Changes in plasma antioxidant status following consumption of diets high or low in fruit and vegetables or following dietary supplementation with an antioxidant mixture.

The aim of the present study was to examine the effect of consumption of a high-fruit and vegetable diet, or a spray-dried extract of selected fruits and vegetables of high antioxidant content, on indices of antioxidant status of individuals consuming a background diet with minimal antioxidant intake. Plasma antioxidant concentrations were determined in twenty-five men following a 2-week depletion period during which they consumed self-selected low-antioxidant diets (less than three servings of fruit and vegetables with no tea, coffee, red wine or fruit juice). Following this period the volunteers consumed either a self-selected diet containing five to seven servings of fruit and vegetables/d, or 30 g of a spray-dried supplement designed to provide the equivalent antioxidant activity of five to seven servings of fruit and vegetables for 2 weeks in a crossover trial. Following consumption of a high-antioxidant diet for 2 weeks, plasma concentrations of ascorbic acid, alpha- and beta-carotene and lutein+zeaxanthin were all significantly increased (P < 0.05) over the depletion period. However, concentrations of lycopene, retinol and tocopherol were not affected. Consumption of the supplement also raised the concentrations of these same antioxidants in plasma. Despite the increases in the concentrations of measured antioxidant nutrients, the 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid-equivalent antioxidant capacity of plasma, as estimated by inhibition of metmyoglobin activity, was not significantly affected by any of the dietary treatments.

Zinc and the gene.

A significant portion of cellular zinc is found in the nucleus where it appears to be critically involved in maintaining genetic stability and in the process of gene expression. With regard to gene expression zinc functions mechanistically at several levels but recent interest has focussed especially on the involvement of zinc in DNA transcription through the activity of transcription factors which contain specific zinc-finger regions which bind to DNA and, in conjunction with other families of transcription factors, control cell proliferation, differentiation and cell death. Because of the central importance of zinc in cell division and growth, considerable attention is paid to zinc as an essential trace element and much has been written concerning dietary sources of zinc and recommended dietary intakes of the metal.

Recommended dietary intake levels for phytochemicals: Feasible or fanciful?

Phytochemicals are biologically active compounds, found in plants in small amounts, which are not established nutrients but which nevertheless seem to contribute significantly to protection against degenerative disease. At present, most interest in phytochemicals is focused on the polyphenolic flavonoids and on the carotenoids, although allium compounds, glucosinolates, indoles and coumarins have also received attention, especially with respect to cancer. Mechanistically, phytochemicals are thought to act in many ways, which include their activity as anti-oxidants, antibacterial/viral agents, phytoestrogens and as inducers or inhibitors of a variety of key enzymes. Recommended dietary intakes (RDI) are the levels of intakes of essential nutrients considered adequate to meet the known nutritional needs of practically all healthy persons. To be regarded as an essential nutrient, a dietary component must be a single identified compound or a close derivative. It should have a demonstrated key biological role and characteristic deficiency syndrome, both of which should respond to nutritional manipulation and are used as a basis for setting an RDI. In these terms, allocating RDI to phytochemicals is problematic, due in part to the large number of chemically different phytochemicals and the lack of a distinctive deficiency syndrome or inherent physiological role in almost all cases. Accordingly, allocation of a single RDI to a general class of phytochemicals would be impracticable, although for an individual phytochemical it may be feasible if acceptable justification for an RDI is extended to include optimum health and evidence is forthcoming of a key inherent role for that compound in maintaining optimum physiological function. However, a distinction will need to be drawn between phytochemicals that participate as integral components of an essential biological system and become recognized as nutrients and those that act as valuable non-nutrient health-promoting agents. Both classes of phytochemicals occur in foods and both could be incorporated into functional foods. Both could be addressed in recommendations such as dietary guidelines, but at present only established nutrient phytochemicals would be eligible for an RDI.

Protection by black tea and green tea against UVB and UVA + B induced skin cancer in hairless mice.

The effects of green and black tea consumption on the early indices of UVB and UVA + B skin damage in hairless mice have been studied in the absence of any chemical tumour initiators or promoters. Black tea consumption was associated with a reduction in the number of sunburn cells in the epidermis of mice 24 h after UVA + B irradiation, although there was no effect of green tea. Other indices of early damage such as necrotic cells or mitotic figures were not affected. Neutrophil infiltration as a measure of skin redness was slightly lowered by tea consumption in the UVB group. Consumption of either green or black tea resulted in significantly fewer skin papillomas and tumours induced by UVA + B light, however black tea provided better protection against UVB-induced tumours than green tea. This study confirms earlier reports that tea consumption can reduce the incidence of skin cancer in hairless mice, and indicates that black tea may afford more protection against simulated solar irradiation than green tea.

Protection by tea against UV-A + B-induced skin cancers in hairless mice.

Consumption of tea, especially green tea, has been shown to reduce the incidence of ultraviolet (UV)-related skin tumors in hairless mice. Because milk is added to much of the tea consumed in Western cultures, we have studied the effects of including milk in the tea consumed by hairless mice receiving simulated solar radiation. Under these conditions, mice consuming tea with 10% whole milk had 30% fewer papillomas, 50% fewer tumors, and 55% smaller lesions than mice consuming water. Mice consuming tea alone had fewer papillomas and tumors than mice consuming tea with milk; however, the difference in area affected was not statistically significant. In separate experiments, there was a significant dose response to black tea as a preventive against UV-related skin lesions, and also consumption of black tea was associated with a small but significant reduction in the incidence of papillomas in mice previously exposed to UV radiation. The results of these studies demonstrate that, in hairless mice, black tea can inhibit the formation of UV-induced skin tumors in a dose-dependent manner and, even with the addition of milk, can still inhibit the growth of UV-related skin tumors.

Nutrition, cancer, and aging.

The parallel increase in cancer risk with advancing age is well recognized, and several pathophysiological mechanisms common to both conditions have been proposed to explain this interrelationship. The importance of nutrition, both in delaying the aging process and in protecting against cancer is also well recognized, and it is therefore of interest to compare the relative impact several of the more widely studied dietary manipulations may have on each of these conditions. For example, caloric restriction, which putatively reduce oxidative stress and effectively increases life span in animals also seems to reduce the incidence of many cancers, possibly due to diminished mitogenesis. Likewise, oxidative damage to DNA appears to be common to both processes but may be more important in the mitochondria with respect to aging and in the nucleus in relation to cancer. Inadequate dietary folate and impaired DNA methylation status are closely associated with increased cancer risk, and recently defective somatic cell methylation and accumulated genetic instability have been proposed as key mechanisms contributing to senescence. Several other well-established anticancer dietary strategies, which include increased fiber intake and the consumption of more fruits and vegetables, have not been studied extensively in relation to aging, although many of the phytochemicals considered important as chemopreventive agents for cancer may well contribute to delaying the aging process. Although not directly related to nutrition, but nevertheless highly relevant, is the question of physical activity, which has been strongly linked to a reduction in risk of some cancers. Although less is known with respect to exercise and biological markers of aging, physical activity does appear to retard the age-related decline in the muscle strength and in the bone density.

Genistein inhibits growth of B16 melanoma cells in vivo and in vitro and promotes differentiation in vitro.

Consumption of soy products has been linked to a reduced mortality and morbidity from a number of cancers. Genistein, one of the principal soy isoflavones, has been shown to inhibit the growth of a number of tumour cell lines in vitro; however, a role of genistein in retarding tumour growth in vivo is less well documented. In this study, in addition to examining the effects of genistein on the growth of murine B16 melanoma cells in vitro, we have examined the effects of feeding a genistein-rich diet on s.c. growth of these tumour cells in mice. In vitro, the melanoma cells showed an increase in sensitivity to genistein with increasing time of exposure, culminating in a 50% growth inhibition (IC50) at 12.5 microM after 7 days. Genistein at 25 microM induced micronucleus formation after 24 hr and at concentrations as low as 2.5 microM induced morphological changes indicative of differentiation. Growth of solid tumours implanted into female C57BL/6J mice was inhibited by 50% when mice were fed genistein for 1 week before and for 1 week after inoculation with B16 melanoma cells. Plasma genistein concentrations at the time of tumour removal were 1.1 microM, which is similar to levels reported in humans consuming diets high in soybeans or soybean products, while control animals had no detectable genistein in plasma. Our results provide additional in vivo evidence suggesting that genistein retards the growth of implanted tumours, adding further to studies suggesting that this isoflavonoid is a biologically active component of soy foods.

Folate, vitamin B12, homocysteine status and chromosome damage rate in lymphocytes of older men.

Deficient levels of folic acid and vitamin B12 are associated with elevated chromosome damage rate and high concentrations of homocysteine in the blood. We have therefore performed a study to determine the prevalence of folate deficiency, vitamin B12 deficiency and hyperhomocysteinemia in 64 healthy men aged between 50 and 70 years, and evaluate the relationship of these micronutrient levels in the blood with the micronucleus frequency in peripheral blood lymphocytes. We also performed a placebo-controlled, double-blind intervention study to determine whether supplementation of the diet with a daily dose of 0.7 mg (as a supplement in cereal) or 2.0 mg (in a tablet) over a period of 4 months resulted in a significant alteration of folate status, homocysteine status and the micronucleus index. Twenty-three per cent of the men were serum folate deficient (<6.8 nmol/l), 16% were red blood cell folate deficient (<317 nmol/l), 4.7% were vitamin B12 deficient (<150 pmol/l) and 37% has plasma homocysteine levels >10 micromol/l. In total, 56% of the men had one or more abnormal blood values for folate, vitamin B12 or homocysteine. The micronucleus index of these men (n = 34) in cytokinesis-blocked binucleated cells (19.2 +/- 1.1) was significantly elevated (P = 0.02) when compared to the micronucleus index of the rest of the men who had normal levels of folate, vitamin B12 and homocysteine (16.3 +/- 1.3, n = 30). Interestingly, the micronucleus index in men with normal folate and vitamin B12, but homocysteine levels >10 micromol/l (19.4 +/- 1.7, n = 15) was also significantly higher (P = 0.05) when compared to those with normal folate, vitamin B12 and homocysteine. This novel result was also supported by the observation that the micronucleus index and plasma homocysteine were significantly (P = 0.0086) and positively correlated (r2 = 0.172) in those subjects who were not deficient in folate or vitamin B12. The micronucleus index was not significantly correlated with folate indices, but there was a significant (P = 0.013) negative correlation with serum vitamin B12 (r2 = 0.099). Daily supplementation of the diet with 0.7 mg free folic acid in cereal for 2 months followed by 2.0 mg free folic acid via a tablet produced a 4-fold increase in plasma folate, a 2.6-fold increase in red blood cell folate and a 11% reduction in plasma homocysteine; however, these changes were not accompanied by a reduction in the micronucleus index. In conclusion, it is apparent that elevated homocysteine status, in the absence of vitamin deficiency and low, but not deficient, vitamin B12 status are important risk factors for increased chromosome damage in lymphocytes.

Cancer biomarkers in the field of tea.

Suitable intermediate end-point biomarkers are needed in order to reduce the time and expense associated with cancer chemoprevention studies. Because of the multi-step nature of cancer intermediate biomarkers reflect a continuum of events associated with different stages of disease development, and range from genetic damage linked to cancer initiation, to tumor growth and expansion during disease progression. With tea, experimental evidence points to potential protection at several stages of carcinogenesis including endogenous carcinogen formation, carcinogen activation and detoxification, DNA damage and destabilisation, cell proliferation, neoplastic growth and metastatic spread. If used effectively in tea research, biomarkers should contribute to a better understanding of the possible extent of cancer protection afforded by tea, as well as the putative mechanisms involved and the principal components of tea with prophylactic potential.

Inhibition of carcinogenesis by tea: the evidence from experimental studies.

In its various forms, tea is one of the most widely consumed beverages in the world. Elucidation of the chemical components of tea has revealed that the beverage is a rich repository of antioxidants. Among these are the polyphenolics, common to green tea, but also found in black teas together with oxidized polymers that in part account, for the darkened color. Consumption of tea on a regular basis has been associated with reduced risk of several forms of cancer in human populations, with the strongest evidence linking green tea use to reduction in cancer risk in parts of Asia. To understand how tea prevents cancer, studies in animal carcinogenesis models have been done with very encouraging results. This review examines the available data from animal studies on the effects of tea in the prevention of cancer.

Protection by zinc against UVA- and UVB-induced cellular and genomic damage in vivo and in vitro.

For many years, zinc salts have been used both topically and orally to treat minor burns and abrasions as well as to enhance wound repair in man and animals. In this study we describe the protective effects of zinc against UV-induced genotoxicity in vitro and against sunburn cell formation in mouse skin in vivo. Cultured skin cells from neonatal mice showed a dramatic increase in the number of micronuclei as a result of UVA and UVB irradiation. Inclusion of zinc at 5 micrograms/mL in the medium significantly reduced the frequency of micronuclei and of micronucleated cells. In hairless mice, topical application of zinc chloride for 5 consecutive days or a single application 2 h prior to UV exposure reduced the number of sunburn cells in the epidermis as did application of zinc 1 h after exposure. Application 2 h after irradiation also tended to have a protective effect, although there was a large variation between animals. It is proposed that an influx of zinc can protect epidermal cells against some of the more delayed effects of UV-induced damage.

Black tea, green tea, and tea polyphenols. Effects on trace element status in weanling rats.

Previous studies have shown that tea consumption can impair trace element metabolism, particularly iron status, and increase the risk of anemia in humans and animals. More recently, however, evidence has been accumulating to show that, in animals, consumption of green tea or its polyphenols is associated with a reduction of the incidence and severity of a variety of experimentally induced cancers. In this study we have monitored the growth, trace element status, including hematological parameters of weanling rats given either (1) water, (2) 1% black tea, (3) 1% green tea, or (4) 0.2% crude green tea extract as their sole drinking fluid while consuming diets containing either adequate or low amounts of iron. With the exception of manganese, none of the trace elements studied (iron, copper, zinc, and manganese) or the hematological indices measured were affected by the type of beverage supplied, even though the polyphenol extract was shown to chelate metals in vitro and all the animals fed the low iron diet were shown to be anemic. There appeared to be an effect of black and green teas on manganese balance in both the first and last weeks of the study. A lower level of brain manganese was associated with green tea consumption, and a higher level of this element in the kidneys of animals fed black tea. The results demonstrate that both black and green teas and a green tea polyphenol extract do not represent a risk to animals consuming the beverages as their sole fluid intake with respect to iron availability, although the interactions with manganese deserve further study.

Induction of micronucleus formation in mouse splenocytes by the soy isoflavone genistein in vitro but not in vivo.

The effects of genistein (one of the major soybean isoflavones), genistein (the glucosylated form of genistein) and etoposide (a topoisomerase 11 inhibitor) have been studied in mouse splenocytes in culture. Genistein (25 microM), genistein (25 microM) and etoposide (0.1 microM) all induced the production of large numbers of micronuclei; however, genistein at 12.5 or 2.5 microM had no clastogenic effect. In a second study, mice were gavaged with 20 mg genistein/kg body weight/day for 5 days (approximately equivalent to a 70 kg human consuming 2.8 kg soybeans/day) and the micronucleus frequency was determined. There was no observable increase in the micronucleus frequency even though the plasma genistein levels in the treated animals were found to be 9.2 +/- 2.0 microM compared with 0.1 +/- 0.0004 microM in the control animals. The results show that even though genistein is capable of inducing micronucleus formation, an event associated with genetic damage, plasma levels are unlikely to be sufficiently elevated to produce such an effect.

Neural tube defects and maternal serum zinc and copper concentrations in mid-pregnancy: a case-control study.

OBJECTIVE: To assess the relationship between mid-pregnancy maternal serum zinc and copper concentrations and neural tube defects. DESIGN: A prospective case-control study during 1978-1988 within a statewide hospital-based neural tube defect screening program measuring maternal serum alpha-fetoprotein levels at mid-pregnancy. SUBJECTS: Cases were 69 women with fetuses with confirmed neural tube defects. Controls were 592 women with fetuses without neural tube defects who were individually matched to cases for hospital, calendar date of screening, age and parity; there was a variable control-to-case ratio. RESULTS: For both unmatched and adjusted matched analyses, mean maternal serum zinc concentration was higher in cases than controls (P = 0.02 and P = 0.03, respectively). There were no case-control differences for serum copper concentrations. Conditional logistic regression analysis showed a (statistically non-significant) 50% increase in risk of neural tube defects in women whose serum zinc concentration was more than two standard deviations above the population mean. CONCLUSION: Within the normal range of maternal serum zinc and copper concentrations there is no variation in risk of neural tube defects. However, women with very high serum zinc levels may have an increased risk of neural tube defects. This could reflect deficient maternal-to-fetal transfer of zinc in some of those individuals. Any such phenomenon would be manifest in observational, but not experimental, studies.

Recommended dietary intakes of iron, zinc, and other inorganic nutrients and their chemical form and bioavailability.

The bioavailability of a nutrient can, in some cases, be significantly influenced by the chemical form in which it appears in the diet and by the presence of other factors in food that may enhance or depress mineral absorption and utilization. With inorganic nutrients in particular, it is important to understand the physicochemical factors that underlie the differences in bioavailability of some trace elements from diets of differing composition, so that meaningful recommended dietary intakes (RDIs) may be set for these nutrients in relation to the dietary profiles of the countries in which they are to be used. Iron and zinc are especially noteworthy in this regard because their absorption can be greatly influenced by chemical form, ionic competition for uptake mechanisms, and the presence of favorable or unfavorable chelating and binding agents in the diet. Recognizing this variability, several national and international authorities recommending RDIs for these nutrients list a range of desirable intakes that relate to the estimated bioavailability of the mineral from various dietary regimens.

The teratogenic effects of salicylic acid on the developing nervous system in rats in vitro.

Aspirin ingestion in humans and animals has been reported to lead to a range of undesirable outcomes, including fetal death, growth retardation, and congenital abnormalities. Rat embryos were cultured for 48 h in 100-300 micrograms/ml of salicylic acid, a metabolite of aspirin, days 9.5-11.5 of gestation. When compared to growth in control embryos, a significant dose-dependent decrease in crown-rump lengths, somite numbers, and yolk sac diameters was observed. There was also a significant increase in overall dysmorphology, including eye, brachial arch, and heart anomalies, and an absence of forelimb buds. The neural tube was especially vulnerable and had frequently failed to close. Cellular and ultrastructural examination revealed extensive cell death in the neuroepithelium, with a lesser effect on the mesenchymal cells. Large condensed blebs projected into the ventricular lumen, and cell membranes as well as the basal lamina were severely disrupted, with all cytoplasmic organelles affected in dying cells. It is likely that the extensive cell necrosis and blebbing in the developing neuroepithelium at the site of neural tube fusion may be involved in failed neurulation, while necrosis at other sites in the cranial neuroepithelium may be linked with previously reported intellectual and behavioural abnormalities.