1. LIPIDS
Study Guide
Pg 171

2. Module Focus Lipids are organic compounds that are insoluble in water
They dissolve in organic solvents such as alcohol
Hyperlipidemia is a biochemical state often associated with the development of atherosclerosis and coronary heart disease (CHD)

3. CLASSIFICATION OF LIPIDS
Fatty acids
Triglycerides
Cholesterol
Phospholipids
Lipoproteins

4. FATTY ACIDS Functions: Metabolic energy
Building blocks for trigylerides & phospholipids
Not routinely measured

5. TRIGLYCERIDES Functions: Main form of lipid storage
Provides energy for cell
Insulator of vital organs
Trigs are broken down by the enzyme lipase, epinephrine & cortisol
Trigs are transported in plasma as chylomicrons or VLDL

6. CHOLESTEROL 75% Esterified form, 25% Free cholesterol Function:
Precursor of steroid hormones (estrogens, androgens)
Cell membrane structure
Formation of bile acids
Transported in plasma by LDL & HDL

7. PHOSPHOLIPIDS Function: Increases solubility of cholesterol
Forms a coating that surrounds cholesterol and triglycerides and glues the lipoprotein coreBilayer of cell membranes
Sphingolipids/sphingomyelin
Not measured routinely

8. LIPOPROTEINS Lipid + Protein (apoprotein)
Protein provides an outer coating for the lipids making them soluble
Transports lipids in the plasma
Chylomicrons
Very low density lipoproteins (VLDL)
Low density lipoproteins (LDL)
High density lipoproteins (HDL)

9. LIPOPROTEIN METABOLISM
In the small intestine, food is digested by pancreatic enzymes and bile acids.
Fats (triglycerides) are hydrolyzed by lipase to fatty acids and glycerol.
They are absorbed & reassembled by the intestinal mucosa into new triglycerides, which are packaged into a structure called CHYLOMICRONS that provide a protein coating to make them soluble.

10. Chylomicrons enter the lymphatics and eventually reach the circulation
Chylomicrons enter the lymphatics and eventually reach the circulation. Peak triglyceride levels usually occur 30 – 90 minutes after a meal.
In the blood, chylomicrons are exposed to the enzyme lipoprotein lipase (LPL) on the surface of the endothelial cells. LPL hydrolyzes triglycerides into monoglycerides and fatty acids that can be absorbed by cells for energy and/or reassembled into trigs and stored in adipose tissue for future energy.

11. LPL leaves a chylomicron remnant that is taken up by the liver.

12. ENDOGENOUS PATHWAY
The liver is the principle site for endogenous lipid metabolism as well as apoprotein formation.
Triglycerides and cholesterol are packaged into triglyceride rich very low density lipoproteins (VLDL).
LPL hydrolyzes the VLDL trigs leaving an intermediate density lipoprotein (IDL

13. As triglyceride content decreases, cholesterol content increases, until the end product low density lipoprotein (LDL) is formed.
LDL is rich in cholesterol and transports cholesterol to tissues.
High Density Lipoprotein (HDL) is produced by the liver and intestine, it transports cholesterol from the cells back to the liver to be excreted.

16. LIPOPROTEINS Chylomicrons Very Low Density Lipoprotein (VLDL)
Low Density Lipoprotein (LDL)
High Density Lipoprotein (HDL)

17. FUNCTION Essential for the transport of insoluble lipids in plasma
Provides cholesterol, phospholipids and triglycerides to tissues for:
Energy
Membrane synthesis
Hormone synthesis

18. LIPOPROTEIN STRUCTURE
Lipids + Protein
Inner core of lipids
Outer coating of protein to make them soluble in plasma
PROTEIN COATING

19. CHYLOMICRONS Protein content = 1%
Not normally present in fasting specimens
Transport dietary lipids (exogenous triglycerides) to body tissues
Responsible for most postprandial lipidemia
Major lipid = 95% trigs
Protein content = 1%

20. VERY LOW DENISTY LIPOPROTEIN “VLDL”
Transports body-made (endogenous) trigs to adipose tissue
55% trigylcerides
10% protein
Causes lipemia in serum specimens

21. LOW DENSITY LIPOPROTEIN LDL
Major transporter of cholesterol
Transports cholesterol to tissues to be deposited
45% cholesterol
20% protein
 levels associated with Coronary Heart Disease (CHD)
Remember: Low density is “LETHAL”

22. HIGH DENSITY LIPOPROTEIN HDL
Transports cholesterol from tissues to the liver to be excreted
Increased levels help prevent CHD
50% protein
20% cholesterol
30% phospholipids

23. CLASSIFICATION
LIPOPROTEINS

24. 4 CLASSES BASED ON: Electrophoretic mobility
Flotation characteristics: reference method
Chemical composition

25. ELECTROPHORETIC MOBILITY
(+)Anode (-) Cathode
CHYLOMICRONS
ALPHA
HDL
BETA
LDL
PRE BETA
VLDL

26. FLOTATION OR ULTRACENTRIFUGATION
The greater the protein content, the higher the density.
Current
reference method
chylomicrons
VLDL
LDL
HDL

27. CHEMICAL COMPOSITION Chylomicrons = dietary triglycerides
VLDL = body-made trigs
LDL = carries the “BAD” cholesterol
HDL = carries the “GOOD” cholesterol

28. CHOLESTEROL Specimen requirements: Fasting not required No alcohol
No medications
No special diet for 2 weeks prior to analysis

29. METHODS OF MEASUREMENT
Abell-Kendall = current reference method
Utilizes Liebermann-Burchard reagent
Acetic anhydride & sulfuric acid
Not practical for routine use
Enzymatic is most commonly utilized

30. ENZYMATIC
Cholesterol esters are hydrolyzed to FREE cholesterol and fatty acids by the enzyme cholesterol esterase
FREE cholesterol is oxidized by the enzyme cholesterol oxidase to form hydrogen peroxide
The H2O2 reacts with a dye to form a colored compound.

31. REFERENCE RANGES <200 MG/dl is desirable 200 – 239 is borderline
> 240 is considered high risk

32. CLINICAL INTERPRETATION Increased Cholesterol
Coronary artery disease
Genetic defects in liver or lack of apoprotein B receptors on cell surface
Diabetes mellitus
Liver disease
Hypothyroidism
Nephrotic syndrome
Menopause due to  estrogen
Increases in cholesterol levels do not change the appearance of plasma

33. TRIGLYCERIDES Specimen requirements 12 – 14 hr fast is mandatory
Nonfasting specimens = lipemic plasma due to the presence of chylomicrons which carry dietary trigs
Triglyceride levels peak 4 – 6 hrs after a meal

34. ENZYMATIC METHOD (detects glycerol)
Lipase
Trigs GLYCEROL + fatty acids
Trigs are hydrolyzed by lipase to form glycerol
Glycerol dehydrogenase
Glycerol + NAD NADH dihydroxyacetone
Glycerol is quantitated by various methods

35. Reference Ranges < 150- 200 mg/dl for a fasting specimen
200 – 400 mg/dl = borderline high
>400 mg/dl = high

36. CLINICAL INTERPRETATION
Increased Triglycerides
One of the most common causes is a nonfasting specimen
Oversynthesis
Diabetes mellitus, pancreatitis, alcoholism, obesity
Impaired catabolism – lack of lipoprotein lipase, present in capillary walls, it hydrolyzes trigs
Remember: increased trigs causes lipemia

37. HDL CHOLESTEROL The “GOOD” Cholesterol ?
Remember: “H” is for healthy lower the risk of coronary artery disease
The higher the level of HDL, the lower the risk of coronary artery disease.

38. HDL METHODS
Precipitation of LDL, VLDL, and chylomicrons from serum specimen by the addition:
Phosphotungstate + MgCl2 or
Dextran sulfate + MgCl2

39. After centrifugation, HDL is the only lipoprotein remaining in the supernatant
A cholesterol method is then performed on the supernatant to determine the HDL
HDL Cholesterol
VLDL,LDL & Chylos

40. HDL Precipitation method is time consuming
Newer methods utilize immunoseparation techniques that do not require an extraction step.

41. CLINICAL INTERPRETATION
> 55 mg/dl = Low risk for CAD
< 40 mg/dl = High risk for CAD
HDL carries cholesterol from tissues to the liver for excretion

42. LDL CHOLESTEROL The BAD cholesterol Remember: “L” is for LETHAL
LDL carries cholesterol to tissues to be deposited
The higher the LDL, the greater the risk for CAD

43. Total Cholesterol – (HDL chol + Trigs)
LDL METHODS
LDL can be calculated:
Total Cholesterol – (HDL chol + Trigs) 5
Calculation is not very accurate if triglyceride measurement is nonfasting
Newer methods utilize immunoseparation and directly measure LDL

44. REFERENCE RANGE
< mg/dl

45. CLINICAL INTERPRETATION
< mg/dl = Low risk for CAD
135 – 159 = Possible risk for CAD
> 160 = High Risk for CAD

46. VLDL CHOLESTEROL Can be calculated by: TRIGYLERIDES 5
Not very accurate

47. Total Cholesterol HDL Cholesterol
RATIO
Divide the patient’s total cholesterol by their HDL cholesterol level
Example:
Cholesterol = 240 mg/dl
HDL Cholesterol = 40 mg/dl
< 4 = Low risk for CAD
>5 = High risk for CAD
240 = 6 40

48. LIPOPROTEIN ELECTROPHORESIS
Principle: Same as SPE.
Lipoproteins carry a net negative charge at a pH of 8.6
Lipoproteins can be separated into 4 components
CHYLOS
PREBETA
BETA
Alpha
- pole
+ pole

49. LPE Specimen: EDTA fasting plasma Support medium: agarose gel
Lipid Stains: Oil Red O, Fat Red B, Sudan Black

50. CLINICAL INTERPRETATION
LPE is infrequently performed since total cholesterol, HDL, LDL & triglycerides provides sufficient information.
Normal SPE pattern: only alpha, beta & a small amount of prebeta will be visible
Chylomicrons should not be present in a normal fasting specimen

51. Apoproteins pg 178
Proteins associated with lipids.

52. SERUM APPEARANCE AFTER 18 HRS at 40C
CREAMY LAYER
CLEAR
LIPEMIC

53. INTERPRETATION Normal appearance is clear
If a creamy layer forms at the top of the tube after 18 hrs, this is indicative of chylomicrons (dietary trigs)
If the serum is cloudy or lipemic throughout the tube after 18 hrs, this is indicative of body made trigs (VLDL)
REMEMBER: Increases in cholesterol do not change the serum appearance.