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1) ANALYSIS OF INHIBITION OF GLUCOSE BY SELECTED NATURAL


AND ARTIFICIAL AGENTS WITH REFERENCE TO DIABETES MELLITUS.

2ND SLIDE

INTRODUCTION :- Glucose an monosaccaride (or) simple sugar is an


important carbohydrate in Biology. The living cell uses it as a source of
energy, and starts cellular respiration in both prokaryotes and
Eukaryotes.

AS A ENERGY SOURCE :- Glucose is a ubiquitous fuel in Biology through


glycolysis and later in citric acid cycle (TCAC) Glucose is oxidized
eventually to form CO2 and water, yielding energy mostly in the form of
ATP.
As an energy source glucose is a primary source of energy for
the brain.

AS A PRECURSOR :- Glucose is critical in the production of proteins and


in lipid metabolism, precursor for Vitamin C.
GLUCOSE ROLE IN BLOOD SUGAR HOMEOSTASIS :- Blood sugar is the
amount of glucose in Blood. Glucose transported via the blood stream.
Blood sugar concentration is tightly regulated in the human body
between 80 – 120 mg/dl.
If blood sugar levels drop too low, a potentially fatal condition
called hypoglycemia.
If the blood sugar levels remain too high condition called
hyperglycemia.

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GLUCOSE TRANSPORT (OR) GLUCOSE UPTAKE :- Glucose cannot diffuse


directly into the cell but enters by one of the two transport
mechanisms.
I. BY FACILITATED TRANSPORT :- For Glucose entry into cells,
facilitated transport is mediated by a family of at least five glucose
transporters in the cell membrane, designated GLUT – 1 to GLUT – 5.
Extra cellular glucose binds to the transporter which then alters its
confirmation, discharging glucose with in the cell. This mechanism is
mainly seen in adipose tissue and RBC.
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SLIDE

II. BY COTRANSPORT :- Cotransport is a carrier – mediated process


in which the movement of glucose is coupled to the concentration
gradient of Na+ which is transported into the cell at the same time.
This mechanism is mainly seen in epithelial cells of intestine, renal
tubules etc.

AIM & OBJECTIVES :-

1) The aim of study was to investigate the influence of various


natural and artificial compounds on the uptake of glucose by the cells
of Diabetic patients.
2) Subjects will be selected according to the degree of blood
glucose level, age and sex.

3) In the study usually the subjects must have increased fasting


glucose level.

6TH SLIDE

GLUCOSE UPTAKE INHIBITORS :

There are various kinds of materials were found to inhibit the uptake of
glucose by the cell both In Vivo and In Vitro. The glucose uptake
inhibitors are two types.

1) Natural glucose uptake inhibitors.


2) Artificial glucose uptake inhibitors.

I. NATURAL GLUCOSE UPTAKE INHIBITORS :

These are obtained from Natural Sources.

1) Several natural flavonoids blocked glucose uptake in myelocytic


cells and lymphocytic cells in vitro. (Jae. B. Park 1999).
2) Turmeric :-

Abstract

The effects of the turmeric ethyl acetate fraction (TEF) from the
methanolic extract from Curcuma longa L. on lipid metabolism and
underlying mechanisms of lipolysis were investigated in 3T3-L1
adipocytes. The intracellular lipid droplets were stained with Oil red O
dye and quantified. Compared to the control, lipid accumulation was
significantly decreased by 46.6% with treatment by TEF at the
concentration of 20 microg/mL. The intracellular triglyceride (TG) level
was also reduced by 37.9% at the concentration of 20 microg/mL. To
determine the mechanism for TG content reduction, levels of glucose
uptake and glycerol release were measured. Incubation of the 3T3-L1
adipocytes with TEF for 4 hours significantly lowered the cellular level
of glucose in a dose-dependent manner. Furthermore, cellular
expression of insulin-responsive glucose transporter (GLUT)-4 was
decreased by 46%, indicating that reduced glucose uptake was due to
a decrease in cellular GLUT-4 expression. In addition, the level of free
glycerol released into the cultured medium was increased by 36.4%
with the treatment by TEF. In subsequent measurements using
quantitative real-time polymerase chain reaction, mRNA levels of
hormone-sensitive lipase (HSL) and adipose TG lipase (ATGL) were
elevated by 34.8% and 16.9%, respectively, at the concentration of 20
microg/mL. These results suggest that TEF partially inhibits lipogenesis
by the suppression of glucose uptake via the decreased expression of
cellular GLUT-4 and stimulates lipolysis through the induction of HSL
and/or ATGL gene expression, resulting in the increased glycerol
release.

8th slide

II. ARTIFICIAL GLUCOSE UPTAKE INHIBITORS :

1. Sodium Fluoride – Potassium Oxalate :-

This is an anticoagulant used in clinical laboratory for prepairing


blood specimens for plasma glucose determination.

Fluoride is an inhibitor of glycolytic enzymes and thus prevents


the loss of glucose from plasma by inhibiting the glucose uptake by the
RBC.
2. Two synthetic detergents :- Triton X – 100 and do decyl trimethyl
ammonium bromide have been found very strong inhibitors.

Kinetic studies showed that these detergents behaved as mixed


type inhibitors i.e. inhibits unspecifically all the transport mechanisms
studied. (Favill F etal., 1988)

3. Arsenicals.

4. 5 – Hydroxy indoleacetic Acid.

5. α - Glycosidase inhibitors etc.

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WORK DONE TILL DATE :-

Insulin, contraction, and the nitric oxide (NO) donor, sodium


nitroprusside (SNP), all increase glucose transport in skeletal muscle.
Some reports suggest that NO is a critical mediator of insulin – and/or
contraction – stimulated transport. Isolated soleus and extensor
digitorum longus (EDL) muscles from rats were treated with various
combinations of SNP (maximum 10 mmol/1), insulin (maximum 50
mU/ml), electrical stimulation to produce contractions (maximum 10
min), wortmannin (100 nmol/l), and/or the NO synthase (NOS) inhibitor
NG – monomethyl-L-arginine (L-NMMA) (0.1 mmol/l). The combinations
of SNP plus insulin and SNP plus insulin and SNP plus contraction both
had fully additive effects on 2-deoxyglucose uptake. Wortmannin
completely inhibited insulinstimulated glucose transport and only
slightly inhibited SNP-stimulated 2-deoxyglucose uptake.

Parameters Analysed :- In the study effect of Sodium Nitroprusside on


glucose uptake by RBC was studied.

Bio chemical Parameters used :-

1. Estimation of glucose conc. in RBC cells by GOD – POD


Method.

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INITIAL STUDY
Step – I. Blood of 2 ml is taken and centrifuse to get plasma and
estimate the Blood glucose immediately. It is (Initial control).

Step – II. Blood of 2 ml is taken and add trisodium citrate which is


anti coagulant. Kept aside for 30 min. to separate the plasma, estimate
the Blood glucose. It is (Final Control)

Glucose
Glucose
level
Body wt. in level (final
S.No. Sex Age in yrs (initial
kg. control in
control in
mg/dl)
mg / dl)
P1 FM 54 65 202 154
P2 M 63 70 212 160
P3 M 65 68 223 156
P4 FM 67 66 240 190
P5 M 72 75 261 178

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Discussion :-

In the initial control there is a increase in Blood sugar levels because


glycolysis is not taken place because immediately we are estimating
RBS.

But in final control the blood is allowed to stand for 30 min at 37 oC


glucose uptake by the RBC takes place i.e. Glycolysis have taken place
and results in decrease in Blood sugar levels.
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Blood glucose levels in 0.1 m SNP treated sample (mg/dl) :

Glucose
Glucose
level
Body wt. in level (final
S.No. Sex Age in yrs (initial
kg. control in
control in
mg/dl)
mg / dl)
P1 FM 54 65 154 160
P2 M 63 70 160 165
P3 M 65 68 156 163
P4 FM 67 66 190 198
P5 M 72 75 178 186

Discussion :-

In the Test sample there is an increase in Blood sugar level because


glucose uptake by RBC is inhibited by the SNP.
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Advantages :

1) Used in clinical Biochemistry according to Biochemical


parameters selected.

2) Inhibit the glucose uptake.

3) To keep the glucose in equilibrium mainly in hypoglycemic


condition.

4) used in long term investigation.

Disadvantages :

1) Artificial glucose uptake inhibitors are toxic, care should be taken


while handling.

2) They may interfere with the Reagents sometimes.

Future plan :

The glucose concentration is different cells of diabetic patients


like RBC, lever cells etc. will be studied by using different inhibitors in
vitro and in vivo by using natural sources like Turmeric and Medicinal
plants, which will not have any side effects.

Conclusion:

The in Vitro glucose uptake inhibition study was carried out in


Blood cell of diabetic patients by using Sodium Nitro prusside markedly
reduce the glucose uptake.
ANALYSIS OF INHIBITION OF GLUCOSE BY SELECTED
NATURAL AND ARTIFICIAL AGENTS WITH REFERENCE TO
DIABETES MELLITUS.

Abstract : The living cells uses glucose an monosaccharide as a source


of energy for the brain, which is ubiquitous fuel in Biology. Yield energy
mostly in the form of ATP.

Glucose is critical in production of proteins, lipid metabolism precursor


for Vitamin C. Play a Role in Blood Sugar Homeostasis. Transported via
blood stream and maintain levels tightly between 80 – 120 mg/dl.
Below normal level called Hypoglycemic and above 180 mg/dl
Hyperglycemic conditions.

Glucose uptake takes place by facilitated Transport with the help of


GLUT – 1 to GLUT – 5 and through concentration gradient of Na+.
Various materials inhibit glucose uptake in Vivo and in Vitro.

Inhibitors which are two types, Turmeric which is a natural source of


inhibition and Sodium Nitro prusside which is a chemical with various
concentration influence the Blood glucose uptake by the RBC. Initial
and final values of samples of diabetic patients are taken and
observed. And it is seen that the glucose uptake was inhibited by the
above two inhibitors and studied further.