.
3 mL DNS reagent was added and boiled for 15
minutes.
After boiled, 1 mL 40% sodium tartarate was
added in hot tubes.
20 mL of distilled water was added.
The tubes were cooled at room temperature and
the absorbance was measured at 550 nm.

ENZYME KINETIC

Stock dilute
solution

1 ml dilute
sample was
added in test
tube

1 ml of citrate
buffer pH 4.8
and 1 ml of
different
concentration of
CMC was added.

Incubated at
50C for 1 hour

3 ml of DNS
was added.

Incubated at
boiling
temperature for
15 minutes

1 ml of 40%
potassium
tartarate was
added.

20 ml of distilled
water was added.

The sample was

cooled at room
temperature

Absorbance at 540 nm using

spectrophotometer.

RESULTS

MOISTURE CONTENT TEST

Rice Bran

Rice Straw

Weight of Petri Dish

94.39 gram

93.62 gram

Sample

1 gram

1 gram

Weight after incubation

95.25 gram

94.52 gram

Difference of weight

0.84 gram

0.90 gram

Moisture %

14%

10%

Moisture% = Weight of original sample weight of dried sample x 100

weight of original sample

RESULT OF LIGNIN TEST

Sample

Weigh of
sample

Weight of
Petri Dish
before
incubation

Weight of
Petri Dish
after
incubation

Differences
in Weight
(%)

Rice Bran

1 gram

2.69

3.78

0.09 gram
(9%)

Rice Straw

1 gram

2.74

3.70

0.08 gram
(8%)

ALPHA, BETA AND GAMMA CELLULOSE

Rice Bran
Rice Straw

- cellulose

- cellulose

- cellulose

62.34877666
59.28866667

26.26777657
25.21842677

11.38344693
9.49290666

GLUCOSE IN ENZYME ACTIVITY TEST

Abs (550nm)
y = 0.5045x - 0.6154
R = 0.9738

2.5

2.4862

Absorbance

1.9699

1.5

Abs (550nm)

1.2463
1

Linear (Abs (550nm))

0.9263

0.5

0
0
-0.5

0.1828

0.0909
1

[Glucose], mg/ml

[CMC HTec],
mg/ml
Blank
1
2
3
4
5

[CMC CTec],
mg/ml
Blank
1
2
3
4
5

Abs (540nm)
0.4262
0.8128
0.9263
1.2463
1.9699
2.48862

Abs (540nm)

0.3574
1.5441
1.8277
2.1311
2.7854
3.7484

Standard Factor CMC-ase Activity

1.23
1.641856
2.02
1.871126
2.41
2.517526
2.02
3.979198
2.02
5.0270124

Standard Factor CMC-ase Activity

0.65
2.177181
1.04
2.577057
1.41
3.004851
1.41
3.927414
1.33
5.285244

[Filter Paper Assay HTec], mg/ml

Blank
1
2
3
4
5

[Filter Paper Assay CTec], mg/ml

Blank
1
2
3
4
5

Abs (550nm)
0.07
0.9722
1.7059
2.0465
2.438
3.5793
Abs (550nm)

0.0747
0.8794
1.4484
1.5541
1.9328
2.673

[Enzyme Kinetics HTec], mg/ml

Blank

Abs (540nm)
0.0875

0.8
1
1.2
1.4
1.6
1.8
2

0.4899
0.6062
0.8879
2.1804
2.6088
3.1877
4.461

[Enzyme Kinetics CTec], mg/ml

Blank

Abs (540nm)
0.0904

0.8
1
1.2
1.4
1.6
1.8
2

0.3191
0.9931
1.5585
1.7958
2.3257
2.9981
4.3732

CONCLUSION

Diet of rice straw alone is not sufficient even to

maintain the animals weight unless supplementary
protein is provided. (Food and Agriculture
Organization)
All of the nutrients in rice bran were significantly
effective and can be useful in food application as
edible oil extraction and food supplement
(Mohammed et al. 2014).

Learn about the process and method of analysis the

chemical content in sample.
Know how to use equipment in proper way.