1. Evidence is Insufficient to Support A Role for Chocolate or Cocoa in Increasing High Density Lipoprotein Cholesterol (HDL-C) in Healthy Normal Weight or Overweight Adults Abstract High density lipoprotein cholesterol (HDL-C) is a risk factor for coronary heart disease (CHD). There is an indication that cocoa and chocolate products increase HDL-C level. The purpose of this evidence analysis is to evaluate whether there is strong evidence to support this relationship in healthy normal-weight and overweight adults. Eleven randomized controlled trails (RCT) were identified in which cocoa, chocolate, or a combination were an intervention and reported HDL-C level as a dependent factor. Each study was reviewed by four reviewers using the Evidence Analysis Manual adopted by the Evidence Analysis Library of the American Dietetic Association. Evidence is insufficient that daily chocolate and cocoa consumption increases HDL-C in healthy normal-weight and overweight adults (grade II). Only four (three neutral and one negative quality) studies observed a significant increase in HDL-C compared to control groups. Cocoa and chocolate may elicit this effect by increasing the level of Apolipoprotein A1 (ApoA1) production. Lack of positive results in the remaining studies might be explained by selection bias, normal baseline HDL-C level, or small sample sizes. Nevertheless, cocoa and chocolate may have other cardiovascular benefits, or at least have a neutral effect on lipids, and can be incorporated in the diet without weight gain. Future research that focuses on HDL-C level in relation to other lipids and in people with low HDL-C might lead to different conclusions. Table 1. A Summary of the evidence analysis results. Articles listed in the order they appear in Figure 2. Author, year (reference), quality rating Study Participants/ Treatment Duration InterventionChange in HDL-C (significance level set at p=0.05) Shiina, 2009, (6) neutral N=39 males. Japanese. Average age= 29.8 14 days 45 g dark chocolate vs. control (35 g white chocolate) No significant change in HDL-C from baseline or difference in HDL-C at the end of study between groups Engler, 2004, (7) neutral N=11 males, 11 females. Caucasian, Asian, Pacific Islander. Age=21-55 14 days 46 g high-flavonoid dark chocolate vs. 46 g low-flavonoid dark chocolate No significant change in HDL-C from baseline in either group Kurlandsky, 2006, (8) positive N=52 females. Age=22-65 42 days 41 g dark chocolate vs. 60 g almonds vs. dark chocolate and almonds, vs. control HDL-C levels were not significantly affected by any of the study products Fraga, 2005, (9) neutral N=28 male soccer players. Latin. Average age=18 14 days 105 g milk chocolate vs. 105 g cocoa butter chocolate No significant change in HDL-C level between baseline and end of study Kris-Etherton, 1994, (10) neutral N= 55 males. Age=21-35. 27 days 46 g milk chocolate bar vs. high carbohydrate snack (pretzels with soft drink) HDL-C level higher in milk chocolate bar compared to high carbohydrate snack (0.08 ± 0.03 mmol/L, P < 0.01) Joo, 1997, (11) neutral N=7 males, 8 females. Age=21-48 7 days 85 g of: milk chocolate vs. dark chocolate vs. white chocolate vs. imitation chocolate vs. imitation white chocolate vs. carob milk bar Compared to baseline, serum HDL-C significantly increased only in the milk chocolate and the carob milk groups but not in other groups (p<0.05) Baba, 2007, (12) neutral N=25 males. Japanese. Age= 38  1 48 days 26 g cocoa powder with 12 g sugar vs. control (12 g sugar) HDL-C increased by 23.4% in cocoa group from baseline compared to 5.1% in control group. The difference in increase between the two groups was significant (p<0.001) Wan, 2001, (13) neutral N= 10 males, 13 females. Average age=36. Mostly Caucasian 28 days 22 g cocoa powder and 16 g dark chocolate vs. control (no cocoa or chocolate) At the end of the study, serum HDL-C in cocoa and dark chocolate group was significantly greater by 4% than the control group (p=0.02) Crews, 2008, (14) positive N= 38 males, 52 females. Age= over 60, average age=68 42 days 8 oz cocoa drink and 37 g dark chocolate vs. placebo The change in HDL-C from baseline in the chocolate and cocoa group was not significantly different from the change in the control group (p=0.902) Davison, 2008, (15) positive N= 17 males, 32 females. Age=18-65. Overweight 84 days High-flavanol cocoa drink vs. low-flavanol cocoa drink vs. high- flavanol cocoa drink with exercise vs. low-flavanol cocoa drink with exercise The change in HDL-C between baseline and end of study was not significant in any of the groups. No significant 3-way interaction (cocoa x exercise x time) (p=0.17) Njike, 2011, (16) positive N=6 males, 33 females. Average age=52.2. Mostly Caucasian 42 days 22 g sugar-free cocoa drink vs. 22 g cocoa drink with 91 g sugar vs. placebo (110 g sugar) The change in HDL-C between baseline and end of study in either cocoa groups was not significantly different from the change in the placebo group Results Eleven articles met the inclusion criteria. The results are summarized in Figure 2 and Table 1. Methods Electronic and manual searches were conducted between August 1, 2011 and September 9, 2011 using eight search combinations in two databases (Figure 1). Inclusion criteria were limited to RCT; healthy human adults of normal weight or overweight; cocoa, or chocolate, or a combination as the independent factor; and HDL-C level as a dependent factor. Exclusion criteria were studies written in languages other than English; interventions that used fortified products or extracts; or participants with hypertension, diabetes, dyslipidemia, or other chronic conditions. No restriction was made on publication date. Chocolate and HDL Chocolate and cholesterol Chocolate and lipoprotein Chocolate and cardiovascular risk Cocoa and HDL Cocoa and cholesterol Cocoa and lipoprotein Cocoa and cardiovascular risk Figure 1. Search term combinations. The analysis followed methods adopted by the Evidence Analysis Library of the American Dietetic Association detailed in the Evidence Analysis Manual, 2010. Four reviewers independently reviewed each study then came to consensus on its rating Discussion Evidence is insufficient that daily chocolate and cocoa consumption increases HDL-C in healthy normal-weight and overweight adults (grade II). Only four of the eleven studies found statistically significant changes in HDL-C (10-13). Kris-Etherton (10) substituted a carbohydrate snack with milk chocolate or another carbohydrate snack, which agrees with evidence that substituting carbohydrates with fats improves HDL-C levels (3). While Joo (11) found that milk chocolate significantly increased HDL-C, the study received negative rating because the amount of chocolate, statistical analysis, and recruitment methods were unclear. Baba (12) found that a cocoa drink increases HDL-C significantly compared to a sugar drink. This neutral study also lacked information on participant selection, statistics, and specifics of the intervention. Wan (13) found HDL-C to be greater by 4% compared to the control group. However, baseline HDL-C was reported for all participants and not for each group separately, thus baseline levels may not have been similar. Chocolate is gaining a healthy food reputation. Stearic acid, a SFA and the main fatty acid in chocolate, has a neutral effect on total cholesterol and LDL-C (2, 17), possibly because it converts to oleic acid in the body. Other SFAs, such as lauric, myristic, and palmitic, raise HDL-C and LDL-C concurrently. Furthermore, cocoa and chocolate contain more flavonoids (potent antioxidants) than black tea, red wine, cranberry juice, or apples (18). Cocoa may raise HDL-C level by increasing ApoA1 production, the main protein in HDL-C particles. In one in vitro study, human hepatic HepG2 and intestinal Caco2 cells were incubated with cocoa polyphenols. After 24 hours, ApoA1 level increased (19). This hypothesis may be supported by another study that found increased expression and production of ApoA1 in a human hepatic HepG2 cells by genistein, a flavonoid in soy beans and coffee (12). There are several potential explanations for the lack of association between cocoa or chocolate and HDL-C in the remaining studies (6-9, 14-16). Recruitment, inclusion/exclusion, and demographic information were not reported adequately, raising the possibility of selection bias. Baseline HDL-C was <40 mg/dL in one study (9), and normal (6-8, 10, 12-16) or unreported (11) in the remaining. It may not be possible to increase HDL-C beyond the normal range. Sample sizes were small, and power was not calculated to determine the number needed to observe a significant change in HDL-C, since it was not the main outcome in some studies. Cocoa and chocolate may have other cardiovascular benefits. Their consumption improved endothelial function (7, 15, 16), increased coronary flow velocity reserve (6), reduced LDL-C oxidation (12, 13), reduced stress markers (12), and lowered serum triglycerides (8, 10). Total cholesterol and LDL-C were either lowered (9, 12) or remained unchanged (6-8, 10, 14-16). All eleven studies reported no weight gain. Introduction Coronary heart disease is the most common type of cardiovascular disease, which affects more than 16 million American adults (1). In 2007, one out of six deaths in the United States was attributed to CHD, and in a given year, 34% of those who experience a coronary attack will die from it (1). There is an inverse relation between HDL-C level and the risk of a CHD event. A low HDL-C level— less than 40 mg/dL in men and women—is a strong independent predictor of CHD (2). A 1-mg/dL increase in HDL-C significantly lowers the risk for CHD by 2% in men and 3% in women (3). It is well established that the diet affects HDL-C level (4). Very high intake of carbohydrates (> 60% of calories) and consumption of trans fatty acids lower HDL-C, while diets high in unsaturated fatty acids and soy protein raise it (2). Saturated fatty acids (SFA) may or may not affect HDL-C level (2). The effect of chocolate on HDL-C level is inconclusive. There’s an indication that cocoa and chocolate may raise HDL-C; however, a recent meta-analysis found otherwise (5). Therefore, the purpose of this analysis is to examine the strength of the evidence that chocolate and cocoa consumption increases HDL-C in healthy normal-weight and overweight adults. References 1.Roger VL, Go AS, Lloyd-Jones DM, Adams RJ, Berry JD, Brown TM, et al. Heart Disease and Stroke Statistics 2011 Update: A Report from the American Heart Association. Circulation. 2011;123:e18-e209. 2.Expert panel on detection, evaluation, and treatment of high blood cholesterol in adults. Executive summary of the third report of the National Cholesterol Education Program (NCEP) expert panel on detection, evaluation, and treatment of high blood cholesterol in adults (Adult Treatment Panel III). JAMA. 2001;285:2486-2497. 3.Gordon DJ, Probstfield JL, Garrison RJ, Neaton JD, Castelli WP, Knoke JD, Jacobs Jr. DR, Bangdiwala S, Tyroler HA. High-density lipoprotein and cardiovascular disease, four perspective American studies. Circulation. 1989;79:8-15. 4.Van Horn L, McCoin M, Kris-Etherton PM, Burke F, Carson JS, Champagne CM, Karmally W, Sikand G. The evidence for dietary prevention and treatment of cardiovascular disease. J Am Diet Assoc. 2008;08;287-331. 5.Jia L, Liu X, Bai YY, Li SH, Sun K, He C, Hui R. Short-term effect of cocoa product consumption on lipid profile: a meta-analysis of randomized controlled trials. Am J Clin Nutr. 2010 Jul;92(1):218-25. 6.Shiina Y, Funabashi N, Lee K, Murayama T, Nakamura K, Wakatsuki Y, Daimon M, Komuro I. Acute effect of oral flavonoid-rich dark chocolate intake on coronary circulation, as compared with non-flavonoid white chocolate, by transthoracic Doppler echocardiography in healthy adults. Int J Cardiol. 2009;131:424-429. 7.Engler MB, Engler MM, Chen CY, Malloy MJ, Browne A, Chiu EY, Kwak HK, Milbury P, Paul SM, Blumberg J, Mietus-Snyder ML. Flavonoid-Rich Dark Chocolate Improves Endothelial Function and Increases Plasma Epicatechin Concentrations in Healthy Adults. J Am Coll Nutr. 2004;23:197-204. 8.Kurlandsky SB, Stote KS. Cardioprotective effects of chocolate and Almond consumption in healthy women. Nutr Res. 2006;26:509-516. 9.Fraga CG, Actis-Goretta L, Ottaviani JI, Carrasquedo F, Lotito SB, Lazarus S, Schmitz HH, Keen CL. Regular consumption of a flavanol-rich chocolate can improve oxidant stress in young soccer players. Clin Dev Immunol. 2005;12:11-17. 10.Kris-Etherton PM, Derr JA, Mustad VA, Seligson FH, Pearson TA. Effects of a milk chocolate bar per day substituted for a high-carbohydrate snack in you men on an NCEP/AHA Step 1 Diet. Am J Clin Nutr. 1997;60:1037S-1042S. 11.Joo SJ, Kies C, Schnepf MS. Chocolate and chocolate-like products: Impact on serum lipid profiles and serum fatty acid composition of humans. J Appl Nutr. 1997;49:46-55. 12.Baba S, Osakabe N, Kato Y, Natsume M, Yasuda A, Kido T, Fukuda K, Muto Y, Kondo K. Continuous intake of polyphenolic compounds containing cocoa powder reduces LDL oxidative susceptibility and has beneficial effects on plasma HDL-cholesterol concentrations in humans. Am J Clin Nutr. 2007;85:709-717. 13.Wan Y, Vinson JA, Etherton TD, Proch J, Lazarus SA, Kris-Etherton PM. Effects of cocoa powder and dark chocolate on LDL oxidative susceptibility and prostaglandin concentrations in humans. Am J Clin Nutr. 2001;74:596-602. 14.Crews Jr WD, Harrison DW, Wright JW. A double-blind, placebo-controlled, randomized trial of the effects of dark chocolate and cocoa on variables associated with neuropsychological functioning and cardiovascular health: clinical findings from a sample of healthy, cognitively intact older adults. Am J Clin Nutr. 2008;87:872-880. 15.Davison K, Coates AM, Buckley JD, Howe PRC. Effect of cocoa flavanols and exercise on cardiometabolic risk factors in overweight and obese subjects. Int J Obes. 2008;32:1289-1296. 16.Njike VY, Faridi Z, Shuval K, Dutta S, Kay CD, West SG, Kris-Etherton PM, Katz DL. Effects of sugar-sweetened and sugar-free cocoa on endothelial function in overweight adults. Int J Cardiol 2011;149:83-88. 17.Kris-Etherton PM, Mustad VA. Chocolate feeding studies: a novel approach for evaluation the plasma lipid effects of stearic acid. Am J Clin Nutr. 1994;60:1029S-1036S. 18.Steinberg FM, Bearden MM, Keen CL. Cocoa and chocolate flavonoids: Implications for cardiovascular health. J Am Diet Assoc. 2003;103:215-223. 19.Yasuda A, Natsume M, Osakabe N, Kawahata K, Koga J. Cacao Polyphenols Influence the Regulation of Apolipoprotein in HepG2 and Caco2 Cells. J Agric Food Chem. 2011;59:1470-1476. Conclusion The current body of evidence is insufficient to support cocoa or chocolate’s role in increasing HDL-C level in healthy normal weight or overweight adults. Future research should focus on HDL-C as the main outcome and how it is related to other lipids and lipoproteins. In addition, research should focus on individuals with low HDL-C levels to determine if HDL-C increases with chocolate or cocoa consumption in this group. 33 Studies Identified 24 RCTs 11 RCTs Healthy Adults 9 Not RCT 4 diabetic 4 hypercholesterolemic 4 hypertensive 1 people with CAD 4 diabetic 4 hypercholesterolemic 4 hypertensive 1 people with CAD 9 RCTs in Normal Weight 2 RCTs in Overweight 3 dark chocolate (6-8) 2 milk chocolate (9, 10) 1 cocoa powder (12) 2 dark chocolate and cocoa powder as one treatment (13, 14) 1 milk or dark chocolate as two different treatments (11) 2 cocoa powder (15, 16) Figure 2. Article inclusion and exclusion. Articles grouped based on the type of cocoa or chocolate used for the intervention. The numbers in parenthesis reflect the reference number.