The Research work was done at The Federal Medical Centre Asaba and The University of Benin.

Introduction

Hypertension has emerged as a significant public health problem in most developing countries of the world ₁ . Uncontrolled hypertension remains a major threat to global health especially in developing countries such as Nigeria, where cardiovascular diseases mainly resulting from hypertension are still responsible for significant rates of death and disability ₂ .

The major risk factors for cardiovascular disorder is associated with plasma cholesterol concentration and fractions known as low density lipoprotein (LDL) cholesterol and very low density lipoprotein (VLDL) cholesterol, whereas the fraction known as high density lipoprotein (HDL) cholesterol appears to have a protective role ₃ . Also peroxidation of these lipids by free radicals is believed to be one of the important causes of cell membrane destruction and could be implicated in the pathogenesis of cardiovascular disorder ₄₅ .

This study is aimed at evaluating the lipids most commonly implicated in hypertensives; assessing the lipid levels in hypertensives and normotensives and to find out if there is any variation in the lipid concentrations of hypertensive patients compared to normotensive patients and also to investigate lipid peroxidation assessed by plasma malonydialdehyde level in hypertensive patients attending the cardiovascular clinic at Federal Medical Centre Asaba.

Materials And Methods

Subjects For The Study

Preparation Of Plasma

Estimation Of Plasma Total Cholesterol

The enzymatic colorimetric method bt Tinder was used ₆ .

Reagents

4-Aminoantipyrine 0.5 mM, Phenol 6mM, 3,5-dichlorophenol 0.2mM, Cholesterol esterase ≥ 500kµ/l, Cholesterol oxidase ≥ 300kµ/l, Peroxidase ≥ 1200kµ/l , Phosphate buffer (pH 6.7) 70mM.

Principle

Figure 1

Procedure

To 1000ul of reagent in tubes meant for samples, standard and blank, 10µl of samples, standard and distilled water were added respectively. The content was mixed and incubated for 5mins at 37 ° C. The absorbance of the samples and standard was measured against reagent blank at 546nm

Calculation

Figure 2

Estimation Of Hdl-Cholesterol

Reagents

HDL-cholesterol precipitant (Phosphotungstic 4.8%, Magnesium chloride 3M)

Principle

Low density lipoproteins (LDL and VLDL) and chylomicron fractions are precipitated quantitatively by the addition of acid phosphotungstic in the presence of magnesium ions. After centrifugation the cholesterol concentration in the HDL (high density lipoprotein) fraction, which remains in the supernatant is determined.

Procedure

Into centrifuged tubes, 500µl of sample and 100µl of HDL-cholesterol precipitant was added, mixed and allowed to stand for 10mins at room temperature .The tubes were centrifuged at 4,000rpm.100µl of supernatant were assayed for cholesterol estimation with the procedure given above for total cholesterol estimation.

Calculation

Figure 3

Estimation Of LDL-Cholesterol

Estimation Of Triglyceride In Plasma

The enzymatic colorimetric method by McGowan was used ₇ .

Reagent

Pipes buffer pH 6.8 50mM, 4-chloro-phenol 4.2 mM, Magnesium aspartate 40 mM, 4-aminophenzone 0.35mM, ATP 2mM, lipase ≥9000 µ/l, Glycerol-Kinase ≥ 500µ/l, Glycerol-3-phosphate oxidase≥ 200µ/l, peroxidase ≥ 500µ/l.

Principle

Figure 4

Procedure

To 100µl of reagent in tubes meant for samples, standard and blank, 10µl of samples, standard and distilled water was added respectively. The content were mixed and incubated for 5mins at 37ºc.The absorbance of the samples and standard was measured against reagent blank at 546nm.

Calculation

Figure 5

Determination Of Malonydialdehyde, A Product Of Lipid Peroxidation

Estimation Of Plasma Phospholipids

Reagent

1. Chromogenic reagent

Solution i) 16g Ammonium molybdate was dissolved in 120ml distilled water.

Solution ii) 80ml of solution 1 was added to 40ml conc HCl and 10ml of mercury. The solution was shaken constantly for 30 minutes and filtered.

Solution iii) 200ml conc H₂SO₄ was added to the remaining solution i above cooled and added to the filtered solution ii. The chromogenic solution was obtained by adding 45ml methanol, 5ml chloroform and 20 ml distilled water.

2. Phospholipid standard; L- α phosphatidylcholine (1mg/ml)

3. Isopropanol

Principle

This involves complexing intact phospholipids with mercury-ammonium molybdate chromogenic reagent ₉ . The absorbance was measured spectrophotometrically at 710nm

Procedure

0.2ml plasma was added to 3.8ml isopropanol. The mixture was centrifuged for 10 minutes at 1000g. 0.5ml of supernatant was pipetted into centrifuged tubes and evaporated to dryness at 90 ° C.0.4ml chloroform and 0.1ml chromogenic solution were added after thorough mixing, the tubes were heated at 100 ° C for 1 minute. After cooling at room temperature, the tubes were mixed and 3ml chloroform was added. At the end of 15minutes, the tubes were centrifuged at 1000g for 5minutes and the absorbance of the supernatants read at 710nm. Isopropanol was used as blank.

Calculation

Figure 6

Statistical Analysis

Results

The plasma lipid concentration and malonydialdehyde were determined from a total of 130 subjects made up of normotensives, hypertensives on anti hypertensive drugs and hypertensives not on any anti hypertensive drugs and they were aged between 30-60 years.

Figure 7

Table 1: Plasma Lipid Values For Normotensives (MEAN ± SD)

Figure 8

Table 2: Plasma Lipid Values For Hypertensives On Drugs (MEAN ± SD)

Figure 9

Table 3: Plasma Lipid Values For Hypertensives Not On Any Anti-Hypertensive Drugs (MEAN ± SD)

Figure 10

Table 4: Statistical Comparisons (P-VALUES)

KEYS

BPS = Systolic Blood Pressure in mmHg
BPD = Diastolic Blood Pressure in mmHg
HDL = High Density Lipoprotein Cholesterol
LDL = Low Density Lipoprotein Cholesterol
HOD = Hypertensives on Drugs
HND = Hypertensive not on Drugs
n = Number of subjects investigated

Discussion

The result of the work has shown that there is some variation in plasma lipids in relation to sex. The total plasma cholesterol was significantly higher in females than in the males. There was a significant difference in the lipid concentration and malonydialdehyde concentration between normotensives, hypertensive on hypertensive drugs and those not on any anti-hypertensive drugs.

Total cholesterol, triglycerides and LDL cholesterol were significantly higher in hypertensives not on any drug therapy than normotensives (P< 0.001). There was a significant difference in blood pressures, total cholesterol, HDL-cholesterol and LDL-cholesterol in hypertensives on antihypertensive drugs when compared to hypertensives not on any antihypertensive drugs, but triglyceride concentration increased marginally in hypertensives on antihypertensive drug than hypertensive not on any anti-hypertensive drug. The reason may be attributed to the effect of antihypertensive drug on triglycerides.

There were high levels of total cholesterol, triglycerides and LDL-cholesterol and low levels of HDL-cholesterol were observed. There was no significant difference in the phospholipids concentration (P>0.05) of normotensives compared to hypertensives.

In our study, we observed an increase in malonydialdehyde, a product of lipid concentration among the normotensives, hypertensives on anti-hypertensives and hypertensives not on any anti hypertensive drug. However there was no significant difference observed in the malonydialdehyde concentration of hypertensives on anti-hypertensive drugs compared to those not on any anti-hypertensive drugs. This may be attributed to the fact that anti-hypertensive drugs may not have any effect on lipid peroxidation.

Conclusion

The results obtained from this study indicate that lipid concentration are increased in hypertensive patients and if not controlled could lead to other cardiovascular disorders or possibly death. Regular laboratory investigations of lipid profile should be investigated along with blood pressure in hypertensives since lipid and lipoproteins are important diagnostic and predictive value in coronary heart diseases.

Correspondence to

Patrick Igbinaduwa, Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Benin, Benin City. Phone: +2348037422420 Email: pigbinaduwa@yahoo.com