Print version ISSN 0212-1611
Nutr. Hosp. vol.26 n.5 Sep./Oct. 2011
Hematologic and immunological indicators are altered by chronic intake of flaxseed in Wistar rats
Indicadores hematológicos e inmunológicos alterados por el consumo crónico de linaza en ratas Wistar
L. Ferreira Medeiros de França Cardozo1, L. Leal Soares1, L.H. Cardozo Brant1, M. Alves Chagas2, V. Alves Pereira2, L. G. Coca Velarde3 and G. Teles Boaventura1
1Laboratory of Experimental Nutrition (LabNE). College of Nutrition. Federal Fluminense University (UFF).
2Laboratory of Cellular and Extracellular Biomorphology. Federal Fluminense University (UFF).
3Statistical Department. Federal Fluminense University (UFF). Brazil.
The authors would like to thank Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) for financial support.
This work sought to evaluate the effects of chronic intake of flaxseed upon hematologic parameters and immunological findings on body development of Wistar rats. Female Wistar rats were used after gestation. They were randomly assigned into two groups during lactation period: Control group (CG), fed with casein based diet, made up of 17% protein and flaxseed group (FG), fed with casein based diet with the addition of 25% flaxseed. At weaning, 12 male pups of each group continued to receive the experimental diets of their mothers (with only 10% of protein) until adult age, when they were killed at 250 days of life aiming at blood collection. At 250 days old FG presented significant reduction in body mass (p < 0.000) and higher levels of hemoglobin (p = 0.019) and albumin (p = 0.030) than CG. It was observed smaller percentage of segmented lymphocytes (p = 0.016) in rats from FG and bigger percentage of segmented leucocytes (p = 0.023) when compared to CG. The chronic consumption of flaxseed altered hematologic and immunological indicators in adult Wistar rats. Supplementation with flaxseed seems to be beneficial to maintenance or reduction of body mass.
Key words: Body weight. Flaxseed. Hematologic indicators. Leukocytes. Rats.
Este trabajo pretendía evaluar el efecto de la ingestión crónica de linaza sobre parámetros hematológicos y hallazgos inmunológicos del desarrollo corporal de ratas Wistar. Se emplearon ratas hembra Wistar tras la gestación. Se las distribuyó al azar en dos grupos durante el período de lactancia: grupo control (GC), alimentado con una dieta basada en caseína, con un 17% de proteína y el grupo linaza (GF), alimentado con una dieta basada en caseína con la adición de un 25% de linaza. En el destete, 12 ratas macho continuaron recibiendo las dietas experimentales consumidas por sus madres (con sólo el 10% de proteína) hasta la edad adulta, en que fueron sacrificados a los 250 días de vida para la recogida de las muestras. A los 250 días de edad, el GF presentaba una reducción significativa de la masa corporal (p < 0,000) y mayores concentraciones de hemoglobina (p = 0,019) y albúmina (p = 0,030) que el GC. Se observó un menor porcentaje de linfocitos segmentados (p = 0,016) en las ratas del GF y un mayor porcentaje de leucocitos segmentados (p = 0,023) en comparación con el GC. El consumo crónico de linaza alteró los indicadores hematológicos e inmunológicos en las ratas Wistar adultas. La suplementación con linaza parece ser beneficiosa en el mantenimiento o la reducción de la masa corporal.
Palabras clave: Peso corporal. Linaza. Indicadores hematológicos. Leucocitos. Ratas.
AA: Arachidonic acid.
CG: Control group.
DHA: Docosahexaenoic acid.
EPA: Eicosapentaenoic acid.
FEC: Food Efficiency Coefficient.
FG: Flaxseed group.
GI: Growth index.
LabNE: Experimental Nutrition Laboratory.
PER: Protein Efficiency Ratio.
PUFAs: Polyunsaturated fatty acids.
Functional food, such as flaxseed, has attracted great attention due to beneficial effects in preventing diseases.1 Previous studies justify its utilization to ameliorate lipid profile,2,3 reduce glycemia,4 diminish tumor growth5 and autoimmune diseases.6
This seed is made up of 41% lipids, 28% fibers, 21% protein, 4% minerals and 6% carbohydrates.7 Benefits to health, mediated by flaxseed, are mainly attributed to its main components: high content of linolenic acid (50-55%) and the secoisolariciresinol diglucoside, which is a lignan present in flaxseed 100 times more than in other food source.8,9
Little is known about chronic use of flaxseed upon hematologic indicators, once it has got good amino acids profile despite its low biologic value when compared to animal derived protein.10 Furthermore, in its composition there are anti-nutritional factors that can cause adverse effects. Linatin can interfere with B6 vitamin absorption, causing deficiency; cyanogenic compound and phytic acid which can chelate minerals such as zinc, iron and calcium.11,12
It is important to determine protein concentrations into blood so as to evaluate if flaxseed intake can provoke hematologic disorders. In humans, it has been studied the short term effects of flaxseed intake upon hematopoietic system, albumin and serum proteins, demonstrating that flaxseed do not provoke deleterious effects when consumed during four weeks.13 In rodents, a different result was reported when 10% flaxseed diet was offered to rats during 56 days. Authors report increase in hematocrit and unchanged values of hemoglobin. 14 Supplementation with a lignan complex from flaxseed in humans for 2 months showed no adverse effects upon hematopoietic system. However, a reduction of leukocytes at the end of the study was found.15 In this way, it is known that diet is important to maintain adequate body lipid composition. There is increasing interest in utilization of polyunsaturated fatty acids (PUFAs) as natural anti-inflammatory agents against inflammatory response and destructive auto-immunity. Greater interest is drawn to n-3 family PUFAs, eicosa - pentaenoic acid (EPA) and docosahexaenoic acid (DHA), whose precursor is alpha-linolenic acid, which is also in flaxseed composition. These PUFAs are widely considered to suppress lymphocyte proliferation and activate macrophages, limiting arachidonic acid (AA).16 A cohort study revealed the presence of abnormal leukocytes counting, which when increased is associated to higher mortality in successive generations. 17 This counting seems to be an independent risk factor, being a cheap and handy indicator to evaluate inflammatory process related to countless diseases.18
No study on possible effects of chronic intake of diet containing 25% of flaxseed was found in the literature. Therefore, the present work aimed to evaluate whether chronic flaxseed intake yields alterations upon Wistar rats hematologic and immunological indicators, besides effects upon body development.
Material and methods
The research project was submitted to Ethics Committee in Animal Research of Fluminense Federal University (UFF), No 00105-09. All procedures were carried out according to Brazilian Scientific Society of Laboratory Animals (SBCAL).
Female Wistar rats were obtained from colonies kept at Experimental Nutrition Laboratory (LabNE) at UFF, nulliparous, 90 days old, matched in a proportion of 3 females to 1 male, receiving commercial chow (23% protein, Nuvilab®, Nuvital Ltda, Paraná, Brazil). After deliver, mothers were randomly assigned to 2 groups during lactation period: Control group (CG), with casein based diet, containing 17% of protein and Flaxseed group (FG), with casein based diet, containing 17% protein with the addition of 25% of flaxseed. At weaning, 12 male pups from each group continued receiving the same experimental diets of their original groups (with only 10% of protein, AIN-93M) until adult age, when they were sacrificed at 250 days old. Body mass and diet intake were evaluated 3 times a week. All animals were kept under controlled temperature (21-23oC) and dark/light cycle (12/12 h), receiving water and diet ad libitum. Rats were anesthetized with intraperitoneal injection of Thiopentax (sodic thiopental 1 g, Cristália Produtos Químicos Farmacêuticos Ltda, Brazil) at a dose of 5% (0.15 mL/100 g of body mass) so as to obtain blood sample through cardiac puncture, being part of the sample placed into tubes containing ethylenediamine tetraacetic acid (EDTA) in order to determine hemoglobin and hematocrit concentrations.
Flaxseed was ground into a blender to obtain a flour, which afterwards was weighted and immediately used to manufacture diet. Experimental isocaloric diets were prepared at LabNE, with 17% of protein and the addition of recommended amounts of vitamins and minerals, following patterns established by Committee on Laboratory Animal Diets, 1979, modified by American Institute of Nutrition-93 (AIN-93G) so as to guarantee that each nutrient exerts its specific function during nursing.19 Diet offered to FG had a concentration of 25% flaxseed, aiming at reaching the recommendation of fiber. This amount of flaxseed had been previously used in other study.20 Ingredients of experimental diets (table I) were weighted and homogenized in industrial mixer Hobart® (São Paulo, SP, Brazil) with boiling water to allow amid gelatinization. The resulting mass was transformed into pellets and dried into ventilated oven (Fabbe-Primar® no171, São Paulo, SP, Brazil) at 60oC for 24h, and after identification, diet was kept under refrigeration until be used. After lactation phase, rats received diets containing 10% of protein, following AIN-93M (table I).
Evaluation of biological value of diets
In order to evaluate biological value of diets, the following indexes were used: Protein Efficiency Ratio (PER), this method is defined as PER = variation of weight gain (g)/protein intake (g). Evaluation of PER was made into a group of animals being fed with the studied protein during 28 days. Considering total variation of total body protein stemmed from differences in the quality of dietary proteins, it is common to measure variation of body mass as a reflection of global actuation of protein intake.21 Afterwards, growth index (GI) was used, which is represented by the application of the same formula of PER, but using data concerning 28 post weaning days.22 The analysis of Food Efficiency Coefficient (FEC) is determined by the relationship between weight variation of animals and dietary intake during 28 days following weaning and demonstrates to which extent one gram of diet promotes increase of body weight.23
In order to calculate the above mentioned indexes, body weight and diet intake were collected each 2 days during the whole experiment. For body mass determination, an electronic digital scale with precision of 0.05 g, Gehara®, was used.
Blood collection and sample preparation
Rats were anesthetized with an intraperitoneal injection of Thiopentax (sodic thiopental 1 g, Cristália Produtos Químicos Farmacêuticos Ltda, Brazil) at a dose of 5% (0.15 mL/100 g of body mass) so as to obtain blood sample through cardiac puncture. Blood sample placed into EDTA containing tube was used to determination of hemoglobin and hematocrit; blood sample without reagents was used to determination of albumin and total protein. Blood was centrifuged (Sigma centrifugal) at 3,500 rpm during 15 minutes to obtain serum, which was stored at -20oC. Analyses of hemoglobin, albumin and total proteins were carried out using BIOCLIN kits (Quibasa industry-Química Básica Ltda/Belo Horizonte-MG). On the other hand, hematocrit was determined with total blood sample (with EDTA), using microhematocrit technique through disposable microcapillary.
Differential counting of leukocytes
For differential counting of leukocytes, blood smear technique was used, being staining of slides made by Instant-Prov kit (Newprov). Different types of cells were evaluated by manual counter ELO´S, with Keys correspondent to each sort of cell.24
Data is presented as average and standard deviation. The normal distribution of the values found was tested through Kolmogorov-Smirnov test. Once the normality of data was verified, it was submitted to comparison between groups using Student T test to independent data. In the results that did not follow normal distribution, non-parametric Wilcoxon test was chosen. The established significance level was p < 0,05. All these analysis were made by S-Plus 8.0.
Evaluation of biological value of diets
It was verified that FG had inferior values of PER (p < 0.000) and CEA (p < 0.000) when compared to CG (table II). Diet intake (p < 0.000) and protein intake (p < 0.000) during this period were significantly inferior in FG, being both evaluations related to consumption if the animal was maintained until 250 days of life (table II). Considering GI, both groups behaved in a similar way (table II). At 250 days old, FG presented body smaller mass than CG (table II).
In table III, FG showed the highest values of hemoglobin (p = 0.019). It was not observed alterations regarding hematocrit percentage and total proteins at 250 days old animals. Higher albumin values (p = 0.030) were found in the group fed onto a diet containing 25% flaxseed.
Table IV shows results of differential counting of leukocytes at the end of the experiment. It was observed smaller percentage of lymphocytes (p = 0.016) in FG and higher percentage of segmented leukocytes (p = 0.023) when compared to CG. It was not verified differences between means of any following parameter: band neutrophils, monocytes, basophils and myelocytes.
Taking PER and FEC results into account, it can be perceived that flaxseed based diet resulted in reduced growth during 28 days after weaning. Lenzi-Almeida et al.25 concluded that flaxseed had inferior impact upon growth than casein diet, indicating that flaxseed cannot be used as exclusive protein source in human diet.
Despite having inferior diet and protein intake during all the experiment, FG presented GI similar to CG at 250 days. This result suggests that even if FG consumed smaller amount of protein and diet, it did not result in any difference concerning growth trajectory after the initial period evaluated by PER. Similar result was found when a flaxseed based diet was offered for 180 days to rats immediately after weaning.26
At the end of experimental period, FG showed a reduction in body mass. Countless authors state that this effect or body mass control can be accounted for intake of integral seed or its isolated components. A previous study showed that after the consumption of isocaloric diets, the group which did not receive flaxseed had greater body mass than the group whose diet contained flaxseed.27 In comparison with control group, female Wistar rats presented smaller weight gain when supplemented with SDG (Secoisolariciresinol diglucoside) and SECO (secoisolariciresinol) for 4 weeks, both of which were obtained from flaxseed.28 Less weight gain was also detected in rats fed with high fat diet made up with flaxseed oil.29 Another study in humans, where three different groups received margarine supplemented with ALA, EPA or DHA for six weeks, demonstrated that effects upon body mass and body mass index seem to be related to duration of supplementation, given that authors did not found differences after treatment.30 In this way, despite being inadequate during growth and development periods, flaxseed and/or its components are capable of reducing body mass gain, being an important tool to control risk factor associated with chronic degenerative diseases.
Serum proteins are cheap and accessible indicators, frequently used in clinical routine to evaluate nutritional status of patients. Furthermore, they are directly related with the intake of some nutrients.31 For instance, hypoalbuminemia is a preditor of bad prognostic in a wide range of situations, not only during the course of a disease but also in healthy population.32 Flaxseed intake seemed to improve these indicators once FG presented higher values of hemoglobin and albumin when compared to CG at 250 days. Alterations in hematocrit percentage and total protein were not observed. In a previous study from our group, pregnant rats were fed with flaxseed based diet during lactation period exclusively and the resulting offspring showed smaller values of hemoglobin at adult age when compared to CG.33 Flaxseed flour when offered at a concentration of 40% to pregnant rats also provoked reduction, but in serum protein.34 In humans, daily supplementation of lignan complex from flaxseed for 12 weeks reduced hemoglobin values, without effect upon albumin, total protein and hematócrito.35
Chronic intake of flaxseed yielded less percentage of lymphocytes together with less segmented leukocytes. Experimental researches revealed that inhibition of limphocytes proliferation only when flaxseed was present at very high concentrations (40%) and this result was attributed to the presence of ALA, once the group that consumed the defatted seed did not show these differences.36 Docosahexaenoic acid (DHA) is a fatty acid from ALA family, which has shown antiinflammatory properties. A recent study determined the effect of DHA upon phagocytic and chemotactic action of peritoneal macrophages, being described a reduction of these activities in rats supplemented with DHA. These results demonstrate the effects of DHA upon immunological system modulation in rats.37 Less deposition of lymphocytes was found in gut mucosa of Wistar rats that received flaxseed oil.38 Supplementation of a complex containing SDG in humans did not alter total leukocytes.35 Likewise, a study in vitro showed that lignans do not modulate leukocyte function in humans.39
Our results suggest that chronic intake of 25% flaxseed into diet alters hematologic and immunological indicators in adult Wistar rats. Supplementation with flaxseed seems to be beneficial to maintenance or reduction of body mass. Further studies are required to elucidate the effects of this seed upon hematologic and immunological findings, considering that literature is scarce.
1. Prasad K. Flax lignan complex slows down the progression of atherosclerosis in hyperlipidemic rabbits. J Cardiovasc Pharmacol Ther 2009; 14 (1): 38-48. [ Links ]
2. Cardozo LFMF, Chagas MA, Soares LL, Troina AA, Boaventura GT. Exposure to flaxseed during lactation does not alter prostate area or epithelium height but changes lipid profile in mts. Nutr Hosp 2010; 25 (2): 250-5. [ Links ]
3. Riediger ND, Othman R, Fitz E, Pierce GN, Suh M, Moghadasian MH. Low n-6:n-3 fatty acid ratio, with fish- or flaxseed oil, in a high fat diet improves plasma lipids and beneficially alters tissue fatty acid composition in mice. Eur J Nutr 2008;47(3):153-60. [ Links ]
4. Zhang W, Wang X, Liu Y, Tian H, Flickinger B, Empie MW, Sun SZ. Dietary flaxseed lignan extract lowers plasma cholesterol and glucose concentrations in hypercholesterolaemic subjects. Br J Nutr 2008; 99 (6): 1301-9. [ Links ]
5. Truan JS, Chen JM, Thompson LU. Flaxseed oil reduces the growth of human breast tumors (MCF-7) at high levels of circulating estrogen. Mol Nutr Food Res 2010; 54 (10): 1414-21. [ Links ]
6. Clark WF, Kortas C, Heidenheim AP, Garland J, Spanner E, Parbtani A. Flaxseed in lupus nephritis: a two-year nonplacebo-controlled crossover study. J Am Coll Nutr 2001; 20 (2 Suppl.): 143-8. [ Links ]
7. Zheng Y, Weisenborn DP, Tostenson K, Kangas N. Energy analysis in the screw pressing of whole and dehulled flaxseed. J Food Eng 2005; 66 (2): 193-202. [ Links ]
8. Carter JF. Potencial of flaxseed and flaxseed oil in baked goods and other products in human nutrition. Cereal Foods World 1993; 38 (10): 753-9. [ Links ]
9. Thompson LU, Robb P, Serraino M, Cheung F. Mammalian lignan production from various foods. Nutr Cancer 1991; 16 (1): 43-52. [ Links ]
10. Oomah BD, Mazza G. Flaxseed proteins- a review. Food Chern 1993; 48 (2): 109-114. [ Links ]
11. Thompson LU. Potential Health benefits and problems associated with antinutrients in foods. Food Res Int 1993; 26 (2): 131-49. [ Links ]
12. Oomah BD, Mazza G, Kenaschuk EO. Cyanogenic compounds in flaxseed. J Agric Food Chem 1992; 40 (8): 1346-48. Doi: 10.1021/jf00020a010. [ Links ]
13. Stuglin C, Prasad K. Effect of flaxseed consumption on blood pressure, serum lipids, hemopoietic system and liver and kidney enzymes in healthy humans. J Cardiovasc Pharmacol Ther 2005; 10(1): 23-7. [ Links ]
14. Babu US, Mitchell GV, Wiesenfeld P, Jenkins MY, Gowda H. Nutritional and hematological impact of dietary flaxseed and defatted flaxseed meal in rats. Int J Food Sci Nutr 2000; 51 (2): 109-17. [ Links ]
15. Prasad K. Effect of chronic administration of lignan complex isolated from flaxseed on the hemopoietic system. Mol Cell Biochem 2005; 270 (1-2): 139-45. [ Links ]
16. Calder PC. N-3 polyunsaturated fatty acids and cytokine production in health and disease. Ann Nutr Metab 1997; 41 (4): 203-34. [ Links ]
17. Ruggiero C, Metter EJ, Cherubini A, Maggio M, Sen R, Najjar SS, Windham GB, Ble A, Senin U, Ferrucci L. White Blood Cell Count and Mortality in the Baltimore Longitudinal Study of Aging. J Am Coll Cardiol 2007; 49 (18): 1841-50. [ Links ]
18. Madjid M, Awan I, Willerson JT, Casscells SW. Leukocyte count and coronary heart disease: Implications for risk assessment. J Am Coll Cardiol 2004; 44 (10): 1945-56. [ Links ]
19. Reeves PG, Nielsen FH, Fahey GC Jr. AIN-93 purified diet of laboratory rodents: final report of the American Institute of Nutrition ad hoc Writing Committee on the Reformulation of the AIN-76A rodents diet. J Nutr 1993; 123(11): 1939-51. [ Links ]
20. Figueiredo MS, Moura EG, Lisboa PC, Troina AA, Trevenzoli IH, Oliveira E, Boaventura GT, Passos MCF. Flaxseed supplementation of rats during lactation changes the adiposity and glucose homeostasis of their offspring. Life Sci 2009; 85 (9-10): 365-71. [ Links ]
21. Angelis RC. Valor nutricional das proteínas: métodos e avaliação. Cad Nutr 1995; 10: 8-29. [ Links ]
22. Soares LL, Lucas AMM, Boaventura GT. Can organic and transgenic soy be used as a substitute for animal protein by rats? Braz J Med Biol Res 2005; 38 (4): 583-86. [ Links ]
23. Madruga MS, Santos HB, Bion FM, Antunes NLM. Avaliação Nutricional de urna dieta suplementada com Multimistura: Estudo em ratos. Ciênc Tecnol Aliment 2004; 24(1): 129-33. [ Links ]
24. Dacie JV, Lewis SM. Practical haematology. 9th ed. Churchill Livingstone, London 2001; 19-46. [ Links ]
25. Almeida KCL, Fernandes FS, Boaventura GT, Guzmán-Silva MA. Efecto de la semilla de linaza (Linum Usitatissimum) em el crecimiento de ratas wistar. Rev Chil Nutr 2008; 35 (4): 443-51. [ Links ]
26. Daleprane JB, Batista A, Pacheco JT, Silva AFE, Costa CA, Resende AC, Boaventura GT. Dietary flaxseed supplementation improves endothelial function in the mesenteric arterial bed. Food Res Inter 2010; 43 (8): 2052-56. [ Links ]
27. Taylor CG, Noto AD, Stringer DM, Froese S, Malcolmson L. Dietary milled flaxseed and flaxseed oil improve N-3 fatty acid status and do not affect glycemic control in individuals with well-controlled type 2 diabetes. J Am Coll Nutr 2010; 29 (1): 72-80. [ Links ]
28. Felmlee MA, Woo G, Simko E, Krol ES, Muir AD, Alcorn J. Effects of the flaxseed lignans secoisolariciresinol diglucoside and its aglycone on serum and hepatic lipids in hyperlipidaemic rats. Br J Nutr 2009; 102 (3): 361-9. [ Links ]
29. Vijaimohan K, Jainu M, Sabitha KE, Subramaniyam S, Anandhan C, Shyamala Devi CS. Beneficial effects of alpha linolenic acid rich flaxseed oil on growth performance and hepatic cholesterol metabolism in high fat diet fed rats. Life Sci 2006; 79 (5): 448-54. [ Links ]
30. Egert S, Kannenberg F, Somoza V, Erbersdobler HF, Wahrburg U. Dietary a-Linolenic Acid, EPA, and DHA have differential effects on LDL fatty acid composition but similar effects on serum lipid profiles in normolipidemic humans. J Nutr 2009; 139(5): 861-8. [ Links ]
31. Fuhrman MP, Charney P, Mueller CM. Hepatic proteins and nutrition assessment. J Am Diet Assoc 2004; 104(8): 1258-64. [ Links ]
32. Franch-Arcas G. The meaning of hypoalbuminaemia in clinical practice. Clin Nutr 2001; 20 (3): 265-9. [ Links ]
33. Cardozo LFMF, Soares LL, Chagas MA, Boaventura GT. Maternal consumption of flaxseed during lactation affects weight and hemoglobin level of offspring in rats. J Pediatr (Rio J) 2010;86(2):126-30. [ Links ]
34. Wiesenfeld PW, Babu US, Collins TRX, Sprando R, O'Don-nell MW, Flynn TJ, Black T, Olejnik N. Flaxseed increased alpha-linolenic and eicosapentaenoic acid and decreased arachidonic acid in serum and tissues of rat dams and offspring. Food Chem Toxicol 2003; 41 (6): 841-55. [ Links ]
35. Fukumitsu S, Aida K, Shimizu H, Toyoda K. Flaxseed lignan lowers blood cholesterol and decreases liver disease risk factors in moderately hypercholesterolemic men. Nutr Res 2010; 30 (7): 441-6. [ Links ]
36. Babu US, Wiesenfeld PW, Collins TFX, Sprando R, Flynn TJ, Black T, Olejnik N, Raybourne RB. Impact of high flaxseed diet on mitogen-induced proliferation, IL-2 production, cell subsets and fatty acid composition of spleen cells from pregnant and Fl generation Sprague-Dawley rats. Food Chem Toxicol 2003; 41 (6): 905-15. [ Links ]
37. Bulbul M, Tan R, Gemici B, Hacioglu G, Agar A, Izgut-Uysal VN. Effect of docosahexaenoic acid on macrophage functions of rats. Immunobiology 2007; 212 (7): 583-7. [ Links ]
38. Rosa DD, Sales RL, Moraes LFS, Lourenço FC, Neves CA, Sabarense CM, Ribeiro SMR, Peluzio MCG. Flaxseed, olive and fish oil influence plasmatic lipids, lymphocyte migration and morphometry of the intestinal of Wistar rats. Acta Cir Bras 2010;25(3): 275-80. [ Links ]
39. Gredel S, Grad C, Rechkemmer G, Watzl B. Phytoestrogens and phytoestrogen metabolites differentially modulate immune parameters in human leukocytes. Food Chem Toxicol 2008; 46 (12): 3691-6. [ Links ]
Ludmila Ferreira Medeiros de França Cardozo.
Universidade Federal Fluminense.
Departamemto de Nutrição e Dietética. Faculdade de Nutrição.
Rúa Mario Santos Braga, 30/5o andar.
CEP 24020-140 Niterói/RJ/Brasil.