INTERPRETATION OF LIPID

PARAMETERS
(LIPID PROFILE)
Integrated Multisystem and Therapeutics Block (CMD 342)
Phase III, Year III
By
Prof. Dr. Muhammad Jawed
MBBS, M.Phil, Ph.D

Department of Medical Biochemistry

College of Medicine
Qassim University 1
 INTERPRETATION OF LIPID PARAMETERS
(LIPID PROFILE)

Objectives
By the end of the session student will be able to:
§ List the different parameters evaluating serum lipid profile.

§ Describe the principle of estimation of triacylglycerols (TAG) and total

HDL, LDL & VLDL-cholesterol.

§ Estimate serum TAG, total cholesterol and calculate LDL-cholesterol as well

as the ratio between LDL- & HDL- cholesterol.

§ Describe the diagnostic importance of each of the lipid profile parameters &
their relations to atherogenicity

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 LIPID PROFILE
It is a group of tests that measure
the concentration of various lipids that circulate in the blood
Cholesterol
HDL Cholesterol

LDL Cholesterol

Lipid Profile VLDL Cholesterol

Triglycerides

Cholesterol/HDL ratio

LDL /HDL ratio

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The parameters are often used to assess the risk of cardiovascular diseases
 LIPID PROFILE

CAD Family history

Hypertension

Obesity

Diabetes
Indications

Renal diseases

Liver diseases

Thyroid related disorders

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 LIPID PROFILE
Cholesterol
• Normal Values: vary with age, sex, diet and geographic or cultural region
– Adult, Fasting:
• Desirable level: 140-199 mg/dL
• Borderline high: 200-239 mg/dL
• High: >240 mg/dL
(Ref: A Manual of laboratory and diagnostic tests. 8th edition. Frances Fischbach
&Marshall Dunning)

• Waxy, fat-like substance required for the normal functioning of the body
• Naturally present in cell walls or membranes everywhere in the body, including the
brain, nerves, muscles, skin, liver, intestines and heart.
• The cholesterol in a person's blood originates from two major sources; the diet and
the liver
– Dietary cholesterol comes primarily from meat, poultry, fish, and dairy products.

Cholesterol is carried in the blood as particles of differing sizes and densities along
with lipoproteins.
• HDL Cholesterol (Good cholesterol)
• LDL Cholesterol (Bad cholesterol)
• VLDL Cholesterol

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 LIPID PROFILE
Cholesterol
If cholesterol is present in higher Elevated Cholesterol Levels:
concentrations, the excess may be deposited in 1. Type II familial hypercholesterolemia
arteries, including the coronary (heart) arteries, 2. Hyperlipoproteinemias type I, IV and V
3. Cholestasis
where it contributes to the narrowing and 4. Hepatocellular disease
blockages that cause the signs and symptoms 5. Nephrotic syndrome, glomerulonephritis
of heart disease. 6. Chronic renal failure
7. Alcoholism
8. Diet high in fat and cholesterol
9. Obesity
10. Poorly controlled diabetes mellitus

Decreased Cholesterol Levels:

1. Hypo-a-lipoproteinemia
2. Myeloproliferative disease
3. Malabsorption syndrome
4. Severe burns, inflammation
5. COPD

Ref: A Manual of laboratory and diagnostic tests. 8th

edition. Frances Fischbach &Marshall Dunning

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 LIPID PROFILE
Cholesterol

• It is recommended that healthy adults with no other risk factors

for heart disease be tested with a fasting lipid profile once
every five years. They may be screened using only a
cholesterol test and not a full lipid profile.

• However, if the cholesterol test result is high, then they may

have follow-up testing with a lipid profile.

• If they have other risk factors or have had a high cholesterol

level in the past, they should be tested more regularly and
should have a full lipid profile.

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 Estimation of cholesterol in the serum
Principle

In the presence of cholesterol esterase, cholesterol esters in the serum sample are
hydrolyzed to cholesterol and free fatty acids. The cholesterol produced is oxidized by
cholesterol oxidase to cholestenone and hydrogen peroxide. Hydrogen peroxide is
detected by a chromogenic oxygen acceptor, phenol-4-aminoantipyrine, in the
presence of peroxidase. The red quinone imine formed is proportional to the amount
of cholesterol present in the sample , and absorbs maximally at 505 nm.

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 Estimation of cholesterol in the serum
Procedure (Protocol)
Blank Standard Sample
Working reagent 1.0 ml 1.0 ml 1.0 ml
Working reagent contains cholesterol esterase, cholesterol oxidase and
peroxidase in addition to phenol and antipyrine. Warm the reagent at 37oC
for 3 minutes
Cholesterol (Standard) soln. - 20 μl -
Serum / sample - - 20 μl
Distilled water 20 μl - -
Mix thoroughly; incubate at 37oC for 10 minutes. Record the absorbance of
test and standard against blank at 505 nm. (The final color is stable for at
least 30 minutes and should be analyzed within time)
• Calculate the concentration of cholesterol (mg/dL) in the sample using the following
relationship:

Concentration of cholesterol (mg/dL) = absorbance of sample × Concentration of standard

absorbance of standard

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 Estimation of HDL cholesterol
Principle

When serum is reacted with the Phosphotungstic acid and Magnesium chloride , it
causes the precipitation of LDL and VLDL proteins. HDL fraction however
remains in the supernatant. The HDL Cholesterol is then estimated by the same
procedure as described for total cholesterol above and follows the same principle

Two Steps

1. Separation of HDL Cholesterol fraction

2. Estimation of HDL Cholesterol

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 1. Separation of HDL Cholesterol fraction

Protocol
Reagent Volume

Precipitating reagent (Phospho

tungstic acid, 0.55 mmol/L;
0.5 ml
Magnesium chloride, 25 mmol/L)

Serum 0.2 ml

Mix thoroughly, Allow to stand at Room temperature for 10 minutes.

Centrifuge at 4000 rpm for 20 minutes. Decant the supernatant carefully.

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 2. Estimation of HDL Cholesterol
Protocol
Blank Standard Sample
Working reagent 1.0 ml 1.0 ml 1.0 ml
Working reagent contains cholesterol esterase, cholesterol oxidase and peroxidase in
addition to phenol and antipyrine . Warm the reagent at 37oC for 3 minutes
Cholesterol (Standard) soln. - 100 μl -
Supernatant (Obtained from serum sample) - - 100 μl
Distilled water 100 μl - -
Mix thoroughly; incubate at 37oC for 10 minutes. Record the absorbance of test and
standard against blank at 505 nm. (The final color is stable for at least 30 minutes and
should be analyzed within time)
• Calculate the concentration of HDL cholesterol (mg/dL) in the sample using the following
relationship:
Concentration of HDLcholesterol (mg/dL) = absorbance of sample × Concentration of standard
absorbance of standard

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 HDL cholesterol
Decreased levels are associated with:
Normal Values 1. Familial hypo-alpha-lipoproteinemia
2. a-β lipoproteinemia
• HDL
3. Hypertriglyceridemia
– Male: 35 - 65 mg/dL 4. Poorly controlled diabetes mellitus
– Female: 35 - 80 mg/dL 5. Hepatocellular disease
6. Cholestasis
7. Chronic renal failure, uremia, nephrotic
syndrome
Increased levels:
1. Familial hyper- alpha-lipoproteinemia
2. Chronic liver disease (cirrhosis,
alcoholism, hepatitis)
3. Long term aerobic or vigorous exercise
(Ref: A Manual of laboratory and diagnostic tests. 8th edition. Frances Fischbach &Marshall Dunning)

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 Estimation of triglycerides in serum
Principle
The triglycerides in the serum sample are hydrolyzed enzymatically by the action
of lipase to glycerol and fatty acids. The glycerol formed is converted to glycerol
phosphate by glycerol kinase (GK). Glycerol phosphate is then oxidized to
dihydroxyacetone phosphate by glycerol phosphate oxidase (GPO). The librated
hydrogen peroxide is detected by a chromogenic acceptor, chlorophenol-4-
aminoantipyrine, in the presence of peroxidase (POD). The red quinone imine
formed is proportional to the amount of triglycerides present in the sample.

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 Estimation of triglycerides in serum
Protocol
Blank Standard Sample
Working reagent 1.0 ml 1.0 ml 1.0 ml
Working reagent contains Glycerol 3 phosphate oxidase, lipase, Glycerol kinase,
peroxidase, 4-amino antipyrine, and ATP dissolved in PIPES (piperazine-
N,N′bis[2-ethanesulfonic acid]) buffer, pH 7.5 conataining 4-chlorophenol and
Magnesium ions.
Cholesterol (Standard) soln. - 20 μl -
Serum / sample - - 20 μl
Distilled water 20 μl - -
Mix thoroughly; incubate at 37oC for 10 minutes. Record the absorbance of test
and standard against blank at 505 nm. (The final color is stable for at least 30
minutes and should be analyzed within time)

Calculate the concentration of triglycerides (mg/dL) in the sample using the following
relationship:
Concentration of triglycerides (mg/dL) = absorbance of sample × Concentration of standard
absorbance of standard 15
 Triglyceride values
mg/dL Increased levels in:
1. Hyperlipoproteinemia type I, IIb, III, IV and V
Desirable <150 2. Liver disease, alcoholism
Borderline high 150-199 3. Nephrotic syndrome, renal disease
4. Hypothyroidism
High 200-499
5. Poorly controlled diabetes mellitus
Very High (hypertrig) >= 500 6. Pancreatitis
7. Anorexia nervosa
• >90% of dietary fat
• 95% of fat stored in tissues Decreased levels:
• 80% of Triglycerides are in VLDL 1. Malnutrition
and 15% are in LDL 2. Hyperthyroidism
• Elevated triglycerides along with 3. Brain infarction
elevated cholesterol are risk factor 4. COPD
for atherosclerotic disease. 5. Congenital a β lipoproteinemia
• Needed to calculate LDL-C

(Ref: A Manual of laboratory and diagnostic tests. 8th edition. Frances Fischbach &Marshall Dunning)
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 Estimation of LDL Cholesterol

Direct method: Costly and cumbersome

Indirect method: Needs no additional procedures, only
calculations are required
Friedwald formula:
valid only if cholesterol and TG values are from a fasting specimen and TG must be <
400 mg/dl)
LDL C = TC – TG – HDL
5
Total Lipids (mg/dL) = TC + HDL C + LDL C + TG + 335
Where LDL C represents LDL cholesterol, HDL C represents HDL
Cholesterol, TC represents total cholesterol and TG represents
triglycerides
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 Estimation of LDL Cholesterol
Example 1: Lipid profile of 50-years-old male showed TC 290 mg/dL, TG
370 mg/dL and HDL-C 29 mg/dL. Calculate LDL-C of this patient.

LDL C = TC – TG – HDL
5
LDL C = (290 – 370) – 29
5
LDL C = (290 –74) – 29

LDL C = 216 – 29

LDL C = 187 mg/dL

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 Estimation of LDL Cholesterol
Example 2: Lipid profile of 45-years-old male showed TC 250 mg/dL, TG
350 mg/dL and HDL-C 28 mg/dL. Calculate LDL-C of this patient.

LDL C = TC – TG – HDL
5
LDL C = (250 – 350) – 28
5
LDL C = (250 –70) – 28

LDL C = 280 – 28

LDL C = 152 mg/dL

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 Interpretations
Increased levels of LDL-C: Normal Values (LDL-C):
1. Familial type 2 hyperlipidemia Adults:
2. Secondary causes:
1. High saturated fat and high cholesterol diet
2. Nephrotic syndrome mg/dL
3. Diabetes mellitus Optimal <100
4. Nephrotic syndrome
5. CRF Near Optimal 100-129
6. Hyperlipidemia secondary to hypothyroidism
7. Anorexia nervosa
Borderline high 130-159
8. Porphyria High 160-189
Decreased levels in: Very High >190
1. Hypolipoproteinemia
2. Type 1 hyperlipidemia
3. Apo-C-II deficiency
4. Chronic anemias
5. Hyperthyroidism
6. Acute stress (burns, illness)
7. Inflammatory joint disease

(Ref: A Manual of laboratory and diagnostic tests. 8th edition. Frances Fischbach &Marshall Dunning)
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 Estimation of Total Lipids
Example 3: Lipid profile of 52-years-old male showed TC 285 mg/dL, TG
360 mg/dL and HDL-C 30 mg/dL. Calculate serum LDL-C and total
lipid of this patient.
LDL C = TC – TG – HDL Total Lipids = TC + HDL C + LDL C + TG + 335
5 (mg/dL)
LDL C = (285 – 360) – 30
= 285 + 30 + 183 + 360 + 335
5
= 1193 mg/dL
LDL C = (285 –72) – 30

LDL C = 213 – 30

LDL C = 183 mg/dL

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 The total cholesterol / HDL cholesterol
ratio

• The total chol /HDL ratio is helpful in estimating the risk

of developing atherosclerosis.
• High ratios indicate a higher risk of heart attacks, whereas
low ratios indicate a lower risk.

Average 2 x average risk 3 x average

Risk risk
Male 5.0 9.6 23.4
Female 4.4 7.1 11.0

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 LDL/HDL ratio
• The LDL/HDL ratio is actually a purer ratio than total cholesterol/HDL,
because LDL is a measure of "bad' cholesterol and HDL is a measure of
"good" cholesterol.

• The LDL/HDL cholesterol ratio is more important than individual levels of

HDL cholesterol and LDL cholesterol. However, recent heart Foundation
recommendation is that irrespective of the ratio, one should try to reduce
the LDL cholesterol.

Risk Level Low risk Average risk Moderate High risk

risk

LDL/HDL
Ratio 3.3 – 4.4 4.4 – 7.1 7.1 – 11.0 >11.0

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 INTERPRETATION OF LIPID
PARAMETERS
(LIPID PROFILE

• Lippincott’s Illustrated Review: Biochemistry 7th

Ed.
• A Manual of laboratory and diagnostic tests. 8th
edition. Frances Fischbach &Marshall Dunning
• Current Medical Diagnosis & Treatment-2015,
McGraw Hill.
• Harper’s Biochemistry (30th edition)
• Mark’s basic Medical Biochemistry: A Clinical
Approach

THANK YOU
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