Estimation of Serum Triglycerides, Practical demonstration lecture for 2nd year MBBS students of Bannu Medical College, Bannu. Lecture delivered on 13/03/2018

1. ESTIMATION OF
SERUM
TRIGLYCERIDES
By:
Dr. Tehmas Ahmad Khan,
Demonstrator Biochemistry Department,
Bannu Medical College, Bannu.

2. Background
 Simple Lipid --- Easter of glycerol with 3 fatty acids
 Main Lipid of Plasma.
 Takes less space to store in body as compared to glycogen
 Formation:
 Synthesized in the intestinal mucosa and Liver by combining glycerol with 3 fatty
acids
 Cleaved by Lipase and free fatty acids are released, that helps in Energy
requirements of body.
 Dietary components are absorbed via chylomicrons.

3. Normal Range
Adults
 Normal Level: 60-200 mg/dL
 Borderline Level: 200-499 mg/dL
 High: >500mg/dL

4. Functions of Triglycerides
 Contribution to Cell Membrane.
 They also provide thermal insulation.
 Imparts flavor to food items.
 Source of Energy.

5. CLINICAL CORRELATION

6. Hypo-Triglyceridemia
 Autoimmune Causes: Systemic Lupus Erythematosus
 Iatrogenic: Ascorbic acid, asparaginase, clofibrate, fenofibrate, fish oil, gemfibrozil,
nicotinic acid, statins
 Hyperthyroidism
 GI: Celiac Disease, Crohn’s disease, Malabsorption syndrome
 Malnutrition
 Low Fat diet

7. Hyper-Triglyceridemia
Causes
 High Carbohydrate Diet
 High fat diet
 Obesity
 Diabetes Mellitus
 Alcohol Consumption
 Kidney Failure
 Lysosome diseases
 Glycogen Storage Diseases
 HIV Medications
 Familial Disorders
Note: Hypertriglyceridemia is usually asymptomatic until triglycerides are greater
than 1000-2000 mg/dL

8. Hyper-Triglyceridemia
 Signs and Symptoms:
 GI: Pain in the mid-epigastric, chest, or back regions; nausea, vomiting
 Respiratory: Dyspnoea
 Dermatologic: Xanthomas
 Ophthalmologic: Corneal arcus, xanthelasmas
 Hypertriglyceridemia Leads to:
 Cerebrovascular/Cardiovascular Events
 Acute Pancreatitis
 Chylomicron Syndrome

9. Treatment
 Medications used in the management of hypertriglyceridemia include the following:
 Fibric acid derivatives (eg, gemfibrozil, fenofibrate)
 Niacin (slow-release, immediate-release, extended-release formulations)
 Omega-3 fatty acids (eg, omega-3-acid ethyl esters)
 HMG-CoA reductase inhibitors (eg, atorvastatin, pravastatin, lovastatin, simvastatin,
rosuvastatin)

10. PRACTICAL / PROCEDURE

11. Requirements
 Standard Solution
 Standard Solution: Concentration of 200mg/dL
 Working Reagent:
 Pipes Buffer pH 7.0 50 mmol/L
 4-chlorophenol 6.0 mmol/L
 Magnesium aspartate >0.5 mmol/L
 Lipase >10 K U/L
 Peroxidase >2.0 KU/L
 4-Aminoantipyrine 1.0 mmol/L
 Glycerol-3-phosphate oxidase >3.5 K U/L
 Glycerol kinase >750 U/L
 ATP 1.0 mmol/L
 Sodium Azide 8.0 mmol/L

12. Principle
 Measurement of triglycerides is based on determination of Light absorbance by the
color compound formed. (Quinoneimine formation)
 Forms a Red Color compound
 The intensity of the red color is directly proportional to the concentration of
triglycerides in the sample.
 It is determined by monitoring light absorbance at 546 nm.
 Reaction:

13. Reactions

14. Precautions
 The only acceptable anticoagulants are heparin and EDTA.
 Fresh Serum to be obtained for Experiment.
 Ideally, after Centrifugation, serum should be immediately separated from blood
cells.
 Do not try to ingest or inhale the Working Solution. In case of contact, wash
thoroughly and seek medical help.
 Reagent (R) contains sodium azide which may react with copper or
lead plumbing.

15. Procedure
 Take Three Test Tubes and Mark then Unknown(U), Standard(S) and Blank(B).
 Take 1ml of Working Reagent in each tube.
 Add 10 µl of distilled water into Blank(B).
 Add 10 µl of Standard Solution into Test tube marked as Standard(S).
 Add 10 µl of Serum into the test tube marked as Unknown(U).
 Mix and Incubate for 10minutes at 20—25 degree Celsius.
 Check light absorbance via a calorimeter at 546nm.
 Unknown Absorbance
Calculation: Triglycérides concentration (mg/dl) = ------------------------------ x 200
 Standard Absorbance

16. Limitations
 Minimum detectable limit of this process in 5 mg/dl.
 Maximum detectable limit of this process is 1000 mg/dl.
 No significant effect of hemolysis
 In Case of Jaundice, Bilirubin Value of more then 10mg/dl leads to decrease in
triglycerides value estimation.
 Of the drugs tested in-vitro, methyldopa and levodopa cause artificially low
triglyceride values at the tested drug Level
 Analytical Value: 5 - 1000 mg/dL

17. Resources Used:
 Resources used:
 Textbook of Medical Biochemistry by M.N.CHATTERJEA
 Spectrum Diagnostics Literature
 Wikipedia and Internet