Doi:10.1016/j.nutres.2008.01.004

Available online at www.sciencedirect.com Brazil nut ingestion increased plasma selenium but had minimal effects on lipids, apolipoproteins, and high-density lipoprotein function Célia C. Strunza, Tatiane V. Oliveiraa, Juliana C.M. Vinagrea, Adriana Limab, Silvia Cozzolinob, Raul C. Maranhãoa,b,⁎ a Lipid Metabolism Laboratory, the Heart Institute (InCor) of the Medical School Hospital, São Paulo, Brazil b Faculty of Pharmaceutical Sciences, University of São Paulo, São Paulo, Brazil Received 1 August 2007; revised 21 December 2007; accepted 16 January 2008 The Brazil nut (Bertholletia excelsa) of the Amazon region is consumed worldwide. It is rich in both monounsaturated fatty acids and polyunsaturated fatty acids and is known for its high seleniumcontent. This study tested the hypothesis whether the consumption of this nut could affect the plasmalipids and apolipoproteins and some functional properties of the antiatherogenic high-densitylipoprotein (HDL). Fifteen normolipidemic subjects aged 27.3 ± 3.9 years and with body mass indexof 23.8 ± 2.8 kg/m2 consumed 45 g of Brazil nuts per day during a 15-day period. On days 0 and 15,blood was collected for biochemical analysis, determination of HDL particle size, paraoxonase1 activity, and lipid transfer from a lipoprotein-like nanoparticle to the HDL fraction. Brazil nutingestion did not alter HDL, low-density lipoprotein cholesterol, triacylglycerols, apolipoproteinA-I, or apolipoprotein B concentrations. HDL particle diameter and the activity of antioxidativeparaoxonase 1, mostly found in the HDL fraction, were also unaffected. Supplementationincreased the reception of cholesteryl esters (P b .05) by the HDL yet did not alter the reception ofphospholipids, free cholesterol, or triacylglycerols. As expected, plasma selenium was significantlyincreased. However, the consumption of Brazil nuts for short duration by normolipidemic subjectsin comparable amounts to those tested for other nuts did not alter serum lipid profile. The onlyalteration in HDL function was the increase in cholesteryl ester transfer. This latter finding may bebeneficial because it would improve the nonatherogenic reverse cholesterol transport pathway.
2008 Elsevier Inc. All rights reserved.
Humans; Cholesterol; Dietary selenium; Lipoproteins; Nanoparticles; Emulsions Apo A-I, apolipoprotein A-I; Apo B, apolipoprotein B; HDL, high-density lipoprotein; LDL, low-density lipoprotein; MUFA, monounsaturated fatty acids; PON1, paraoxonase 1; PUFA, polyunsaturated fatty acids; SFA,saturated fatty acids.
Epidemiological studies indicate that frequent consump- tion of nuts may decrease the risk of coronary artery disease. A lipid-lowering effect has been documented in studies ⁎ Corresponding author. Laboratório de Metabolismo de Lípides with almonds , walnuts , pecans pistachios Instituto do Coração do HC-FMUSP, São Paulo, Brazil, Tel.: +55 11 and peanuts consumed in daily amounts ranging from 0271-5317/$ – see front matter 2008 Elsevier Inc. All rights reserved.
doi: C.C. Strunz et al. / Nutrition Research 28 (2008) 151–155 Brazil nut (Bertholletia excelsa) found in the Amazon Participants had not ingested Brazil nuts for at least the region in South America is consumed worldwide. As a distinct characteristic, this fruit has a high-selenium All participants were simultaneously included in the content . Selenium is found in the active site of many study protocol that consisted in the consumption of 45 g/ enzymes such as thioredoxin reductase, which catalyzes d of Brazil nuts, roughly 11 nuts, for a 15-day period.
oxidation/reduction reactions and is essential for the The nuts were consumed as snacks or with meals in synthesis if glutathione peroxidase that prevents oxidative salads. On the last day of ingestion, the dietary inquiry processes . Diminished plasma selenium levels have was once again applied to verify whether or not nut con- been associated with increased of risk of coronary artery sumption had altered the composition of the diet. Blood samples were drawn after a 12-hour fast on the first and last With regard to fat composition, the Brazil nut is rich in day of the study for the biochemical determinations monounsaturated fatty acids (MUFA) and polyunsaturated fatty acids (PUFA) that have the potential of reducing As the nut consumption period was relatively short, it proatherogenic low-density lipoprotein (LDL) cholesterol was not necessary to design a control group without However, when compared with other nuts such as supplementation of Brazil nuts. The specified supplement of almonds, walnuts, pecans, pistachios, and peanuts, the Brazil 45g Brazil nuts used for the study contains 6.44 g of nut has a relatively higher content of LDL cholesterol– protein, 5.52 g of carbohydrates, and 29.89 g of total fat raising saturated fatty acids (SFA) . Brazil nuts are also (6.81 g of SFA, 11.06 g of MUFA, 9.26 g of PUFA), as well rich in magnesium and sulfur amino acids.
as 3.38 g of dietary fiber and 862.65 μg of selenium, for a To the best of our knowledge, the effects of Brazil nut consumption on plasma lipids have not yet been explored.
Because of the nutritional composition of this nut, the health 2.3. Plasma lipid and apolipoprotein determinations benefits of this food are of special interest. Few investiga- Plasma total cholesterol (CHOD-PAP; Roche, Basel, tions have evaluated the effects of selenium on plasma lipids CHE) and triacylglycerols (Triacylglycerol Rapid, Roche) with the interaction of PUFA, MUFA, and SFA in the Brazil were determined by enzymatic methods using a Cobas nut. Therefore, this study was designed to evaluate the Mira analyzer (Roche). High-density lipoprotein choles- effects of consumption of this nut at a level similar to other terol was measured after precipitation of the very low- edible nuts reported so to make an important density lipoproteins and LDL with HDL reagent (Roche; comparison. Some functional properties of high-density phosphotungsten acid/MgCl2 method) with automatic lipoprotein (HDL), HDL particle size, and the antioxidant equipment. Low-density lipoprotein C was estimated by enzyme paraoxonase 1 (PON1) that is associated with the the Friedewald formula Apolipoproteins A-I and B HDL fraction were also tested. These measurements are were determined by immunoturbidimetric assay (Roche) valuable in understanding the HDL antiatherogenic proper- ties that cannot be evaluated by simply measuring HDLcholesterol concentration.
2.4. High-density lipoprotein particle size The mean particle diameter of the HDL fraction was measured by laser-light scattering analysis with a ZetaPALSZeta Potential Analyzer (Brookhaven Instruments, Holts- ville, NY) after the separation of HDL from the plasma by Fifteen healthy volunteers (5 males and 10 females) chemical precipitation of the apo B–containing lipoprotein participated in the study. All were personnel of our laboratory, aged 27.3 ± 3.9 (23-34) years, and with body 2.5. Preparation of the labeled nanoemulsion for the lipid mass index 23.8 ± 2.8 (19.8-28.4) kg/m2. Inclusion criteria were plasma total cholesterol concentration of less than200 mg/dL, LDL cholesterol of less than 100 mg/dL, and The lipid nanoemulsion was prepared from a lipid triacylglycerols of less than 200 mg/dL. None were taking mixture of 40 mg cholesteryl oleate, 20 mg egg phospha- vitamin supplements, hypolipidemic agents, or any other tidylcholine, 1 mg triolein, and 0.5 mg cholesterol purchased type of medication. The study was approved by the ethics from the Sigma Chemical Company (St Louis, Mo). Lipid committee of the Heart Institute of the Medical School emulsification by prolonged ultrasonic irradiation in aqueous Hospital (São Paulo, Brazil), and an informed written media and a 2-step ultracentrifugation of the crude emulsion consent was obtained from each participant.
with density adjustment by addition of KBr to obtain thenanoemulsion was carried out by the method previously described and modified by Maranhão et al The Before the commencement of the study, the usual food nanoemulsion was dialyzed against a 0.9% NaCl solution.
intake of each participant was assessed by a dietary inquiry.
Trace amounts of cholesteryl [1-14C] oleate and glycerol tri C.C. Strunz et al. / Nutrition Research 28 (2008) 151–155 Corp., Tokyo) with generation of hydrides coupled with Estimate by dietary inquiry approach of the daily composition of the diet the cell of quartz (HGQTAAS), as described previously immediately before and on the last day of Brazil nut consumption (45 g/d) Results are expressed as means and standard deviations (mean ± SD). Differences of P b .05 were considered significant. All variables were compared using paired Student t test. Sigma Stat 3.11 for Windows software (Systat Software Inc, Calif) was used in the statistical calculations.
shows that, according to the dietary inquiry, energy intake increased by about 18% after the introduction Results are expressed as means ± SD (n = 15; 10 women and 5 men).
⁎ P b .05 compared to baseline diet (paired t test).
of the Brazil nut supplementation. This was due to the ⁎⁎ P b .01 compared to baseline diet (paired t test).
increase in the ingestion of MUFA, PUFA, and SFAcontained in the nuts.
[9,10(n)-3H] oleate or [7(n)-3H] cholesterol and shows that the participant body weight did not phatidylcholine, 1-stearoyl-2-[1-14C]arachidonyl (Amer- change at the end of the supplementation period and the sham, Little Chalfont, Buckinghanshier, UK) were added concentration of selenium in the serum increased 4-fold. also shows that the nut supplementation did not alter HDL orLDL cholesterol and triacylglycerol concentration. Apolipopro- 2.6. Assay for the lipid transfer from the donor tein A-I concentration, which together with HDL cholesterol is a marker for the HDL levels in the plasma, also remainedunchanged. There was, however, a trend for increased apo B Plasma with EDTA in a volume of 200 μL was incubated but did not attain statistical significance (P = .07).
with 50 μL of the nanoemulsion labeled with 14C-cholesterol As shown in , neither the HDL average particle and 3H-triacylglycerols or with 3H-cholesteryl esters and14 diameter nor the activity of the antioxidative enzyme PON 1 C-phospholipids. After a 1-hour shaking bath at 37°C, was affected by the nut supplementation. The ability of HDL 250 μL dextran sulfate/MgCl2 was added as a precipitation to simultaneously receive (normal lipoprotein metabolism reagent. The mixture was then mixed for 30 seconds and for lipid transfer enzymes) the 4 major lipid classes from the centrifuged for 10 minutes (3000g). Finally, 250 μL of the supernatant was added to counting vials containing 5 mL supplementation resulted in increased reception by HDL of scintillation solution (Packard BioScience, Groningen, The cholesteryl esters (P b .05) but did not alter the reception of Netherlands) . Radioactivity was measured with a liquid phospholipids, free cholesterol, or triacylglycerols ).
scintillation analyzer (1600 TR model, Packard, Palo Alto, During the study, the participants did not complain of CA). The blank samples consisted of 200 μL Tris solution symptoms associated with selenosis, such as gastrointest- with added labeled nanoemulsion and the precipitation inal disturbances, rashes, fatigue, irritability, and nervous reagent after incubation, as described above. The results of the radioactive transfer from the lipid nanoemulsion to theHDL fraction were expressed as a percentage of the totalincubated radioactivity, determined in a plasma sample containing no precipitation reagents.
Body weight and serum measurements of the 15 study subjects immediatelybefore and on the last day of the 15-day period of Brazil nut consumption Paraoxonase 1 activity was measured by adding serum to 1 mL Tris-HCl buffer (100 mmol/L, pH 8.0) containing 2 mmol/L CaCl2 and 5.5 mmol/L paraoxon (Sigma Chemical Company, London, UK). The generation of p-nitrophenol was measured at 405 nm, at 37°C in a Bio-Rad Benchmark Microplate Reader (Nippon Bio-Rad, Tokyo) .
The selenium concentration in the plasma was determined Results are expressed as means ± SD (n = 15; 10 women and 5 men).
through atomic absorption spectrometry, using a HITACHI model Z-5000 spectrometer (Hitachi High-Technologies ⁎⁎ P b .0001 compared to baseline (paired t test).
C.C. Strunz et al. / Nutrition Research 28 (2008) 151–155 as beneficial. The transfer of cholesterol into the HDL pool High-density lipoprotein particle size, PON1 activity, and ability of HDL can be considered an antiatherogenic mechanism because it to receive lipids from the artificial nanoemulsion in normolipidemic would facilitate cholesterol elimination into bile while subjects immediately before and on the last day of the 15-day period of avoiding arterial deposition that may occur with cholesterol transported in apo B–containing lipoproteins The lack of effect of nut consumption on the size of the HDL fraction suggests that Brazil nuts did not change the HDL subclass profile. In respect to PON1, this enzyme is related to the antioxidant functions of HDL, and the fact that no alterations of the enzyme activity were found under nut consumption suggests that, at least in part, HDL antiox- As a limitation of the study, we should point out the short Results are expressed as means ± SD (n = 15; 10 women and 5 men).
period of nut consumption was adopted because of the potential toxicity of high-selenium consumption for longperiods and the small study sample. However, as the High MUFA and PUFA content in foodstuffs is participants were employees of the institution, a strict day-to- potentially beneficial in terms of fat composition compared day control of the dietary protocol was possible.
to the SFA Daily ingestion of equivalent amounts of In summary, the consumption of 45 g/d of Brazil nuts by other nuts, such as almonds walnuts pecans , normolipidemic subjects during a short period, at the daily pistachios and peanuts , has led to the improvement of amount range tested for other nuts, did not affect the the plasma lipid profile, with diminished atherogenic LDL established atherosclerosis lipid and apolipoprotein mar- cholesterol. The increase in HDL cholesterol, which is a kers. The only alteration in HDL functional parameters protective factor, occurred less frequently under the inges- measured herein was the increase in cholesteryl ester tion of those nuts . In this study, the lack of reception by the lipoprotein, which may be a protective effectiveness of Brazil nut feeding in reducing LDL cholesterol might be attributed to the relatively greater SFAcontent of this nut compared with other nuts .
As expected, Brazil nut consumption pronouncedly This study was supported by the Fundação do Amparo increased the plasma selenium concentration. Some studies à Pesquisa do Estado de São Paulo, São Paulo, Brazil.
have found a positive correlation between plasma selenium Maranhão is a Research Awardee of the Conselho and HDL cholesterol The plasma levels of selenium Nacional de Desenvolvimento Científico e Tecnológico, attained in our study, 208 ± 55 μg/L, are above the recommended concentrations for this micronutrient, rangingfrom 53 to 161 μg/L However, in our study, the highlevels attained were not harmful to the participants because of the short, 15-day supplementation period and the fact thatin Brazil nut selenium is mainly found in the less toxic [1] Kelly Jr JH, Sabaté J. Nuts and coronary heart disease: an epidemiological perspective. Br J Nutr 2006;96:S61-7.
selenomethionine form . Selenium intoxication was [2] Jenkins DJ, Kendall CW, Marchie A, Parker TL, Connelly PW, Qian described after consumption excesses that lead to a higher W, et al. Dose response of almonds on coronary heart disease risk than 1000 μg/L concentration in the plasma for prolonged, factors: blood lipids, oxidized low-density lipoproteins, lipoprotein(a), long-term periods in areas of endemic selenosis .
homocysteine, and pulmonary nitric oxide. A Randomized, controlled, In this study, in addition to the determination of HDL crossover trial. Circulation 2002;106(11):1327-32.
[3] Ros E, Núñez I, Pérez-Heras A, Serra M, Gilabert R, Casals E, et al. A cholesterol and apo A-I, some fundamental properties of walnut diet improves endothelial function in hypercholesterolemic HDL were also evaluated. Because of the many antiathero- subjects: a randomized crossover trial. Circulation 2004;109(13): genic functions of the lipoprotein, the importance of functional approaches has been highlighted. The ability to [4] Morgan WA, Clayshulte BJ. Pecans lower low-density lipoprotein receive lipids, mediated by transfer proteins such as cholesterol in people with normal lipid levels. J Am Diet Assoc 2000;100(3):312-8.
cholesteryl ester transfer protein and phospholipid transfer [5] Edwards K, Kwaw I, Matud J, Kurtz I. Effect of pistachio nuts on protein, is a fundamental property of HDL because the serum lipid levels in patients with moderate hypercholesterolemia.
metabolism and function of this lipoprotein in the reverse cholesterol transportation depends heavily on lipid [6] Alper CM, Mattes RD. Peanut consumption improves indices of cardiovascular disease risk in healthy adults. J Am Coll Nutr 2003;22(2):133-41.
The single alteration in the lipid transfers to HDL elicited [7] Chunhieng T, Pétritis K, Elfakir C, Brochier J, Goli T, Montet D. Study by nut consumption, that is, the increased influx of of selenium distribution in the protein fractions of the Brazil nuts, cholesteryl esters into the HDL fraction may be interpreted Bertholletia excelsa. J Agric Food Chem 2004;52(13):4318-22.
C.C. Strunz et al. / Nutrition Research 28 (2008) 151–155 [8] Brenneisen P, Steinbrenner H, Sies H. Selenium, oxidative stress, and [18] Mackness MI, Harty D, Bhatnagar D, Winocour PH, Arrol S, Ishola M, health aspects. Mol Aspects Med 2005;26(4-5):256-67.
et al. Serum paraoxonase activity in familial hypercholesterolaemia [9] Flores-Mateo G, Navas-Acien A, Pastor-Barriuso R, Guallar E.
and insulin-dependent diabetes mellitus. Atherosclerosis 1991;86: Selenium and coronary heart disease: a meta-analysis. Am J Clin [19] Hao D, Xie G, Zhang Y, Tian G. Determination of serum selenium by [10] Kris-Etherton PM, Hecker KD, Binkoski AE. Polyunsaturated fatty hydride generation flame atomic absorption spectrometry. Talanta acids and cardiovascular health. Nutr Rev 2004;62(11):414-26.
[11] Mukuddem-Petersen J, Oosthuizen W, Jerling JC. A systematic review [20] Ros E, Mataix J. Fatty acid composition of nuts—implications for of the effects of nuts on blood lipid profiles in humans. J Nutr 2005;135 cardiovascular health. Br J Nutr 2006;96:S29-S35.
[21] Lee O, Moon J, Chung Y. The relationship between serum selenium [12] USDA national nutrient database for standard reference. levels and lipid profiles in adult women. J Nutr Sci Vitaminol 2003;49 [13] Friedewald WT, Levy RI, Fredrickson DS. Estimation of the [22] Spagnolo A, Morisi G, Marano G, Righetti G, Maietta A, Menotti A.
concentration of low-density lipoprotein cholesterol in plasma, without Serum selenium and precursors of cardiovascular risk factors in use of the preparative ultracentrifuge. Clin Chem 1972;18:499-20802.
adolescents. Eur J Epidemiol 1991;7(6):654-7.
[14] Lima ES, Maranhão RC. Rapid, simple laser-light–scattering method [23] Versieck J, Cornelis R, editors. Trace elements in human plasma or for HDL particle sizing in whole plasma. Clin Chem 2004;50(6): serum. Boca Raton (Fla): CRC Press; 1989.
[24] Dietary reference intakes for vitamin C, vitamin E, selenium, and [15] Ginsburg GS, Small DM, Atkinson D. Microemulsions of phospho- lipids and cholesterol esters. Protein-free models of low-density lipoprotein. J Biol Chem 1982;257:8216-27.
[25] Sviridov D, Nestel P. Dynamics of cholesterol transport: protection [16] Maranhão RC, Cesar TB, Pedroso-Mariani SR, Hirata MH, Mesquita against atherosclerosis. Atherosclerosis 2002;161(2):245-54.
CH. Metabolic behavior in rats of a nonprotein microemulsion [26] de Grooth GJ, Klerkx AH, Stroes ES, Stalenhoef AF, Kastelein JJ, resembling low-density lipoprotein. Lipids 1993;28(8):691-6.
Kuivenhoven JA. A review of CETP and its relation to atherosclerosis.
[17] Dina CH, Lo Prete AC, Azevedo CHM, Hueb W, Lopes N, Maranhão RC. Transfer of lipids to high density lipoprotein in coronary artery [27] Lewis GF, Rader DJ. New insights into the regulation of HDL disease patients: effects of simvastatin. XVI International Symposium metabolism and reverse cholesterol transport. Circ Res 2005;96(12): on Drugs Affecting Lipid Metabolism, Nova Iorgue, vol. 1. 2007. p. 485.

Source: http://www.vianutricia.com.br/imagens/namidiavianutricia-nutrition_research.pdf