NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350

BIOCHEMISTRY PRACTICAL REPORT

PRACTICAL 1
(A) BLOOD GLUCOSE MEASUREMENT
Aim of practical :
To analayse blood glucose and investigate bodys ability to deal with
carbohydrate load (glucose tolerance test)
Methods :
1. The subject need to fast overnight if the test is to be carried out
in the morning or the subject is allowed to have a light breakfast or
no lunch if test is to be carried out in the afternoon.
2. Blood sample for determination of fasting blood sugar is taken
with washed hands.
3. Approximately 75 g glucose solution is drank immediately after
fasting sample has been taken. The time is noted.
4. Hands are washed well to remove any traces of glucose before
the fingertip is pricked with the microlance provided.
5. One drop of blood is applied to pad of glucose meter test strip at
following times thereafter :
15 minutes, 30 minutes, 45 minutes, 60 minutes, 90 minutes and
120 minutes
6. The readings for each test subject are noted at each time point.
7. A graph of glucose concentration against time is plotted.

NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350

Result :
Time
(mins)
Glucose
concentrati
on (mg/dL)

15

30

45

60

90

120

Table 1 shows : glucose concentration (mg/dL) recorded at each

time point (mins)
Graph of glucose concentration against time

Glucose concentration (mg/dL)

15.0

30.0

45.0

60.0

90.0

Time (mins)

Graph 1 : shows glucose concentration (mg/dL) recorded for each

time point (mins)
Conclusion :

120.0

NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350
SELF ASSESSMENT QUESTIONS
1. Would analysis of urine be helpful in glucose tolerance test?
Yes because presence of glucose in urine might indicate diabetes
mellitus.
2. How would a normal, moderately diabetic and severely diabetic
condition be differentiated by glucose tolerance test?
The highest blood glucose level in blood after glucose drinking
might indicate severely diabetic condition and for moderately
diabetic condition, the blood glucose level might be slightly lower
than that . For normal condition, the blood glucose level should be
the lowest one.

3. The hormone insulin, secreted by the pancreas, increased rate of

uptake of glucose by the tissues. Hormones such as growth
hormone of anterior pituitary, epinephrine from the adrenal medulla,
glucocorticoids of adrenal cortex, glucagon from the pancreas and
thyroid hormone tend to elevate blood glucose. What combinations
of hypo- and hypersecretion would increase or decrease the
circulating blood glucose.
The hypersecretion of growth hormone, epinephrine,
glucocorticoids, glucagon and thyroid hormone will increase the
circulating blood glucose.
4. Why must the patient be fasting before beginning the glucose
tolerance test?
To ensure that the blood glucose level of the patient is not affected
by her/his own intake of diet so that the results obtained is the best.
5. Why is an amount of 1.75 g/kg body weight specified for children
as opposed to a standard 75 g glucose in the glucose tolerance test?
It is because children need less energy/glucose than the adult.
6. What are the steps whereby the carbohydrates of the diet are
converted to glucose in the body?
Digestion of carbohydrates starts in the mouth, when the saliva we
secrete which contains ptyalin starts to work on the carbohydrates.
As it gets to the stomach, further digestion and breakdown with
acids continue. In the intestines, intestinal juices further break down

NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350
the carbohydrates. It is absorbed in the blood stream and carried to
the liver which converts it into glucose.

(B) ANALYSIS OF PLASMA LIPOPROTEINS

Aims of practical :
1. To carry out a biochemical analysis of the cholesterol and
triglyceride profile of the serum prepared from three donor
volunteers
2. To make a clinical assessment of the results with respect to
risk of development of cardiovascular disease
Methods :
i. Determination of total cholesterol in plasma (or serum)
1. Two cuvettes are prepared and labeled as A and B respectively.
2. 1 ml of reagent solution which are phosphate buffer, 4aminophenazone, phenol, 3,3-dichlorophenol, dydroxyethoxy-nalkanes, cholesterol, esterase, cholesterol oxidase, peroxidase are
pipetted into Cuvette A and Cuvette B. (1 cm light path)
3. However, for Cuvette B, 10 l plasma is also added into Cuvette
B.
Cuvette
A
B
Reagent
10 l plasma
1 ml reagent solution 1 ml reagent solution
Table 2 : shows two cuvettes labeled as Cuvette A and Cubette B
and its reagent
4. The reagents are mixed in both Cuvette A and Cuvette B and
these two cuvettes are incubated for 20 minutes at room
temperature.
5. Absorbance of sample against blank at 500 nm is read within 10
minutes.
Results :

NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350
Cuvette
Absorbance

Table 3 : shows absorbance of sample against blank for Cuvette A

and Cuvette B.

Calculation : Concentration of cholesterol in the plasma (mmol/l)

c= 14.3 x A500nm sample
Cuvette
A
B
Concentration of
cholesterol in the
plasma (mmol/L)
Table 4: shows concentration of cholesterol In plasma (mmol/L) in
Cuvette A and Cuvette B
ii. HDL- cholesterol
Methods :
1. 0.5 ml of plasma and 1 ml of precipitating solution (content :
phosphotungstic acid 0.55 mmol/l + MgCl2 25 mmol/l) are pipetted
into a centrifuge tube
2. The content of centrifuge tube is mixed and let stand for 10
minutes at room temperature.
3. The tube is centrifuged for 10 minutes at 400 rpm.
4. After centrifugation, the supernatant is aspirated off into a small
tube. The cholesterol in this supernatant is measured as follows :
Pipette into test tube A
Distilled water
0.1 ml
Supernatant
Reagent solution
1 ml
Table 5 : shows Centrifuge A and Centrifuge B

B
0.1 ml
1 ml
with its content

NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350
5. The content is mixed and both test tube is incubated for 20
minutes at room temperature.
6. Absorbance of sample against blank is measured within 10
minutes.
Results :
Test tube
A
B
Absorbance
Table 6 : shows absorbance of sample against blank for each test
tube

Calculation of HDL-Cholestrol :
Concentration of HDL-cholestrol (mmol/L) = 4.65 x A500nm sample
A

Concentration of
HDL-cholestrol
(mmol/L)

Table 6 shows : concentration of HDL-cholestrol (mmol/L) for test

tube A and test tube B
iii. Determination of triglycerides in serum or plasma
Methods :
1. The following solutions are mixed and left to stand for 20 minutes
at room temperature.
Blank
1 ml

Standard
1 ml

Chromogen
reagent
Standard
10 l
Plasma
Table 7 shows mixture of each solution
2. Absorbance is read at 546 nm

Single
1 ml
10 l

NAME : MOHAMAD LAWI, NUR AFIFAH

STUDENT ID : 14205350
Results :
Blank

Standard

Single

Absorbance
Table 8 shows absorbance of sample against blank for each solution

Calculation :
Absorbance of sample
Absorbance of standard

x Concentration of standard

Blank
Standard
Single
Concentration
of triglyceride
(mmol/L)
Table 9 shows concentration of triglyceride (mmol/L) for each
solution