Jeevan K. Prasain

Hypoglycemic effect of the water extract of Piper sarmentosum in rats.

The hypoglycemic effect of the water extract of the whole plant of Piper sarmentosum Roxb. (Piperaceae, Thai name: Chaplu) was examined in normal and streptozotocin-diabetic rats. In an oral glucose tolerance test, a single oral administration of the water extract at doses of 0.125 and 0.25 g/kg significantly lowered the plasma glucose level in the normal rats. A reference drug, glibenclamide, at a dose of 5 mg/kg (per os, p.o.) also showed a significant hypoglycemic effect in the normal rats. In contrast, a single oral administration of the water extract at these doses and glibenclamide did not significantly lower the plasma glucose level in the diabetic rats. However, the repeated oral administration of the water extract at a dose of 0.125 g/kg for 7 days produced a significant hypoglycemic effect in the diabetic rats. Glibenclamide (5 mg/kg, p.o.) also caused significant hypoglycemia in the diabetic rats. These results demonstrated that the water extract of whole plant of Piper sarmentosum has a hypoglycemic effect in rats.

Chemoprevention by Grape Seed Extract and Genistein in Carcinogen-induced Mammary Cancer in Rats Is Diet Dependent

Many popular dietary supplements are enriched in polyphenols such as the soy isoflavones, tea catechins, and resveratrol (from grape skins), each of which has been shown to have chemopreventive activity in cellular models of cancer. The proanthocyanidins, which are oligomers of the catechins, are enriched in grape seeds and form the basis of the dietary supplement grape seed extract (GSE). Evidence suggests that the proanthocyanidins may be metabolized to the monomeric catechins. This study was carried out to determine whether GSE added to rodent diets protected against carcinogen-induced mammary tumorigenesis in rats and whether this was affected by the composition of the whole diet. Female rats were begun on 5%, 1.25%, or 0% (control) GSE-supplemented diets at age 35 d. At age 50 d they were administered 7,12-dimethylbenz[a]anthracene (DMBA) in sesame oil at 80 mg/kg body weight. They were weighed and monitored weekly for tumor development until 120 d after DMBA administration. Administration of GSE in AIN-76A diet did not show any protective activity of GSE against DMBA-induced breast cancer. However, administration of GSE in a laboratory dry food diet (Teklad 4% rodent diet) resulted in a 50% reduction in tumor multiplicity. In similar experiments, genistein administered in AIN-76A diet also failed to show chemopreventive activity against the carcinogen N-methyl-N-nitrosourea; however, when administered at the same dose in the Teklad 4% rodent diet, genistein exhibited significant chemopreventive activity (44-61%). These results demonstrate that GSE is chemopreventive in an animal model of breast cancer; moreover, the diet dependency of the chemopreventive activity for both GSE and genistein suggests that whether or not a compound is chemopreventive may depend on the diet in which the agent is administered.

Liquid chromatography tandem mass spectrometry identification of proanthocyanidins in rat plasma after oral administration of grape seed extract.

Proanthocyanidin rich plant extracts derived from grape seed extract (GSE), hawthorn and cranberry are on markets for their preventive effects against cardiovascular diseases and uroinfections in woman. However, the importance of these health beneficial effects of these botanicals remains elusive due to incomplete understanding of uptake, metabolism and bioavailability of proanthocyanidins in vivo. In the present study rats were given GSE orally (300 mg/kg, twice a day) and blood and urine were collected over a 24 h period. Monomeric catechins and their methylated metabolites, and proanthocyanidins up to trimers were detected in blood samples treated with GSE using LC-MS/MS operating in the multiple reaction monitoring (MRM) mode. A new tetramethylated metabolite of dimeric proanthocyanidin (m/z 633) in GSE-treated urine was tentatively identified. Using LC-MS/MS, (+)-catechin and (-)-epicatechin were identified in the brain conclusively. These data suggested that GSE catechins cross the blood brain barrier and may be responsible for the neuroprotective effects of GSE.

Flavonoids and Age Related Disease: Risk, benefits and critical windows

Plant derived products are consumed by a large percentage of the population to prevent, delay and ameliorate disease burden; however, relatively little is known about the efficacy, safety and underlying mechanisms of these traditional health products, especially when taken in concert with pharmaceutical agents. The flavonoids are a group of plant metabolites that are common in the diet and appear to provide some health benefits. While flavonoids are primarily derived from soy, many are found in fruits, nuts and more exotic sources, e.g., kudzu. Perhaps the strongest evidence for the benefits of flavonoids in diseases of aging relates to their effect on components of the metabolic syndrome. Flavonoids from soy, grape seed, kudzu and other sources all lower arterial pressure in hypertensive animal models and in a limited number of tests in humans. They also decrease the plasma concentration of lipids and buffer plasma glucose. The underlying mechanisms appear to include antioxidant actions, central nervous system effects, gut transport alterations, fatty acid sequestration and processing, PPAR activation and increases in insulin sensitivity. In animal models of disease, dietary flavonoids also demonstrate a protective effect against cognitive decline, cancer and metabolic disease. However, research also indicates that the flavonoids can be detrimental in some settings and, therefore, are not universally safe. Thus, as the population ages, it is important to determine the impact of these agents on prevention/attenuation of disease, including optimal exposure (intake, timing/duration) and potential contraindications.

Polyphenols, Inflammatory Response, and Cancer Prevention: Chlorination of Isoflavones by Human Neutrophils

An important aspect of the risk of cancer is the involvement of the inflammatory response. Currently, antiinflammatory agents are used in chemopreventive strategies. For example, aspirin is recommended for the prevention of colon cancer as well as breast and other cancers. The inflammatory response involves the production of cytokines and proinflammatory oxidants such as hypochlorous acid (HOCl) and peroxynitrite (ONO2-) produced by neutrophils and macrophages, respectively. These oxidants react with phenolic tyrosine residues on proteins to form chloro- and nitrotyrosine. Diets rich in polyphenols (green tea catechins, soy isoflavones) have also been shown to be chemopreventive. The aromatic nature of polyphenols makes them potential targets of HOCl and ONO2-. These reactions may create novel pharmacophores at the site of inflammation. Previous studies in the neutrophil-like cell line, differentiated HL-60 cells, demonstrated the formation of chlorinated and nitrated isoflavones. In this study we have examined whether similar reactions occur in freshly isolated human neutrophils. After induction of a respiratory burst with a phorbol ester, isoflavones and their metabolites were identified by liquid chromatography-tandem mass spectrometry and then quantitatively measured by LC-mass spectrometry using multiple-reaction ion monitoring. The data obtained indicate that both chlorinated and nitrated genistein are formed by human neutrophils. The extent of chlorination of genistein was markedly increased by the phorbol ester whereas the low level of nitration of genistein was constitutive and unaffected. These data imply a potential role for modified forms of genistein that would be produced in the inflammatory environment in and around a tumor.

Insulin/FOXO Signaling Regulates Ovarian Prostaglandins Critical for Reproduction

Abnormalities in insulin/IGF-1 signaling are associated with infertility, but the molecular mechanisms are not well understood. Here we use liquid chromatography with electrospray ionization tandem mass spectrometry to show that the C. elegans insulin/FOXO pathway regulates the metabolism of locally acting lipid hormones called prostaglandins. C. elegans prostaglandins are synthesized without prostaglandin G/H synthase homologs, the targets of nonsteroidal anti-inflammatory drugs. Our results support the model that insulin signaling promotes the conversion of oocyte polyunsaturated fatty acids (PUFAs) into F-series prostaglandins that guide sperm to the fertilization site. Reduction in insulin signaling activates DAF-16/FOXO, which represses the transcription of germline and intestinal genes required to deliver PUFAs to oocytes in lipoprotein complexes. Nutritional and neuroendocrine cues target this mechanism to control prostaglandin metabolism and reproductive output. Prostaglandins may be conserved sperm guidance factors and widespread downstream effectors of insulin actions that influence both reproductive and nonreproductive processes.

Effects of botanical dietary supplements on cardiovascular, cognitive, and metabolic function in males and females

BACKGROUND The onset of menopause marks a pivotal time in which the incidence of hypertension and of cardiovascular disease (CVD) begins to increase dramatically in women. Before menopause, the incidences of these diseases are significantly lower in women than in age-matched men. After menopause, the rates of these diseases in women eventually approximate those in men. The loss of endogenous estrogen at menopause has been traditionally believed to be the primary factor involved in these changes. OBJECTIVE This review summarizes recent findings regarding the effectiveness of botanicals in the treatment of some menopausal symptoms and other symptoms of aging (eg, rise in arterial pressure, cognitive decline, insulin resistance, and hyperlipidemia). METHODS Articles were selected for inclusion in this review based on the significance of the research and contribution to the current understanding of how each botanical elicits cardioprotective effects. To this end, PubMed and MEDLINE databases were searched, using terms that included the name of the specific botanical along with the relevant aspects of its action(s), such as blood pressure, glycemic control, and lipids. Most of the articles used were published within the past 5 years, although some older articles that were seminal in advancing the current understanding of botanicals were also included. RESULTS Soy has been found to lower plasma lipid concentrations and arterial pressure in postmenopausal women and age-matched men, and to have protective effects in heart disease and atherosclerosis of the carotid and coronary circulation. Soy was also found to lower fasting insulin concentrations and glycosylated hemoglobin concentrations. Grape seed extract, another frequently used botanical, contains polyphenols that have been found to reduce arterial pressure and salt-sensitive hypertension in estrogendepleted animal models. CONCLUSION Several botanical compounds have been found to have beneficial effects in the treatment of the symptoms of menopause and other symptoms of aging, including CVD, cognitive decline, and metabolic diseases.

A Heterogeneous Mixture of F-Series Prostaglandins Promotes Sperm Guidance in the Caenorhabditis elegans Reproductive Tract

The mechanisms that guide motile sperm through the female reproductive tract to oocytes are not well understood. We have shown that Caenorhabditis elegans oocytes synthesize sperm guiding F-series prostaglandins from polyunsaturated fatty acid (PUFA) precursors provided in yolk lipoprotein complexes. Here we use genetics and electrospray ionization tandem mass spectrometry to partially delineate F-series prostaglandin metabolism pathways. We show that omega-6 and omega-3 PUFAs, including arachidonic and eicosapentaenoic acids, are converted into more than 10 structurally related F-series prostaglandins, which function collectively and largely redundantly to promote sperm guidance. Disruption of omega-3 PUFA synthesis triggers compensatory up-regulation of prostaglandins derived from omega-6 PUFAs. C. elegans F-series prostaglandin synthesis involves biochemical mechanisms distinct from those in mammalian cyclooxygenase-dependent pathways, yet PGF2α stereoisomers are still synthesized. A comparison of F-series prostaglandins in C. elegans and mouse tissues reveals shared features. Finally, we show that a conserved cytochrome P450 enzyme, whose human homolog is implicated in Biettis Crystalline Dystrophy, negatively regulates prostaglandin synthesis. These results support the model that multiple cyclooxygenase-independent prostaglandins function together to promote sperm motility important for fertilization. This cyclooxygenase-independent pathway for F-series synthesis may be conserved.

DING, a Genistein Target in Human Breast Cancer: A Protein Without a Gene

Because most noncancer cells are tolerant to high micromolar concentrations of genistein (GEN), inhibitory or stimulatory effects of GEN have been claimed for a wide variety of biochemical targets that lead to a plethora of potential mechanisms. However, because GEN is present in tissues in the nanomol-per-liter range, most of these mechanisms are unlikely to be relevant in vivo. To better identify proteins that are targets of GEN, we used a GEN-agarose-affinity phase. Cytosols from human breast cancer MCF-7 cells were fractionated over a Sephadex diethylaminoethyl column, and nonabsorbed proteins in the flow-through were affinity absorbed onto a 2-carboxygenistein-agarose column. After proteins were washed with 100 mmol NaCl/L to remove weakly bound proteins, affinity elution was conducted with 1 mmol 2-carboxygenistein/L. Using this method, a p38 protein was recovered from MCF-7 cells. N-terminal chemical sequencing of the first 30 residues of the protein revealed a peptide sequence similar to those that have been discovered in human tissues (a T-cell attractant protein from synovial fluid from patients with osteoarthritis and an analogous human skin fibroblast protein using a hirudin-affinity column) as well as a cotonine-binding protein from rat brain and related proteins in plants. In each case, the corresponding gene has not been found. In conclusion, although much of the human genome has been sequenced, novel proteins that are not described by genome data remain to be found. The DING protein (N-terminal amino acid sequence Asp-Ile-Asn-Gly) that binds to genistein with high affinity is one of these. Its biological role, however, remains to be defined.

Simultaneous determination of 11 phytoestrogens in human serum using a 2 min liquid chromatography/tandem mass spectrometry method

A rapid 2 min liquid chromatography-tandem mass spectrometry (LC-MS/MS) method operating in multiple reaction ion monitoring mode was developed and validated that allows for the characterization and simultaneous quantification of 11 phytoestrogen metabolites with mass transitions m/z 241/119 (equol), 253/132 (daidzein), 255/149 (dihydrodaidzein), 257/108 (O-desmethylangolesin), 269/133 (genistein), 283/184 (glycitein), 267/191 (formononetin), 289/109 (biochanin A), 267/91 (coumestrol), enterodiol (301/253), and enterolactone (297/253). The method was demonstrated to be specific and sensitive, and a linear response for each phytoestrogen was observed over a range of 1-5000 ng/mL in human serum with the exception of dihydrodaidzein, whose lower limit of quantification was 2 ng/mL. The separation was carried out on a Synergi Polar-RP 2.5 micron (50 mm x 2.0 mm i.d.) column at 50 degrees C with water and acetonitrile (both containing 10 mM ammonium acetate) as the mobile phase under gradient conditions at a flow rate of 0.75 mL/min. This LC-MS/MS method is very useful for high-throughput analysis of phytoestrogens and proved to be simple, sensitive, reproducible, and reliable.