The serum phospholipid levels and total cholesterol: phosopholipid ratios of apparently normal adult Filipino students

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Part of Acta Medica Philippina

Title: The serum phospholipid levels and total cholesterol: phosopholipid ratios of apparently normal adult Filipino students
Date: 1958
extracted text: THE SERUM PHOSPHOLIPID LEVELS AND TOTAL CHOLESTEROL: PHOSPHOLIPID RATIOS OF APPARENTLY NORMAL ADULT FILIPINO STUDENTS * • This atudy was supported in part by a grant-in-aid from the U.P. Natural Science Research Center. SO UTA F. CAMAKA-BESA, M.D, M.S. (Biechem.) and JESUS L. CHAN-YUKOCa M.D. Department of Biochemistry Coileye of Medicine, University of the Philippines The interrelationships of serum cholesterol and phospholi pids in health and in coronary heart disease have drawn con siderable interest among many investigators and the work done on the subject from 1950 to July. 1957 has been summarized in an excellent review by Mattill (1). Aspects of the problem that have attracted our interest are the reported findings that the serum total cholesterol: phospholipid ratio, usually designated as the C :P ratio, has been shown to be more constant in a given individual than the actual cholesterol level and this C:P ratio tends to be elevated in patients with coronary heart disease. Some investigators feel that the C:P ratio may be more im portant than serum total cholesterol levels in the study of athe rosclerosis because it has been contended that “normal C :P ratio is a prerequisite for a normal vascular system even if the abso lute concentrations are higher than normal” (1). It was therefore felt that a study of the serum phospholipid levels and the serum cholesterol: phospholipid ratios in Filipino subjects would provide valuable information in addition to those secured in studies we have conducted (2) and are still conduct ing on serum cholesterol levels in this country. Correlation studies were also done on C :P ratio and relative body weight, C :P ratio and 'serum total cholesterol level, serum phospholipid level and amount of total fat habitually taken at breakfast, and serum phospholipid level and amount of coconut oil contained in the same meal. 51 52 ACTA MEDIC A PHILIPPINA MATERIAL AND METHODS The subjects of this study were 66 male and 30 female first year medical students aged from 18 to 24, averaging 19.9 years. They had no overt disease revealed by a physical and medical examination and their blood pressure readings were below 140 nun. Hg. systolic and 90 mm. Hg. diastolic. Detailed dietary diaries were kept for seven days by the subjects. Blood samples were obtained by venipuncture between 2 to 8 hours after breakfast. These were allowed to clot and the serum obtained. Serum total cholesterol was determined on duplicates by the method of Abell st al. (3). The serum phos pholipids were determined on duplicates, on the same day as the blood withdrawal, by the method of Youngburg (4) which em ploys the phosphate procedure of Fiske and Subbarow. RESULTS AND DISCUSSION Serum phospholipids. The serum phospholipids of the 66 male subjects ranged from 120 to 310 mg. per 100 ml. with a mean of 192.4 + 36.6 mg. per 100 ml. (S.D.) Those of 30 female subjects ranged from 110 to 332.5 mg. per 100 ml. with a mean of 207 + 54.8 mg. per 100 ml. <S.D.) Statistical analy sis showed that the difference of 14.6 mg. per 100 ml. between the mean values of the 2 sexes is not significant (t = 1.454). Likewise, Peters and Man {5) did not find any significant dif ference between the serum phospholipids of normal male and female adults. Comparing our results with findings abroad, we find that our values are comparable to those of Kornerup in Denmark (6), but are lower than those of Russ et al. (7) and of Peters and Man (5), and Gertler et al. (8) on American subjects. These are summarized in the following table. SERUM PHOSPHOLIPID AND CHOLESTEROL 53 COMPARISON OF SERUM PHOSPHOLIPID VALUES AND CHOLES TEROL: PHOSPHOLIPID RATIOS FOR ‘'NORMALS” OF THIS SERIES AND THOSE OF WORKERS ABROAD. No. of subjects Serum phospholipid in mg. per 100 ml. Serum total cholesterol: phospholipid ratio Range Mean + S.D. Range Mean Filipinos This series 96 110-332.5 19? ± 45.8 0.564-1.791 1.043 Gertler et al (Boston) Unselected normals 146 213-415 299.3 + 39.3 0.520-1.056 0.7408 Normals ‘’matched” to myocar* dial infarc tion points 90 221-397 305.7 + 39.8 U.5184-1.146 0.776 Peters and Man (New York) 103 152.5-362.5 230.2 ± 35.25 0.775-0.968 0.8556 Russ et al. (New York) 38 187-327 253 0 75-1,14 0.95 Koj*nerup (Denmark) 104 80-260* 130 + 40.0 0.736-2.818* • 1.227** • Range = mean + 2 S.D. * • Computed from data given in paper. q4 ACTA MEDICA PHIL1PMKA The frequency distribution of the serum phospholipid values of the 96 subjects of this study is shown in Figure 1. SERUM PHOSPHOLIPID (MG. PEE )0n ML.) Figure ). Frequency distribution of serum phospholipid values in 96 male and female Filipino students SERUM PHOSPHOLIPID AND CHOLESTEROL 55 Cholesterol: Phospholipid ratios. The C :P ratios in our 66 male subjects varied from 6.564 to 1.469, with a mean of 1.041 + 0.19 (S.D.). In the 30 female subjects, the C :P ratios ranged from 0.660 to 1.791, averaging 1.049 + 0.274 (S.D.). Although the female subjects gave a higher range and mean, the difference between means of the two sexes, 0.008, is not statistically significant (t = 0.157). The frequency distribu tion of the C:P ratios is shown in Figure 2. The mean for the CHOLESTEROLiPHOSPHOLIPID RATIO Figure 2. Frequctncy (list ri hut ion of cholesterol: phospholipid ratios nf DC male and female Filipino uladvnts. 56 ACTA MED1CA PHI LI PPI NA Comparing with findings of other workers shown in the table, the C:P values of our Filipino subjects are strikingly higher than those obtained by American investigators. We com puted the C :P ratios from data given by Kornerup in his paper and found that they were also higher in his Danish subjects than those of Americans. However, the factor of differences in analytical methods used for cholesterol and phospholipid deter minations must be remembered in making this comparison. Se rum cholesterol was determined by means of the LiebermanBurchard reaction after extraction by varying methods in all the investigations included in the table except in that of Peters and Man where the gravimetric method was employed. Serum phospholipid was extracted and digested in different ways and the P analyzed by the colorimetric method of Fiske and Subbarow, in this series and those of Gertler et at. and Russ et al. Kornerup employed a modification of the method of Fiske and Subbarow using amidol instead of aminonaphthol sulfonic acid in the color reaction. In al) the reports being compared here, the lipid P value was multiplied by 25 to get the phospholipid level. It is nevertheless of interest to mention that our mean for this series of normal adults is 1.043 which is the value (1.04) quoted from Oliver and Boyd (1) as the mean of their 50 pa tients with myocardial infarction. It is also higher than those of 60 patients with coronary disease obtained by Gertler et al. (8), to wit: 0.578 to 1.32, with a mean of 0.894. Gertler et al, (8) attribute the importance of the phospho lipid in maintaining the other lipids in solution to the fact that it is a colloid stabilizer. Since phospholipids are hydrophilic, they tend to keep the hydrophobic cholesterol in solution, much like the action of lecithin and bile salts in holding the cholesterol of the bile in suspension. They observed that in normal indi viduals, a rise of serum cholesterol is usually accompanied by a concomitant rise in the serum phospholipids, while in coronary artery disease, the interrelationships between the lipids are dis turbed and the phospholipid does not rise pari passu with the cholesterol. This was corroborated by Steiner et al. (1) in his study comparing 82 coronary infarction patients with 112 SERUM PHOSPHOLIPID ANO CHOLESTEROL 57 healthy controls. They found twenty-five of the coronary groups with cholesterol values within the normal range but with low phospholipid levels resulting in high C:P ratios. Correlation studies. A contributory factor to the high C:P ratios is the fact that the serum phospholipid levels in this series are lower on the average than those reported in the li terature for normal groups. The lower serum phospholipid va. lues were obtained in spite of the fact that the blood samples were withdrawn not in the fasting state but 2 to 3 hours after breakfast. It has been claimed in the literature (6) that unlike cholesterol, phospholipid levels are affected by the precedent meal and rise in the blood to a maximum within four or five hours after a mea! rich in fat. This finding made us look into the quantities of fat habi tually taken by the subjects at breakfast from data gathered through their individual seven-day dietary diaries. We found that total fat of the daily breakfast ranged from 11.8 to 75.2 Gm., with a mean of 29 + 13.4 Gm. <S.D.) This shows that as a group, our subjects took breakfast with moderately high fat content. There was no correlation between the serum phospho lipid level and the total fat intake at breakfast in the subjects of either sex. The statistical data are as follows: * * For thio nnd subsequent correlation studies the symbols used are: n - No. of oases = Standard error of re gression coefficient b = Regression coefficient t = h/sh Male n = 66 b = 0.58 mg. serum phosphilipid per 100 ml. per gram total breakfast fat S, - 0.344 t — 1.682 Female n = 30 b = — 0.303 mg. serum phospholipid per 100 ml. per gram total breakfast fat S, = 1.189 t '=0.255 It was thought of interest to determine the amount of co conut fat taken habitually at breakfast for several reasons. Blood phospholipids normally contain essential fatty acids (9). Because of the low content of such fatty acids in coconut oil. 58 ACTA MEDICA PHILIPP1NA could the common use of coconut oil in food preparation in this country have been a factor in the production of low levels of serum phospholipids in this series? Coconut oil has consistently been reported as a highly saturated fat with only 1 to 2 per cent linoleic acid content (10,11). The coconut oil content of the daily breakfast varied from 1,0 to 16.9 Gm., with a mean of 7.04 + 3.64 Gm. (S.D.) No correlation was found between the serum phospholipid levels and the amount of coconut oil expressed in grams, taken at break fast among the male subjects. However, the female subjects ex hibited a significant decrease of 9.03 mg. serum phospholipids per 100 ml. per gram increase in the coconut oil intake at breakfast. The statistical data are as follows: Male n = 66 b = — 0.106 mg, serum phospholipid per 10O ml. per gram coco nut oil taken at breakfast S„ = 1.315 t =0.081 Female n = 30 b = — 9.03 mg. serum phospholipid per 190 ml. per gram coco nut oil taken at breakfast S„ = 3.559 t = 2.537 Expressing the coconut oil content as per cent of the total fat at breakfast, the range was from 3.7 to 62,6 per cent with a mean of 24.7 per cent among the female subjects. A decrease of 2.08 mg. per 100 ml. serum phospholipid accompanied a rise of 1 per cent in the coconut oil content of the total breakfast fat, significant at the 2 per cent level (t = 2.66), Among the male subjects a corresponding decrease of 0.26 mg. per 100 ml. serum phospholipid was not found to be significant (t = 0,59). The coconut oil constituted from 5.7 to 53.8 per cent with a mean of 26.3 per cent for the male subjects. The influence of body build on the serum lipid level has been brought out in the literature (3, 8). In this series only relative body weights were studied. No correlation was found SERUM PHOSPHOLIPID AND CHOLESTEROL 59 between C:P ratios and relative body weight expressed as per cent of the standard weight for Filipinos of the corresponding height and age {12). The statistical data are as follows: Male Female n = 66 n = 30 b = — 0.0013 units C :P pet b = — 0.0042 units C :P per 1 per cent relative 1 per cent relative body weight body weight Sh = 0.0022 S„ = 0.0053 t'= 0.575 t’= 0.802 In this series, the C :P ratios rose as the serum cholesterol level increased. This means that the serum phospholipid level did not increase at the same rate as the serum cholesterol. A similar trend has been reported by Peters and Man (5), although their different means of C:P ratios for increasing levels of serum cholesterol are below the values obtained in this study. Among our 66 male subjects, there was found to be a rise of C :P value of 0.0017 per mg. per 100 ml. rise in serum cholesterol, significant at the 1 per cent level (t = 2.916). In the 30 female subjects, the rise of C :P ratio was 0.0035 per mg. per 100 ml. rise in serum cholesterol, significant at the 5.5 per cent level (t = 2.034). The scattergrams and regression line calculated by the least squares method are shown in Figures 3 and -1. 60 ACTA MEDICA PHILIPP1NA n>m SERUM PHOSPHOLIPID AND CHOLESTEROL 61 OL: PHOSPHOLIPID RATIOS SERUM CHOLESTEROL (MG. PER 100 ML.) Figure 4. Relation between scrum cholesterol: phospholipid ratio and sc rum total cholesterol level in 30 female Filipino students. 62 ACTA MEDIC A PHtLIPPINA SUMMARY 1. The non-fasting serum phospholipid values of 96 ap parently healthy adult male and female subjects 18 to 24, averaging 19.9 years old and habitually taking.break fast with moderately high fat content (mean 29 Gm, 25 per cent of which was coconut oil) varied from 110 to 332.5 mg. per 100 ml. serum, with a mean of 197 + 45.8 mg. ner 100 ml. (S.D.) No significant sex difference was found. 2. The serum cholesterol: phospholipid ratios ranged from 0.564 to 1.791, with a mean of 1.043 + 0.218 (S.D.). These are high compared to reports in the literature, so much so that our mean is the same as the 1.04 given bv Oliver and Boyd (1) for their subjects with myocar dial infarction. No significant sex difference in C:P ratios was obtained in our study, 3. No correlation was found between the serum phosnholipid level and the total grams of fat taken at breakfast. However, expressed as grams or as per cent of the total fat, there was observed a significant decrease of serum phospholipid levels with an increase in the coco nut oil content of breakfast among the 30 female sub jects. No correlation was obtained among the 66 male subjects. 4. No correlation was found between C :P ratio and relative body weight in both sexes. 5. The C :P ratio rose significantly with the level of serum cholesterol in the subjects of both sexes: 0.0017 per mg. per 100 ml rise of serum cholesterol in the males, and 0.0035 per mg. per 100 ml. in the females. ACKNOWLEDGMENT The authors are indebted to Misses Lourdes Ciudadano and Minerva Bataclan for their technical assistance and to the personnel of the Institute of Nutrition who gave the necessary guidance in the dietary fat studies. Computation of fat content was done with the aid of the Food Composition Table, Institute of Nutrition Handbook, prepared by the staff of the Foods and Nutritional Biochemical Division, June. 1957. SERUM PHOSPHOLIPID AND CHOLESTEROL S3 The authors are also grateful to the members of Class 1963 of the College of Medicine, University of the Philippines who co operated as subjects in this study and to Mr. Anselmo Limbo of the Department of Biochemistry who helped in the preparation of this manuscript. REFERENCES 1. MATTILL, H. L: Cholesterol as related to atherosclerosis, a review of the literature, 1950 to July, 1957. V. Relation between serum cholesterol and phospholipid. Cereal Institute, Inc., 135 South La Sa Ho Street, Chicago, 3. 2. CaMARA-RESA, S. F ? A study of serum cholesterol levels in rela tion to diet and some physical and biochemical variables in the Armed Forces of the Philippines. J. Phil. Med. Assoc. 75:137-152 (March), 1959, 3. ABELL, L. L., LEVY, R. B., BRODIE, B. B. and KENDALL, F, E.: A simplified method for the estimation of total cholesterol in serum and demonstration of its specificity, J. Bio. Chem. 105:357, 1952. 4. HAWK, P. B., OSER, B. L„ and SUMMERSON, W. H.: Practical Physiological Chemistry, 1954. The Blakiston Co., Ine., New York and Toronto, Thirteenth Edition, pp. 589'590. S. PETERS, J. P. and MAN, E. B,: The interrelations of serum lipids in normal persons. Arch. I nt. Med. 55:398-415 (March), 1950. 7. RUSS, E. M., EDER, H. A. and BARR, D. P,; Protein-lipid rela tionships in human plasma. The Amer, Jour, of Mod. 11:4G8-469 (October), 1951. 8. GERTLER, M. M., GARN, S. M. and LEHRMAN, J.: The interrela tionships of serum cholesterol, cholesterol esters and phospholipids in health and in coronary artery disease. Circulation 2:225-214 (August), 1950. 9. VAN JTALLIE, T. B.: Dietary fats and atherosclerosis. Nturition RcV. 15:1-6 (January), 1957. 10. MALM ROS, H, utid WIGAND, G.: The effect on serum cholesterol of diets containing different /ats. Lancet 2:1-7 (July 6), 195?. 11. AHRENS, E. H., HIRSCH, J.T INSULL, W., JR.., TSALTAS, T. T., BLOMSTRAND, R. and PETERSON, M, L.: The influence of dietary fats on scrum lipid levels in man. Lancet 1:943-953 (Moy 11), 1957, 12. CORDERO, N.. JAVIER, B. N., PAULINO-GONZALEZ, C. M„ BAGA BALDO, 7.. and RODR1GUEZ-BUSUEGO, M.s Philippine phy siological standards: body weight in relation to height and age for adults. Acta Med. Phil. 15:417-440. 1956-1957,
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