Omne Ignotum pro Magnifico: characterization of commercial Bilberry extracts to fight adulteration.

Thanks to their chemical composition, extracts from the bilberry are commonly used to manufacture food, health products, supplements and cosmetics. However, in this field, as in others, "the process is the product" and a careful characterization of the entire supply and production chain, from purity and quality of raw material to extraction procedures needs to be implemented by using validated, sensitive and specific techniques of analysis. This position paper discusses the importance of the characterization of bilberry extracts, in order to fight adulteration.

Effects of dietary vitamin E, C and soybean oil supplementation on antioxidant enzyme activities in liver and muscles of rats.

The effect of elevated levels of dietary vitamin E, C and a combination of vitamin E and C (E&C) with soybean oil on activities of antioxidant (AOE) enzymes important in the protection against lipid peroxidation was studied in male rats fed with vitamin C (12 mg/g), vitamin E (3.68 mg/g) or E&C (3.68 mg/kg+12 mg/g) supplemented diets for 28 days. Catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR) activity in liver, pectoralis major (PM) and sartorius (S) muscles was increased significantly in rats fed with dietary vitamin C, E separately, and vitamin C&E combination, except, superoxide dismutase (SOD), which showed no alterations. These results clearly indicated that vitamin E&C separately and E&C together increased AOE activity in liver, PM and S muscle of rats. However, vitamin E and C combination enhanced AOE activity more significantly and our findings suggest the possible role of vitamin C&E and their combination in reducing the risk of chronic diseases related to oxidative stress.

Effect of oral or transdermal hormone replacement therapy on homocysteine levels: a randomized clinical trial.

OBJECTIVE: Aim of this randomized trial was evaluate the effect on homocysteine plasma levels of two different hormone replacement therapy (HRT) formulations in a group of late postmenopausal women. METHODS: Eligible for this study were women: in postmenopause since 5 years or more (confirmed from FSH level > or = 40 mIU/l); with body mass index (BMI) < or = 35; without endocrine, hepatic or renal diseases; not current users of vitamin B or folic acid supplements; not users of any lipid-lowering drugs and sex steroids in the 6 months before trial entry. Group A: oral estradiol valerate 2 mg per day per oral normegestrol acetate 2.5 mg per day (n = 98) for 12 months; Group B: a weekly patch releasing estradiol (50 microg per day) per oral normegestrol acetate 2.5 mg per day (n = 101) for 12 months. RESULTS: The mean values of the homocysteine levels in the group A and B at baseline, 3, 6 and 12 months were 7.9 and 9.1, 8.7 and 8.9, 9.3 and 10.2, 9.6 and 10.2, respectively, the differences between the two treatments were not statistically significant (time by treatment interaction, P = 0.32). Otherwise, the changes of homocysteine level at the four visits was statistically significant (P = 0.0001) in both groups. In particular, in the oral treatment group homocysteine levels increased from baseline of 10.5% at 3 months, of 17.2% after 6 months of therapy and of 21.9% at the end of the study; in the transdermal group, after a little decrease at 3 months (1.5%), the increases were of 12.1 and 12.9%, respectively. CONCLUSIONS: This study does not show any different effect of oral and transdermal treatment with estradiol plus normegestrol acetate on homocysteine levels. Further it does not support previous suggestion of a lowering effect of HRT on plasma homocysteine.

A simple HPLC-UV method for the assay of ginkgolic acids in Ginkgo biloba extracts.

Extracts from the leaves of Ginkgo biloba are among the most widely used phytotherapeutics. Some alkylphenols (ginkgolic acids, cardanols and cardols) have been described as potentially hazardous constituents in Ginkgo extracts. Accordingly, a requirement for a maximum concentration of ginkgolic acids has been proposed in the UE and US pharmacopoeias Ginkgo monographs establishing a limit value of 5 ppm. A novel HPLC-UV method, developed by the use of HPLC-APCI-MS HPLC-DAD techniques and allowing the identification of ginkgolic acids and related phenols, is described. The new analytical method, not requiring enrichment procedures, can be used for the quantification of ginkgolic acids in Ginkgo extracts.

Identification of Ruscus steroidal saponins by HPLC-MS analysis.

A novel HPLC-UV method has been developed for the fingerprint analysis of the steroidal saponins in the rhizomes of three Ruscus species (Ruscus aculeatus, Ruscus hypoglossum and Ruscus colchicus). Saponins were identified by HPLC-ESI-MS. During the study a new major saponin was detected in the rhizomes of R. hypoglossum and R. colchicus. The structure of the new compound was defined as 1-O-[alpha-L-rhamnopyranosyl-(1-->2)-6-O-acetyl-beta-D-galactopyranosyl]-1beta,3beta,22xi,26-tetrahydroxy-furost-5(6)-en-26-O-beta-D-glucopyranoside (8) by spectral analysis.

Determination of ginsenosides in Panax ginseng roots by liquid chromatography with evaporative light-scattering detection.

A liquid chromatographic method was developed and validated for the determination of ginsenosides in Panax ginseng roots by using evaporative light-scattering detection. Eighteen ginsenosides were separated on a reversed-phase C18 column with water-ammonium acetate-acetonitrile as the mobile phase. The method is suitable for the routine determination of ginsenosides in P. ginseng roots and extracts. The validation of the method was comprehensive for efficiency and recovery optimization of the P. ginseng roots extraction, specificity by liquid chromatography/mass spectrometry, linearity, stability, reproducibility, repeatability, intermediate precision, and robustness.

Characterization of proanthocyanidins from grape seeds.

Leucoselect (grape seed selected proanthocyanidins) was analyzed. HPLC thermospray mass spectrometry (TSP-MS) allowed the detection of monomeric flavan-3-ols and dimeric proanthocyanidins. Fractionation over Sephadex LH-20 resin and analysis of the isolated fractions by gel permeation chromatography (GPC) and electrospray mass spectrometry (ESI-MS) led to the complete characterization of the proanthocyanidin constituents of Leucoselect. The analysis revealed the presence of approximately 15% of (+)-catechin (1) and (-)-epicatechin (2), 80% of (-)-epicatechin 3-O-gallate (3), dimers, trimers, tetramers and their gallates and 5% of pentamers, hexamers, heptamers and their gallates.

Liquid chromatography-electrospray mass spectrometric identification of ginsenosides in Panax ginseng roots.

A high-performance liquid chromatographic method was developed for electrospray mass spectrometric analysis of ginsenosides in Panax ginseng roots. The analyses were performed on a reversed-phase C18 column using a binary eluent (aqueous 8 mM NH4OAc, buffered to pH 7 with NH4OH-acetonitrile) under gradient conditions. Twenty-five ginsenosides could be separated and detected. The mass spectra obtained provided information on their molecular masses. A MS-MS experiment was undertaken in order to determine the sugar unit sequences and the aglycone moieties.

Proximate composition and mineral content of selected Tanzanian vegetables and the effect of traditional processing on the retention of ascorbic acid, riboflavin and thiamine.

Proximate composition, mineral content and the effect of traditional processing practices on the retention of ascorbic acid, riboflavin and thiamine were studied using amaranth, cowpea, peanut, pumpkin and sweetpotato leaves. Results of this study indicated that, crude protein, crude fat, carbohydrate and ash contents were in the range of 20.64--46.56 percent, 2.57--4.34 percent, 35.43--63.50 percent and 8.92--15.69 percent respectively. The mineral content per 100 g of fresh vegetables was in the range of 83.64--229.34 mg, 145.97--780.19 mg, 11.5--2l.31 mg, 43.02--110.30 mg, 0.96--5.90 mg and 0.40--2.24 mg for Ca, K, Na, Mg, Fe and Zn respectively. For ascorbic acid, riboflavin and thiamine, concentrations in 100 g of fresh vegetables were in the range of 43.78--89.00 mg, 0.62--1.71 mg and 0.09--0.30 mg respectively. The traditional processing practices of sun/shade drying and storing in ventilated containers resulted in a significant (P <0.05) decrease in ascorbic acid, riboflavin and thiamine for all vegetables. Conventional blanching and cooking for up to 15 minutes resulted in a significant (P <0.05) increase in riboflavin content in cowpea, peanut and pumpkin greens while in amaranth and sweetpotato leaves, thermal processing resulted in a significant (P <0.05) decrease in the vitamin. Based on the results of this study, the vegetables were good dietary sources of minerals, carbohydrate and protein.

Effect of blanching on the content of antinutritional factors in selected vegetables.

The effect of blanching on the antinutritional content was studied in cabbage, turnip, collard, sweetpotato and peanut leaves. All the vegetables contained various amounts of phytic acid, tannic acid and/or oxalic acid. Tannic acid was found in largest amounts ranging from 1266.00 mg/100 g in cabbage to 491.00 mg/100 g in sweetpotato. Phytic acid content ranged from 0.31 mg/100 g in sweetpotato to 3.97 mg/100 g in collard. Oxalic acid was in trace amounts in cabbage and turnip; but high concentrations were found in sweetpotato (469.67 mg/100 g) and peanut greens (407.00 mg/100 g). Levels of both tannic acid and phytic acid were significantly (p < 0.05) reduced by conventional and microwave blanching methods while oxalic acid levels were not significantly (p < 0.05) reduced in most of the treatments by either of the blanching methods. In general, blanching is recommended as an effective method for reducing the antinutritional factors in green vegetables, however, further investigation on the heating times for both conventional and microwave blanching methods has been suggested.

Chronomodulated 5-day infusion of floxuridine and L-folinic acid in patients with advanced malignancies: a feasibility and tolerability study.

The best schedule for administering floxuridine (FUDR) has not yet been established. Duration of infusion, need (and dosage) of leucovorin (folinic acid, FA), and circadian timing need to be further specified. Nevertheless, FUDR delivery according to circadian rhythms has allowed increase of dose intensity without enhancing the side effects. A 5-day infusional schedule combining FUDR and L-FA was devised as an attempt to increase dose intensity and to provide therapy every 3 weeks to patients with advanced cancer. An ambulatory programmable-in-time pump was used for this purpose. Fourteen patients entered this trial. Two dose levels (mg/kg x 5 days) were evaluated: 0.5 mg/kg/day in six patients and 0.525 mg/kg/day in eight patients. Both patient groups received a concurrent infusion of L-FA 10 mg/m2/day i.v. The delivery patterns of both FUDR and L-FA varied sinusoidally during the 24 hours with a maximum at 18.00 hours. Courses were repeated every 3 weeks. Of 35 courses, treatment produced mucosites greater than grade 2 in only two of them. No severe diarrhea, the dose-limiting toxicity of FUDR when infused over 14 days, was encountered at the dose levels tested. This 5-day chronotherapy schedule allowed delivery of a larger amount of FUDR than the flat delivery described in a previous report. A daily dose of 0.525 mg/kg FUDR, combined with 10 mg/mg2 L-FA, with intraindividual dose escalation according to tolerance, is recommended for future investigations of the activity of this chronotherapy schedule.

Establishment of a cell line from a hepatocellular carcinoma from a patient with hemochromatosis.

We describe the establishment and characterization of a novel hepatoma cell line. This cell line, designated RBHF-1, was established from a hepatocellular carcinoma of a 67-yr-old man with a history of genetic hemochromatosis. At this writing, the cells have been maintained in RPMI-1640 tissue-culture medium and fetal calf serum without any additional supplements for 30 mo. The cells form colonies on soft agar and are not tumorigenic in nude mice. The cell line is polymorphic and displays characteristics of mature hepatocytes by synthesizing albumin, alpha 2-macroglobulin, fibronectin and alpha-fetoprotein. Cytogenetic analysis shows multiple chromosomal aberrations, with a consistent deletion in the long arm and deletions or rearrangements in the short arm of chromosome 1. There is no evidence for hepatitis B or hepatitis C virus infection of the cell line. The cells contain no detectable intracellular iron after staining with Perls' stain. Unlike other hepatoma cell lines, there is no detectable binding of epidermal growth factor to RBHF-1 cells. This is the first cell line to be established from a patient with hemochromatosis, and it provides a potentially important model for the study of hepatocyte transformation in association with iron overload.

Antitumor effects of a normal human serum factor (normal human globulin fraction 1) on human tumor cells in vitro.

Normal human globulin fraction 1 (NHG-1) produced cytotoxicity and/or cytostasis as well as inhibition of protein synthesis in 8 well-characterized human tumor cell lines (4 breast cancer, 1 colon cancer, 1 melanoma, and 2 leukemia) and in 2 variants of murine B-16 melanoma. NHG-1 was not cytotoxic for the Chang liver cell line, a normal kidney embryo line, or for normal lymphocytes or macrophages when used in lower concentrations but was growth inhibitory for normal cells in higher concentrations. Although lymphocyte blastogenesis with phytohemagglutinin (PHA) was inhibited by high concentrations of NHG-1, augmentation of the lymphocyte PHA response was seen at lower concentrations, suggesting a lymphokine-like effect. Preincubation with the mitogen partially nullified these NHG-1 effects (suggesting the need for cell surface binding). Although NHG-1 antitumor activity was confirmed in selected human and murine tumor cell lines, the mechanism of its activity is unknown. Occurrence of NHG-1 in the alpha 2-globulin region (an area rich in immune-regulating factors) suggests that NHG-1 may have general "cytokine"-like effects and may be capable of regulating replication of both normal and transformed cells.