Lab 7 Diabetic profile KAUFaculty of Science Biochemistry

Lab (7): Diabetic profile KAU-Faculty of Science- Biochemistry department Clinical biochemistry lab (Bioc 416) 2012 T. A Nouf Alshareef nf. shareef@hotmail. com

Diabetes Mellitus: • It is a chronic disease due to disorder of carbohydrate metabolism, due to insulin deficiency results in hyperglycemia (increased blood glucose level) & glucourea (presence of glucose in urine). • Associated with several changes in metabolism; such as metabolism of proteins & fats.

Clinical Biochemical Findings in Diabetes: • Glucosuria. • Large volume of urine & increase urination frequency (Polyuria) • Polyphagia (eats more frequently). • Several metabolic changes

Metabolic changes in diabetes Include increase in: v Fat catabolisim leads to increase in FFAs in blood & liver. v Acetyl. co. A leads to increase formation of cholesterol & risk of atherosclerosis. v ketone bodies generation in blood and urine leads to acidosis. v catabolism of tissue protein due to energy requirement (because glucose can't uptake by cells) lead to weight loss and increase in level of amino acids in blood & more formation of urea by deamination of amino acid.

Types of diabetes: • • Type I diabetes mellitus (TIDM) Type 2 diabetes mellitus (TIIDM) Gestational diabetes mellitus (GDM) Other "due to drugs or chemicals"

Diabetic profile: - Is group of tests that are used to diagnose diabetes or its complications , it includes: 1. C-peptide. 2. Blood glucose (4 types: FBS, PPBS, RBS, OGGT) 3. Hb. A 1 C 4. Insulin 5. ICA (islent cell antibody) for type I 6. Ketones 7. Microalbuminurea.

1 - C-peptide: - This test is used to differentiate between type I and II diabetes type I DM: low level of insulin and C-peptide type II DM: normal or high level of C-peptide. Advantages of measuring C-peptide than insulin: • It is better indicator of B-cell function than peripheral insulin. Normal value: Fasting 0. 51 -2. 72 ng/ml

2 - Blood sugars: Types of blood glucose tests: (1) Fasting blood sugar (FBS): • measures blood glucose after fasting for at least 8 -12 hrs • It often is the first test done to check for diabetes. • patient with mild or borderline diabetes may present with normal FBG values. • If diabetes is suspected, GTT can confirm the diagnosis. Normal levels: 60 -110 mg/dl

(2) Post-Prandial Blood Sugar (2 -hour PPBS): • After the patient fasts for 12 hours, a meal is given which contains starch and sugar (approx. 100 gm). • Then after 2 hours blood is collected to measure glucose level. • home blood sugar test is the most common way to check 2 hour postprandial blood sugar levels.

(3) Random blood sugar (RBS) • measures blood glucose randomly at any time throughout the day without patient fasting. • it is useful because glucose levels in healthy people don’t vary widely throughout the day. • blood glucose levels that vary widely may indicate a problem.

(4) Oral glucose tolerance test (OGTT) • Glucose Tolerance is defined as the capacity of the body to tolerate an extra load of glucose or it measures the body's ability to use glucose. • it is series of blood glucose measurements taken after drink glucose liquid • It is considered as definitive diagnostic test for DM. • It is ordered to: - confirm the diagnosis, in pre-diabetic - diagnose gestational diabetes (most commonly) • Recommended if 100 -126 mg/dl or (5. 5 mmol/L-7. 0 mmol/L).

Procedure: • Arrive FBS: After an overnight fasting of 12 -16 hrs • Drink: 75 -100 g dissolved in 250 -300 ml of water and given orally. • After drink: blood samples and urine are collected every 30 min for 3 hrs (1 hr, 1. 5 hr , 2 hr, 2. 5 hr, 3 hr ) • A curve between time and blood glucose concentration, is plotted.

Other types of OGTT: • Extended GTT : glucose measured for 4 -5 hrs after giving glucose to see how the curve behaves below the normal fasting glucose limits. Done in some conditions causing hypoglycaemia. • Cortisone Stressed GTT : Can be used for detecting latent DM. • Intrevenous GTT : - is done if oral glucose is not tolerated or oral GTT curve is flat. - In these cases 20% glucose as 0. 5 g glucose/Kg body weight. - Usually peak occurs within 30 min after infusion and returns to normal after 90 min.

Interpretation: • Normal Response : FBS is normal. After 1 hr it will rise, returns to normal fasting level within 2 hours. • Diabetic curve : FBS: 140 mg/dl or 7. 8 mmol/L. After 2 hr: 200 mg/dl (11 mmol/L) or more. Glucosuria is usually seen • Impaired GTT: - with 2 hrs glucose level between 140 mg/dl - 200 mg/dl - it is not abnormal but must be followed up for DM.

3 - Hb. A 1 c: • Hb. A 1 C: is glucose bound to hemoglobin • Measures blood glucose conc. over a longer period of time • it indicates how well diabetes has been controlled in the 2 -3 months before the test. • The A 1 C level is directly related to complications from diabetes (lower the A 1 C level lower risk for complications) • Type of sample: whole blood in EDTA tube Normal Values: • Glycohemoglobin A 1 c: 4. 5%-5. 7% • Total glycohemoglobin: 5. 3%-7. 5%

Expected range of Hb A 1 c: • Sugar: 90 -150 5 -0% to 7. 0% • Sugar: 150 -180 7. 0% to 8. 0% • Sugar: 180 -360 9. 0% to 14. 0%

Hypoglycaemia : • When blood glucose falls below 60 mg/dl. • Causes: 1. Most commonly seen in overdose of insulin in treatment of DM. 2. Hypothroidism. 3. Insulin secreting tumours of pancrease – rare. 4. Hypoadrenahsm (Addison's disease) 5. Hypopitruitism. 6. Severe exercise. 7. Starvation.

Experiment: Measuring glucose level Principle: GOD Glucose + H O + O Gluconic acid + H 2 2 2 O 2 POD 2 H 2 O 2 + 4 aminoantipyrine + PHBS Quinoneimine dye + H 2 O Red color

Kit components • Glucose Oxidase Reagent : mixture of: glucose oxidase + peroxidase+ aminoantipyrine+ buffer • Glucose standard Reagent : conc. 100 mg/dl or 5. 55 mmol/L

Procedure • Prepare the reaction as the following: Glucose oxidase reagent Sample (serum) Glucose standard Reagent blank Standard Sample 1 ml 1 ml 0. 01 ml or 10 ml - - • Mix, incubates at 37 o. C for 10 min • Read abs at 510 nm 0. 01 ml or 10 ml -

Calculations: Glucose conc. = Abs. sample X conc. standard Abs. standard . .