Effects of non-esterified stanols in a liquid emulsion on cholesterol absorption and synthesis in hypercholesterolemic men

Summary Background Background Numerous studies have shown that dietary plant sterols (phytosterols and phytostanols) and their esters can decrease cholesterol absorption. However, few researchers have examined the effects of plant sterols on cholesterol absorption and synthesis using stable isotope tracers, instead of relying on endogenous pathway precursors. Further, we have worked with non-esterified lecithin-solubilized stanols as opposed to the more frequently studied esterified sterols and stanols. The vehicle was an oil-in-water liquid emulsion rather than the more common spread vehicle typically employed. Aim of the study To determine the effects of relatively low doses of lecithin-solubilized non-esterified stanols in liquid emulsions on cholesterol absorption and synthesis in mildly hypercholesterolemic subjects. Methods In a randomized, double blind crossover design, 12 mildly hypercholesterolemic men received either a free phytostanol supplement (3 g/d in 3 servings) or a control treatment for 3 days. Cholesterol endogenous synthesis rate was determined using the rate of incorporation of deuterium from body water into newly formed cholesterol molecules. Cholesterol absorption at the intestinal level was determined using the dual isotope method using ¹³C cholesterol injected intravenously and ¹⁸O cholesterol given orally. Results Cholesterol absorption was 55.7 ± 6.5 % for the control and 33.5 ± 5.3 % for the phytostanol treatment. This massive reduction of the cholesterol absorption did not induce, on average, a difference in cholesterol endogenous synthesis which was measured at 0.074 ± 0.0015 pool/d for plant sterols and 0.0736 ± 0.0015 pool/d for controls (p > 0.05). Conclusions The results demonstrated that lecithin-solubilized stanols administrated during a short period of time (3 days) in an oil-in-water emulsion can dramatically decrease cholesterol absorption, without a consistent, concomitant increase in synthesis, which is highly suggestive of effective LDL cholesterol lowering. The effects of synthesis should be verified in a longer study with more subjects. Received: 17 September 2001, Accepted: 17 December 2001     

Cholesterol-lowering effect of spreads enriched with microcrystalline plant sterols in hypercholesterolemic subjects

Summary Background Plant sterols have been shown to reduce serum lipid concentrations. The effectiveness is highly dependent on the physical state of the plant sterols. By means of a new crystallizing method, plant sterols can be added into dietary fats and oils homogeneously. In this fat ingredient, plant sterols are in a microcrystalline form. Aims of the study We investigated the cholesterol-lowering effect and possible side effects of vegetable oil-based spreads fortified with two different doses of microcrystalline plant sterols. Methods: This double-blind randomized, placebo-controlled study consisted of a 6-wk run-in and a 6-month experimental period. During the run-in period, all 155 hypercholesterolemic subjects received rapeseed oil-based control spread. In the beginning of the experimental period subjects were randomly assigned into one of three experimental groups. The control group continued to use control spread, and the two test groups used spreads with added plant sterols of either 1.5 g/d or 3.0 g/d. The subjects consumed test spreads as a part of their normal diet without any restrictions in lifestyle and diet. Results Plasma total- and LDL-cholesterol concentrations were significantly reduced by 7.5–11.6 % (0.46–0.62 mmol/l) in groups consuming margarine enriched with free plant sterols, compared with the control group. The effects were similar between the two groups consuming either 1.5 g or 3.0 g plant sterols per day. No effect on HDL-cholesterol or triacylglycerol concentrations occurred. The test spreads did not induce any adverse effects in blood clinical chemistry, hematology or decreases in serum concentrations of lipid soluble vitamins. Conclusions Microcrystalline plant sterols are effective in lowering serum total- and LDL-cholesterol concentrations without obvious side effects. The daily dose of 1.5 g plant sterols is enough to reach the maximum effect. Received: 5 December 2000 / Accepted: 14 March 2001     

The hypocholesterolemic effect of lemon peels,lemon pectin,and the waste stream material of lemon peels in hybrid F<Subscript>1</Subscript>B hamsters

Summary Background We found in preliminary studies with hamsters that citrus peels have a cholesterol lowering effect comparable to that of pectin extracted from these peels. Aim of the study We wanted to examine whether the cholesterol lowering effect of the peels could be completely accounted for by the pectin in the peels. Methods We fed cholesterol enriched (0.1 %,w/w) semipurified diets containing 3 % (w/w) of cellulose, lemon peels, lemon pectin, and the waste stream material of the lemon peels to hybrid F₁B hamsters for a period of 8 weeks. The waste stream of the lemon peels is the left over after extraction of the lemon pectin. Results Feeding the semipurified diets resulted in an increase of plasma cholesterol levels in all the dietary groups after 2 and 4 weeks on the diets. Cholesterol concentrations in the cellulose fed hamsters continued to increase after 4 weeks on the diet, whereas cholesterol levels in the other groups had reached a plateau. As a consequence, the plasma cholesterol levels in the hamsters fed the peels (5.59 ± 0.74 mmol/L, mean ± SD, n = 14), pectin (5.19 ± 0.48 mmol/L), or waste stream (5.53 ± 0.94 mmol/L) were lower than those in the hamsters fed cellulose (6.71 ± 1.52 mmol/L) after 8 weeks on the diets. Differences in total plasma cholesterol were reflected in differences in both VLDL and LDL cholesterol concentration, but this effect was more distinct for the VLDL. There was no effect of the type of fiber on HDL cholesterol levels. Liver cholesterol concentrations paralleled the concentrations of plasma cholesterol and the liver cholesterol concentrations in the hamsters fed the peels (3.57 ± 1.01 μmol/g liver, mean ± SD, n = 14), pectin (4.86 ± 1.42), and the waste stream (4.96 ± 1.89) were lower than those in the cellulose group (7.19 ± 2.32). The hamsters fed the peels, pectin, or waste stream tended to have a higher excretion of fecal bile acids and neutral sterols then the cellulose fed hamsters. Conclusion The results of this study suggest that lemon peels and the waste stream of the lemon peels are as effective in lowering plasma and liver cholesterol in hamsters as the pectin extracted from the peels and that also compounds other than pectin are probably responsible for the cholesterol lowering effect of the citrus peels. Received: 20 September 2001, Accepted: 17 December 2001     

Lipid-altering effects of a dietary supplement tablet containing free plant sterols and stanols in men and women with primary hypercholesterolaemia: a randomized,placebo-controlled crossover trial

This randomized, placebo-controlled, crossover trial assessed the lipid-altering efficacy of a dietary supplement (tablet form) providing 1.8 g/day free (non-esterified) plant sterols and stanols versus placebo for 6 weeks as part of a therapeutic lifestyle changes (TLC) diet in 32 men and women with primary hypercholesterolaemia. Mean ± SE baseline (end of a 5-week TLC diet lead-in) lipid concentrations (mmol/l) were total cholesterol (TC), 5.88 ± 0.08; non-high-density lipoprotein cholesterol (non-HDL-C), 4.71 ± 0.09; low-density lipoprotein cholesterol (LDL-C), 4.02 ± 0.08; HDL-C, 1.17 ± 0.06 and triglycerides (TGs), 1.51 ± 0.12. Differences from control in responses (plant sterol/stanol ? control) were significant (p < 0.05) for LDL-C ( ? 4.9%), non-HDL-C ( ? 3.6%) and TC ( ? 2.8%). HDL-C and TG responses were not significantly different between treatment conditions. These results indicate that 1.8 g/day free plant sterols/stanols administered in a tablet produced favourable lipoprotein lipid changes in men and women with hypercholesterolaemia.     

Effects of insulin and amino acids on leucine metabolism in young and middle-aged humans

Summary Background Aging is characterized by loss of muscle mass. In healthy subjects this process is associated with hormone and nutritional changes which take place over many decades. Aim of the study To investigate the effects of insulin and amino acids on amino acid metabolism in middle-aged humans. Methods We evaluated leucine kinetics by means of the intravenous infusion of [1-¹⁴C]leucine, in 8 young (age 24±2 yr, BMI 21±2 kg/m²) and in 6 middle-aged (age 53±4 yr, BMI 26±1 kg/m²) healthy subjects. Studies were performed under fasting conditions (basal), and after 180 min of euglycemic hyperinsulinemic clamp (study I), or 180 min of euglycemic hyperinsulinemia in combination with an intravenous amino acid infusion (study II). Results In the basal state endogenous leucine flux (ELF, an index of proteolysis), normalized for IBW, averaged 1.71±0.12 and 1.66±0.14 μmol/kg·min in young and middle-aged subjects, respectively. Basal leucine oxidation (0.22±0.03 vs 0.28±0.03 μmol/kg·min, p < 0.05) was lower in middle-aged with respect to young subjects. Non-oxidative leucine disposal (NOLD, an index of protein synthesis: 1.44±0.11 vs 1.43±0.11 μmol/kg·min) was similar in young and middle-aged subjects, respectively. In response to insulin (study I) the absolute and percent decline of ELF and LOX were similar in young and middle-aged subjects: ELF declined to 1.05±0.06 μmol/kg·<,min (−M39±5 %) and 1.07±0.14 μmol/kg·min (−36±4 %), in young and middle-aged, respectively (both p < 0.01 vs basal); LOX declined to 0.21±0.02 μmol/kg·min (−35±3 %), and 0.18±0.05 μmol/kg·min (−28±3 %, p < 0.05 vs basal) in young and middle-aged individuals respectively (both p < 0.01 vs basal). In contrast, insulin-mediated whole-body glucose uptake was lower in middle-aged subjects (6.6±1.4 mg/kg·min) with respect to young individuals (8.1±1.7 mg/kg·min, p < 0.05). During study II (insulin plus AA) a significant rise in NOLD was obtained in both young (1.72±0.10 μmol/kg·min, p < 0.01 vs basal) and middle-aged subjects (1.76±0.25 μmol/kg·min, p < 0.01 vs basal). Similarly, net leucine balance rose significantly in both young (+0.62±0.13 vs −0.25±0.02 μmol/kg·min, p < 0.01 vs basal) and middle-aged subjects (+0.37±0.08 vs −0.22±0.03 μmol/kg·min, p < 0.01 vs basal) suggesting that the anabolic response to amino acids is preserved in middle-aged subjects. Conclusions In middle-aged subjects we observed 1) a moderate decline in basal leucine oxidation; 2) a normal antiproteolytic response to insulin and a reduction in glucose uptake; and 3) a normal anabolic response to AA plus insulin. In conclusion, the data provide evidence for a normal regulation of protein anabolism and an early dissociation between the metabolic effects of insulin on glucose uptake and proteolysis in middle-aged subjects. Received: 6 September 2000, Accepted: 10 July 2001     

Effect of complex polyphenols and tannins from red wine (WCPT) on chemically induced oxidative DNA damage in the rat

Summary Background: Flavonoids are polyphenolic antioxidants occuring in vegetables and fruits as well as beverages such as tea and wine which have been thought to influence oxidative damage. Aim of the study: We wanted to verify whether a complex mixture of wine tannins (wine complex polyphenols and tannins, WCPT) prevent chemically-induced oxidative DNA damage in vivo. Methods: Oxidative DNA damage was evaluated by measuring the ratio of 8-hydroxy-2'-deoxyguanosine (80HdG)/ 2-deoxyguanosine (2dG) × 10⁻⁶ in hydrolyzed DNA using HPLC coupled with electrochemical and UV detectors. Results: We treated rats with WCPT (57 mg/kg p.o.) for 14 d, a dose 10-fold higher than what a moderate wine drinker would be exposed to. WCPT administration significantly reduced the ratio of 80HdG/2dG × 10⁻⁶ in liver DNA obtained from rats treated with 2-nitropropane (2NP) relative to controls administered 2NP only (33.3 ± 2.5 vs. 44.9 ± 3.2 × 10⁻⁶ 2dG; μ± SE; p<0.05). On the contrary, pretreatment with WCPT for 10 d did not protect the colon mucosa from oxidative DNA damage induced by 1,2-dimethylhydrazine (DMH). 2NP and DMH are hepatic and colon carcinogens, respectively, capable of inducing oxidative DNA damage. Conclusions: WCPT have protective action against some types of chemically-induced oxidative DNA damage in vivo. Received: 25 January 1999, Accepted: 27 May 1999     

Triglyceride Recrystallized Phytosterols in Fat-Free Milk Improve Lipoprotein Profiles More Than Unmodified Free Phytosterols in Hypercholesterolemic Men and Women

Objective: Foods incorporating plant sterols (PS) consistently decrease serum low-density lipoprotein cholesterol (LDL-C), although results vary depending on the PS form and food matrix. The objective was to study the effect of a novel triglyceride-recrystallized phystosterol (TRP) incorporated into fat-free milk on markers of cardiovascular risk compared to unmodified free sterols alone in the same fat-free milk.

Methods: Hypercholesterolemic men and women (n = 13 males/7 females; 56 ±10 years; body mass index 27.3 ±5.9 kg/m²) participated in 3 sequential 4-week phases of 480 mL milk consumption. During phase 1 (control) all subjects consumed 2% milk containing no PS, followed by phase 2 with fat-free milk containing free PS (2 g/d fPS) and phase 3 with fat-free milk with TRP (2 g/d). After each phase, determinations of lipoprotein cholesterol distribution, particle concentration via nuclear magnetic resonance (NMR), apolipoproteins, inflammatory markers, and fat-soluble dietary antioxidants were made.

Results: Body mass, body composition, dietary energy and macronutrients, and physical activity were unaffected throughout the study. Compared to the control 2% milk, LDL-C was significantly (p < 0.05) decreased by fPS (?9.1%) and was further decreased by TRP (?15.4%); reductions with TRP were significantly greater. Total LDL particle concentration was decreased to a greater extent after TRP (?8.8%) than fPS (?4.8%; p < 0.05). Only TRP significantly decreased serum levels of apolipoprotein B (apoB; ?6%), interleukin-8 (IL-8; ?11%) and monocyte chemotactic protein-1 (MCP-1; ?19%). Plasma α- and γ-tocopherols and carotenoids, normalized to cholesterol, remained unchanged throughout the study with the exception that β-carotene was lowered by 18%.

Conclusion: In summary, TRP in fat-free milk may provide cardiovascular benefits beyond that of fPS by inducing more substantial decreases in LDL cholesterol and particle concentration, associated with declines in markers of vascular inflammation.     

Magnesium balance studies in premature and term infants

Summary Background: The knowledge of magnesium requirements of premature infants is still very limited, although it is essential for the optimal composition of suitable formulas. Aim of the study: The study concept was 1) to assess physiological magnesium balance data of healthy term infants and longitudinal results from formula-fed premature infants and 2) to deduce conclusions on the magnesium content of the formulas. Methods: Premature infants (n = 14, birth weight ≤ 1500 g, gestational age ≤ 32 weeks) were studied in conventional balance trials with 1) a semi-elemental diet (A), 2) preterm infant formula (B), and 3) infant formula (C). In addition, healthy term formula-fed (n = 11, D) and breast-fed (n = 14, E) infants were investigated. Analysis was performed by flame atomic absorption spectroscopy. Results: The median magnesium intake ranged between 4.84 mg/kg × d⁻¹ (breast-fed infants) and 16.33 mg/kg × d⁻¹ (premature infants). The term breast-fed infants retained nearly as much magnesium as term formula-fed infants (3.37 vs. 3.97 mg/kg × d⁻¹), due to a low percental fecal and urinary excretion. A higher magnesium retention was observed in the premature group: A: 7.97 mg/kg × d⁻¹, B. 5.3 mg/kg × d⁻¹, 3.) 5.54 mg/kg × d⁻¹. Conclusion: In view of the high percental magnesium retention in formula-fed premature infants, excessive supply should be avoided. The long-term effects of lower intakes have to be monitored. Received: 23 July 1999, Accepted: 2 February 2000     

Plant sterols in vegetables and fruits commonly consumed in Sweden

Summary Plant sterols are known to have serum cholesterol lowering effects. A high dietary intake might therefore have a positive impact on health. All food items of vegetable origin contain some amount of plant sterols. The aim of this study was to analyse the plant sterol content of vegetables and fruits commonly consumed in Sweden, and to compare fresh and cooked samples of the same items. Altogether 20 different vegetables and 14 fruits were analysed. All vegetables and fruits were purchased in two shops in the city of Gothenburg, Sweden. Lyophilization was performed within one month of the items being purchased. The samples were frozen at −20 (C and analysed within six months, with a GLC method after acid hydrolysis, alkaline hydrolysis and silylation with tri-methylsilylether. The acid hydrolysis was done in order to detect the fraction of glycosylated plant sterols, which are split during boiling with HCl. The median plant sterol content of vegetables was 14 (3.8–50) mg/100 g edible portion. The highest concentrations were found in broccoli, Brussels sprouts, cauliflower and olives. The median plant sterol content of fruits was 16 (3–44) mg/100 g edible portion. The highest concentrations were found in oranges and passion fruits. The plant sterol concentrations were thus low in vegetables and fruits commonly consumed in Sweden. A serum cholesterol lowering effect attributed to the plant sterols in vegetables and fruits would therefore be of limited significance. Received: 25 September 1998, Accepted: 10 February 1999     

Evidence for an acute rise of intestinal calcium absorption in response to aerobic exercise

Summary. Background: The acute effects of physical activity on intestinal calcium (Ca) uptake and on bone metabolism are not known. Aim of the study: To investigate the consequences of an acute aerobic exercise bout on fractional Ca absorption and on biomarkers of bone turnover. Methods: With the use of a cross over design, eighteen male athletes, aged 25.2 (SE 0.6) years, either had to perform a 60 min run (70 % of maximal speed) or had to rest for 60 min. Intestinal Ca absorption (Fc₂₄₀) was assessed by the use of a stable strontium test. Moreover, calciotropic hormones and serum C-Telopeptide (CTx), a biomarker of bone collagen degradation, and serum C-terminal propeptide of type I collagen (PICP), a marker of bone collagen formation, were measured prior (t₋₆₀) and 3 hours after (t₂₄₀) exercise or rest. Results: Fc₂₄₀ values were significantly enhanced in response to exercise compared to rest (16.2 ± 0.7 % vs. 14.6 ± 0.8 %; P < 0.05). PICP values were significantly lower in response to exercise compared to rest: −9.8 % (P < 0.05). Exercise did not influence serum levels of intact parathyroid hormone and calcitriol. Serum CTx levels decreased markedly between t₋₆₀ and t₂₄₀ during both intervention periods (both P values < 0.001), the results being in line with the circadian rhythm of serum CTx. Conclusions: A moderate exercise bout can induce an acute rise in fractional Ca absorption. Moreover, even in endurance-trained young men a moderate exercise bout acutely decreases bone collagen formation, while the physiologic fluctuations of the bone resorption marker CTx remain unaffected. Received: 3 October 2001, Accepted: 27 June 2002 Correspondence to: Armin Zittermann, PhD     

Plasma levels of advanced glycation end products in healthy, long-term vegetarians and subjects on a western mixed diet

Summary Background Evidence indicates that food-derived Maillard's reaction products are absorbed and yet can be detected in the circulation. Aim of the study We postulated that consumption of the heat-treated food by omnivores could be reflected by higher plasma levels of advanced glycation end products (AGEs) in comparison with vegetarians, who in cooking (by keeping away from meat) use lower temperatures and less time for heating. Methods Plasma fluorescent AGEs (350/450 nm) and Ne-(carboxymethyl)lysine (CML, competitive ELISA) levels were investigated in 3 groups of healthy vegetarians (9 vegans-V, 19 lacto-ovo-vegetarians – VLO and 14 semi-vegetarians – VS) and compared with those of age-matched omnivores (O, n=19). Mean duration of vegetarian diet was V: 7.2±1.0, VLO: 8.2±0.8 and VS: 7.9±1.1 years. Results Both fluorescent AGE (O: 9.9±0.5; V: 10.8±0.7, LO: 13.1±0.8* and SV: 11.6±1.2 ×10³ AU), and CML levels (O: 427.1±15.0, V: 514.8±24.6*, LO: 525.7±29.5**, SV: 492.6±18.0* ng/ml) were significantly lower in omnivores than in vegetarians. Plasma glucose, parameters of renal function (plasma concentration of creatinine and cystatin C, calculated glomerular filtration rate – GFR) as well as C-reactive protein levels were within the normal range and did not differ significantly between the groups. Thus, neither decline of kidney function nor inflammatory processes contributed to the rise in plasma AGEs. Conclusion Enhanced plasma AGE levels in vegetarians in comparison to omnivores are herein presented for the first time. Mechanisms of AGE elevation and potential pathophysiological relevance of this finding are to be elucidated in prospective studies. Received: 18 June 2001, Accepted: 7 November 2001     

A phytosterol-enriched spread improves the lipid profile of subjects with type 2 diabetes mellitus

Summary. Background: Phytosterol-enriched margarines are known to significantly lower total and LDL cholesterol, but little is known about the effect of such margarines in subjects with type 2 diabetes. Aim of the study: Investigation of the effect of a phytosterol-enriched spread in subjects with type 2 diabetes mellitus on serum lipids, Hb_A1c, and blood glucose under free-living conditions. Methods: Randomized, placebo-controlled, double-blind clinical trial in two parallel groups over 12 weeks; 85 type 2 diabetic patients with serum LDL cholesterol levels ≥ 3.60 mmol/l and without hypolipidemic medication were included in the study. Participants consumed 2 × 10 g of spread with or without 8 % phytosterol-esters daily. Fasting blood samples were analyzed at 0, 4, 8, and 12 weeks. Results: After 4 weeks, total and LDL cholesterol were significantly reduced in the phytosterol group by 5.2 % and 6.8 %, respectively, compared to baseline (p < 0.05). After 8 and 12 weeks, these reductions became smaller and were not significant any more compared to baseline or between the groups, but a repeated measurement analysis demonstrated a significant difference for both variables between the two groups (each p < 0.05). HDL cholesterol was significantly increased in the phytosterol group compared to the placebo group after 8 and 12 weeks, but there was no overall difference in the repeated measurement analysis between the two groups. In the phytosterol group, there was a small reduction in Hb_A1c compared to the control group which was only significant after 4 weeks. Conclusions: This clinical study shows that a phytosterol-enriched spread is effective in lowering total and LDL cholesterol in subjects with type 2 diabetes but also illustrates the difficult maintenance under free-living conditions over time. Although this effect is modest, it may contribute to decreasing the elevated risk of cardiovascular disease in type 2 diabetes. Received: 20 September 2002, Accepted: 26 November 2002 Correspondence to: H. Hauner     

Hyperhomocysteinemia,and low intakes of folic acid and vitamin B12 in urban North India

Summary Background and Aim An adverse coronary risk profile has been reported amongst rural-to-urban migrant population living in urban slums undergoing stressful socio-economic transition. These individuals are likely to have low intakes of folic acid and vitamin B12, which may have an adverse impact on serum levels of homocysteine (Hcy). To test this hypothesis, we studied serum levels of Hcy in subjects living in an urban slum of North India and healthy subjects from urban non-slum area. Methods Group I consisted of 46 subjects (22 males and 24 females) living in an urban slum, while group II consisted of healthy subjects (n = 26, 13 males and 13 females) living in the adjacent non-slum area. Anthropometric measurements, biochemical profile (fasting blood glucose, total cholesterol, serum triglycerides, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol) and fasting serum levels of Hcy were measured. Dietary intakes of folic acid, vitamin B12, vitamin B1, and iron were calculated by the 24-hour dietary recall method. Serum levels of Hcy were correlated with dietary intakes of nutrients, anthropometry, and metabolic variables. Results Sex-adjusted serum levels of Hcy in mmol/L (Mean ± SD) were high, though statistically comparable, in both the groups (group I: 20.8 ± 5.9 and group II: 23.2 ± 5.9). Overall, higher than normal serum levels of Hcy (> 15 μmol/L) were recorded in 84 % of the subjects. A substantial proportion of subjects in both groups had daily nutrient intakes below that recommended for the Asian Indian population (folic acid: 93.4 % in group I and 96.7 % in group II, vitamin B12: 76.1 % in group I and 88.4 % in group II). However, between the two groups, average daily dietary intakes of both the nutrients were statistically comparable. As compared to non-vegetarians, vegetarians showed lower intakes of folic acid (p < 0.01) and vitamin B12 (p < 0.01) in both groups. On multivariate linear regression analysis with serum Hcy as the response variable and vegetarian/non-vegetarian status and sex (male/female) as predictor variables, higher serum levels of Hcy were observed in vegetarians vs non-vegetarians (β = 4.6, p < 0.05) and males vs females (β = 5.3, p < 0.01). Conclusions Low intakes of folic acid and vitamin B12, and hyperhomocysteinemia, in both the healthy population living in urban slums and adjacent urban non-slum areas, are important observations for the prevention of nutritional and cardiovascular diseases in the Indian subcontinent. Received: 30 October 2001, Accepted: 14 January 2002     

Effects of a soy milk supplement on plasma cholesterol levels and oxidative DNA damage in men – a pilot study

Summary Background: Phytoestrogens are a major component of Asian diets and may be protective against certain hormone-dependent cancers (breast and prostate) and coronary heart disease. They may also have antioxidant function in scavenging potentially harmful free radicals and thus decreasing oxidative attack on DNA. Aims of the study: A pilot study to determine the effects of a phytoestrogen supplement, in the form of soy milk, on plasma LDL and HDL cholesterol levels and DNA damage in men. Methods: Ten healthy men participated in the study and were assigned to one of three groups consuming 1 litre of either soy milk, rice dream (vegetable protein control) or semi-skimmed cow's milk (animal protein control) each day for 4 weeks. Results: The soy supplement caused significant increases in plasma genistein and daidzein concentrations despite considerable interindividual variation (P<0.001). Supplementation with soy resulted in a decrease in oxidative damage to DNA bases detected using the comet assay compared with controls (P<0.05). However, there was no significant effect of the soy supplement on plasma cholesterol or triglyceride levels in comparison with control groups. Conclusions: A 4 week soy milk supplementation in healthy volunteers does not alter serum cholesterol levels but can have a protective effect against oxidative DNA damage in lymphocytes. Received: 29 December 1998, Accepted: 24 March 1999     

Beta(2)-adrenergic receptor mutation and abdominal obesity risk: effect modification by gender and HDL-cholesterol

Summary Objective and design A case-control study was conducted to examine the association between the 27Glu polymorphism of the β₂-adrenergic receptor gene (ADRB2) and the risk of abdominal obesity (defined by a waist/hip ratio: WHR higher than 0.85). Methods The case series encompassed 112 obese subjects with body mass index (BMI) > 30 kg/m² and WHR > 0.85 and no other major disease except for type 2 diabetes, while the controls were 127 healthy subjects, BMI < 25 kg/m² and WHR < 0.85. Results The association between the risk of abdominal obesity and the 27Glu polymorphism was estimated using multivariate logistic regression. A higher crude odds ratio (OR) of 4.08 (95 % confidence interval: 0.98–16.3) for the 27Glu allele was found among men, while no increased risk was apparent among female participants. Moreover, when the model was adjusted for age, male subjects carriers of the 27Glu allele had a significant ten-fold higher risk of abdominal obesity (OR = 10.31; 95 % CI: 1.4–76.8) and the product-term for the interaction (effect modification) between gender and the ADRB2 mutation was near to the limits of statistical significance (Likelihood ratio test p = 0.056). Interestingly, we also found an effect modification with higher OR among individuals with low HDL-cholesterol (< 1.5 mmol/l) after adjustment for age and gender (OR = 2.87 95 % CI 1.09–7.50) and the product-term for interaction between the 27Glu allele and HDL-cholesterol was statistically significant (Likelihood ratio test p = 0.003). Conclusions. Our results showed that the 27Glu allele of the ADRB2 gene appears to be a risk factor for abdominal obesity among male subjects, specially among those with lower HDL-cholesterol levels. Received: 26 November 2001, Accepted: 2 March 2002     

Study of wheat breakfast rolls fortified with folic acid. The effect on folate status in women during a 3-month intervention

Summary. Background: Folate has come into focus due to its protective role against child birth defects such as neural tube defects (NTD). Swedish authorities recommend all fertile women to increase their folate intake to 400 μg/day by eating folate-rich foods. Because not all women follow these recommendations, there is a discussion today about whether Sweden should introduce folic acid fortification in wheat flour and sifted rye flour. This decision needs knowledge about the bioavailability of folic acid from fortified foods. Aim of the study: To investigate effects of two folic acid fortification levels on folate status in healthy female volunteers and to study the folic acid stability during the baking procedure and storage of the fortified breakfast rolls. Method: Twenty-nine healthy women were recruited. Folic acid-fortified wheat breakfast rolls were baked with the purpose to contain 200 μg folic acid/roll (roll L) and 400 μg folic acid/roll (roll H). Fourteen women were given one roll/day of roll L (group L) and 15 one roll/day of roll H (group H) during 12 weeks of intervention. Fasting venous blood samples were collected on days 0, 30, 60 and 90. Serum homocysteine concentrations were determined using an immunoassay. Serum and erythrocyte folate concentrations were analysed using a protein-binding assay with fluorescent quantification. The folic acid concentration in the breakfast rolls was analysed by HPLC on days 0, 30, 60 and 90. Total folate concentration was measured with microbiological assay on day 45. Results: Group L Group L had initially an average erythrocyte folate concentration of 577 ± 93 nmol/L. After 90 days of intervention, an increase of 20 % (p < 0.05) was observed. At day 0, mean serum folate concentrations were 16.9 ± 4.3 nmol/L. The mean serum folate concentrations increased by 30 % (p < 0.001) after 90 days. At day 0, mean serum homocysteine concentrations were 9.1 ± 2.0 μmol/L, which decreased by 20 % (p < 0.01) after 30 days. Group H Group H had an initial erythrocyte folate concentration of 784 ± 238 nmol/L. After 90 days, an increase of 26 % (p < 0.05) was observed. Serum folate increased at least 22 % after 30 days, from a level of 18.7 ± 4.8 nmol/L at day 0. Thereafter, all women of group H had serum concentrations at or above the upper limit of quantification (23 nmol/L). At day 0, mean serum homocysteine concentrations were 8.4 ± 1.7 μmol/L, which decreased by 16 % (p < 0.05) after 30 days. The baking procedure resulted in 20–25 % loss of fortified folic acid in the rolls used in the present study. The size of the rolls affected the retention of folic acid during baking. No significant loss was seen in folic acid concentration in the rolls during the intervention period. Conclusion: The present study showed that in healthy women, subjected to a 12-week intervention with breakfast rolls fortified with either 166 μg or 355 μg folic acid, serum homocysteine concentration decreased (p < 0.05) and erythrocyte folate increased (p < 0.05). The lower level of fortification seems to be sufficient to improve the folate status. Together with the average daily intake of natural folates, these women reach the recommended intake of 400 μg/day. Folic acid is stable in fortified bread for 90 days storage at −20 °C. Received: 22 April 2002, Accepted: 24 October 2002 Correspondence to: Madelene Johansson     

Deglycosylation by small intestinal epithelial cell beta-glucosidases is a critical step in the absorption and metabolism of dietary flavonoid glycosides in humans

Summary. Background: Pharmacokinetic studies have shown that the small intestine is the major site of absorption for many flavonoid glucosides. Flavonoids are generally present as glycosylated forms in plants and foods, but there is increasing evidence that the forms reaching the systemic circulation are glucuronidated, sulphated and methylated derivatives. Hence, first-pass metabolism (small intestine-liver) appears to involve a critical deglycosylation step for which the mechanisms are not known. Aims: To explore the hypothesis that deglycosylation is a prerequisite to absorption and metabolism of dietary flavonoid glycosides, to identify the enzymes responsible, and relate their specificities with absorption kinetics. Methods: Flavonoid glycoside hydrolysing enzymes were isolated from samples of human small intestine and liver using chromatographic techniques. The proteins were characterised with respect to the cellular fraction with which they were associated, molecular weight, specificity for various substrates, and cross-reactions with antibodies. Cellular models were used to mimic the small intestine. Results: Protein extracts from human jejunal mucosa were highly efficient in hydrolysing flavonoid glycosides, consistent with an enterocyte-mediated deglycosylation process. Considerable inter-individual variation was observed [e. g. range, mean and standard deviation for rate of hydrolysis of quercetin-3-glucoside (n = 10) were 6.7–456, 96, and 134 nmol min⁻¹ (mg protein)⁻¹, respectively]. Two β-glucosidases with activity towards flavonoid glycosides were isolated from human small intestine mucosa: lactase-phlorizin hydrolase (LPH; localised to the apical membrane of small intestinal epithelial cells) and cytosolic β-glucosidase (CBG), indicating a role of human LPH and CBG from small intestine in flavonoid absorption and metabolism. Hydrolysis of flavonoid glycosides was only detected in cultured cells exhibiting β-glucosidase activity. Conclusions: The absorption of dietary flavonoid glycosides in humans involves a critical deglycosylation step that is mediated by epithelial β-glucosidases (LPH and CBG). The significant variation in β-glucosidase activity between individuals may be a factor determining variation in flavonoid bioavailability. Received: 6 July 2002, Accepted: 24 October 2002 Current address: K. Németh, Department of Nutrition and Food Assessment, Faculty of Chemical and Food Technology, Slovak Technical University, Bratislava, Slovak Republic Current address: G. Williamson, Nestec Ltd. Research Centre, Lausanne, Switzerland Correspondence to: Dr. P. A. Kroon     

Plasma concentration response to drinks containing beta-carotene as carrot juice or formulated as a water dispersible powder

Summary. Background: Bioavailability of β-carotene is highly variable and depends on the source, the formulation and other nutritional factors. Objective: It was the aim of the study to compare β-carotene plasma response to b-carotene dosing with two commercially available drinks, containing β-carotene from carrot juice or as water dispersible β-carotene powder. Design In a randomized, parallel group study design, 4 volunteers per group received daily β-carotene doses of 6–7 or 18–22 mg of either drink over 6 weeks. Blood samples for determination of carotenoid and vitamin A plasma concentrations were collected before supplementation and over the dosing period. Results: Apparent steady-state β-carotene concentrations were attained after 40 days of supplementation. Consumption of the beverage containing β-carotene as a water dispersible powder resulted in a higher response of β-carotene plasma concentrations with increments of 3.84 ± 0.60 μmol/L (p < 0.05, dose: 7.2 mg/d) and 5.04 ± 0.72 μmol/L (p < 0.05, dose: 21.6 mg/d), respectively, in comparison to the carrot juice-based drink with increments of 0.42 ± 0.33 μmol/L (dose: 6 mg/d) and 1.71 ± 0.55 μmol/L (dose: 18 mg/d), respectively. β-carotene was cleared from the plasma with an apparent half-life of 6–11 days. Plasma concentrations of α-carotene, β-cryptoxanthin, lutein, zeaxanthin, and lycopene remained almost unchanged, whereas retinol plasma concentrations increased slightly. By contrast, with the exception of elevated 13-cis-retinoic acid in one group (21.6 mg/d, water dispersible powder), the concentrations of all-trans-retinoic acid, and the oxo-derivatives or retinoic acid were not significantly affected by b-carotene supplementation. Conclusions: The results confirm that the relative bioavailability of β-carotene depends largely on the source of b-carotene and demonstrate the superior bioavailability of β-carotene powder in comparison to that in carrot juice. Received: 7 May 2002, Accepted: 22 August 2002 This work was supported by a grant from F. Hoffmann-La Roche Ltd., Vitamins and Nutrition Research, Basle, Switzerland. Correspondence to: Prof. Dr. med. Petra A. Thürmann     

Intestinal absorption of lycopene from different matrices and interactions to other carotenoids, the lipid status, and the antioxidant capacity of human plasma

Summary Background: The bioavailability of carotenoids has been investigated in animal studies as well as in human studies, so far mostly for β-carotene. Only few results exist for lycopene. In recent studies, lycopene was significantly better available from processed tomatoes compared to raw tomatoes, when using daily intakes between 16.5 mg and 75 mg lycopene. Aim of the study In a comparative study the availability of a low oral lycopene dosage of 5 mg/d from different food matrices versus soft gel capsules containing tomato oleoresin was assessed. In addition to the plasma carotenoid content, the effect of lycopene ingestion on other plasma carotenoids, the lipid status parameters, and the antioxidant activitys was estimated. Methods Twenty-two female adults (20 – 27 y) were randomized in three groups and were advized to minimize their carotenoid intake for two weeks. After this initial period, two groups received a portion of tomatoes or tomato juice adjusted to a lycopene dose of 5 mg/d, the third group ingested the same dose comprized in soft gel capsules containing tomato oleoresin. During the test period of 6 weeks, the participants continued reducing the intake of carotenoids from food. Fasting blood samples were withdrawn prior to the study, before supplementation started, and then weekly while supplemented. Seven-day dietary records were prepared before the study started and after one week of supplementation. Carotenoids were analyzed by reversed phase HPLC with diode array detection. Dietary records were evaluated using the computer software EBIS 2.1. The plasma total cholesterol, HDL cholesterol, and triglycerides were determined enzymatically. In addition, the antioxidant activity of plasma was estimated by using the TEAC and the TRAP assays. Results The basal levels of lycopene in plasma were comparable for all groups (0.2 – 0.3 μmol/l) and decreased significantly during the two weeks of depletion to approximately 50 % of the basal values. Other plasma carotenoids such as β-carotene and β-cryptoxanthin decreased significantly, too, whereas lutein and zeaxanthin remained unchanged. After supplementation with tomato oleoresin capsules or tomato juice, the plasma lycopene increased significantly, while it remained unchanged during intake of tomatoes. Normal dietary habits were practized of all volunteers before and during the study except vitamin C whose intake was significantly lower during the study period, because the probands were recommended to reduce the intake of fruits and vegetables. Lycopene supplementation did not affect the lipid status parameters of the three groups. After ingestion of lycopene the antioxidant activity of the plasma was not altered. Mean TEAC values were estimated to 0.33 ± 0.05 mmol/l and TRAP values to 1.0 ± 0.1 mmol/l and showed no significant differences in all groups during the whole study period. Conclusions The bioavailability of lycopene varied significantly depending on the administered matrix. Lycopene from tomato oleoresin capsules and tomato juice (processed tomatoes) was better absorbed from the intestine than lycopene from raw tomatoes. The daily intake of 5 mg lycopene, an intake comparable to the usual daily carotenoid intake, did not affect cholesterol and triglycerides in plasma or its antioxidant capacity. Received: 1 March 1999, Accepted: 30 April 1999     

Multinational study of major breast milk carotenoids of healthy mothers

Summary. Background: Carotenoids in serum vary between countries and within populations with evidence suggesting a qualitative relationship to diet. Breast milk carotenoids furnish a source of vitamin A and potentially provide immunoprotection and other health benefits for infants. There have been numerous studies of milk carotenoid concentrations in undernourished populations; however, carotenoid concentrations have not previously been compared in populations of well-nourished mothers. Aim of Study: To compare concentrations of five major carotenoid groups: α-carotene, β-carotene, β-cryptoxanthin, lutein/zeaxanthin, and lycopene in breast milk of healthy women from Australia, Canada, Chile, China, Japan, Mexico, the Philippines, the United Kingdom, and the United States, and to qualitatively compare patterns of dietary intake with milk carotenoid concentrations. Methods: Breast milk collected from healthy lactating women was analyzed for concentrations of five carotenoids and retinol and quantitated relative to total milk lipid. All determinations were performed in a single research laboratory using standardized methodology. Mothers consumed their usual diets and provided a single 24-h dietary recall. Results: Breast milk carotenoid concentrations varied greatly among countries, with the greatest differences in β-cryptoxanthin (∼ 9-fold) and the least in α-carotene and lycopene (∼ 3-fold). Breast milk retinol concentrations varied ∼ 2-fold across countries. The provitamin A carotenoids α-carotene, β-carotene, and β-cryptoxanthin as a group accounted for > 50 % of the carotenoids measured. Total breast milk carotenoids were highest in Japanese and lowest in Philippine mothers. Breast milk β-carotene concentrations were highest in Chile and lowest in the Philippines. Conclusions: Patterns of breast milk carotenoids were unique to each country and qualitative patterns reflected the dietary carotenoid supply. Received: 24 June 2002, Accepted: 12 December 2002 Supported by a grant from Wyeth Nutrition, Philadelphia, PA, USA. Correspondence to: Louise M. Canfield