A bioactive biflavonoid from Campnosperma panamense

Lanaroflavone (1), a biflavonoid isolated from the methanol extract of the aerial part of Campnosperma panamense by bioguided fractionation, has been assessed for in vitro antiprotozoal activity. Lanaroflavone showed both antimalarial and leishmanicidal activities, but was inactive against Chagas disease vector, Trypanosoma cruzi.     

Authentication of natural vanilla flavorings: isotopic characterization using degradation of vanillin into guaiacol

The isotopic investigation of vanillin has been extended to the new sources of natural precursors of vanillin recently introduced with a view of obtaining natural vanillin by biotechnological processes. To check the consistency of the isotopic composition of vanillin with that of the corresponding aromatic fragment, a selective degradation reaction into guaiacol was carried out. The reaction was shown to proceed without significant isotopic fractionation at the sites of interest, and an optimized procedure was defined from the results of an experimental design involving the quantity of reagent and the temperature and duration of the experiment. Guaiacol, which can be easily obtained in a reasonable time, is an interesting isotopic probe for carbon- and oxygen-isotope ratio mass spectrometry (IRMS) determinations. It provides (13)C information specific to the aromatic fragment and, combined with delta(13)C values measured on vanillin itself, it improves the authentication potential of carbon-IRMS. Thus, the natural status of ferulic acid may be characterized by significant (13)C depletion at the formyl site. Similarly, the oxygen-18 content of guaiacol is a better authentication tool than delta(18)O of vanillin because it does not suffer the drawback of being altered by chemical exchange of the sp(2) oxygen atom with water in industrial or laboratory procedures. Although collaborative studies are still necessary to improve the interlaboratory reproducibility of the delta(18)O parameters, consistent results can be obtained in an intralaboratory context. It is shown in particular that chemical oxidation of ferulic acid is characterized by a relative enrichment of the aromatic moiety of vanillin.     

Inhibition of reactive nitrogen species in vitro and ex vivo by trypsin inhibitor from sweet potato 'Tainong 57' storage roots

Peroxynitrite (ONOO-), formed from a reaction of superoxide and nitric oxide, is one of the most potent cytotoxic species known to oxidize cellular constituents including essential proteins, lipids, and DNA. ONOO- induces cellular and tissue injury, resulting in several human diseases such as Alzheimer's disease, atherosclerosis, and stroke. Due to the lack of endogenous enzymes responsible for ONOO- scavenging activity, finding a specific ONOO- scavenger is of considerable importance. In this study, the ability of trypsin inhibitor (TI), isolated from sweet potato storage roots (SPTI), to scavenge *ON and ONOO- was investigated. The data obtained show that TI generated a dose-dependent inhibition on production of nitrite and superoxide radicals. The IC50 value of TI on superoxide radical was 143.2 +/- 4.29 microg/mL. SOD activity staining was used to confirm SOD activity of SPTI. SPTI also caused a dose-dependent inhibition of the oxidation of dihydrorhodamine 123 (DHR) by peroxynitrite. A calculated IC50 value of 809.1 +/- 32.36 microg/mL was obtained on the inhibition of peroxynitrite radical. Spectrophotometric analyses revealed that TI suppressed the formation of ONOO--mediated tyrosine nitration through an electron donation mechanism. In further studies, TI also showed a significant ability to inhibit nitration of bovine serum albumin (BSA) in a dose-dependent manner. In vivo TI inhibited lipopolysaccharide-induced nitrite production in macrophages in a concentration-dependent manner with an IC50 value of 932.8 +/- 29.85 microg/mL. The present study suggested that TI had an efficient reactive nitrogen species scavenging ability. TI might be a potential effective NO and ONOO- scavenger useful for the prevention of NO- and ONOO--involved diseases.     

Nonfeed application of rendered animal proteins for microbial production of eicosapentaenoic acid by the fungus Pythium irregulare

Rendered animal proteins are well suited for animal nutrition applications, but the market is maturing, and there is a need to develop new uses for these products. The objective of this study is to explore the possibility of using animal proteins as a nutrient source for microbial production of omega-3 polyunsaturated fatty acids by the microalga Schizochytrium limacinum and the fungus Pythium irregulare. To be absorbed by the microorganisms, the proteins needed to be hydrolyzed into small peptides and free amino acids. The utility of the protein hydrolysates for microorganisms depended on the hydrolysis method used and the type of microorganism. The enzymatic hydrolysates supported better cell growth performance than the alkali hydrolysates did. P. irregulare displayed better overall growth performance on the experimental hydrolysates compared to S. limacinum. When P. irregulare was grown in medium containing 10 g/L enzymatic hydrolysate derived from meat and bone meal or feather meal, the performance of cell growth, lipid synthesis, and omega-3 fatty acid production was comparable to the that of culture using commercial yeast extract. The fungal biomass derived from the animal proteins had 26-29% lipid, 32-34% protein, 34-39% carbohydrate, and <2% ash content. The results show that it is possible to develop a nonfeed application for rendered animal protein by hydrolysis of the protein and feeding to industrial microorganisms which can produce omega-3 fatty acids for making omega-3-fortified foods or feeds.     

Glutathione-dependent biotransformation of the fungicide chlorothalonil

A gene responsible for the chlorothalonil biotransformation was cloned from the chromosomal DNA of Ochrobactrum anthropi SH35B, capable of efficiently dissipating the chlorothalonil. The gene encoding glutathione S-transferase (GST) of O. anthropi SH35B was expressed in Escherichia coli, and the GST was subsequently purified by affinity chromatography. The fungicide chlorothalonil was rapidly transformed by the GST in the presence of glutathione. LC-MS analysis supported the formation of mono-, di-, and triglutathione conjugates of chlorothalonil by the GST. The monoglutathione conjugate was observed as an intermediate in the enzymatic reaction. The triglutathione conjugate has not been previously reported and seems to be the final metabolite in the biotransformation of chlorothalonil. The glutathione-dependent biotransformation of chlorothalonil catalyzed by the bacterial GST is reported.     

Study of the influence of alcoholic fermentation and distillation on the oxygen-18/oxygen-16 isotope ratio of ethanol

A laboratory procedure for the analysis of the oxygen-18/oxygen-16 isotope ratios of ethanol derived from sugars and fruit juices by pyrolysis-isotope ratio mass spectrometry (IRMS) has been applied to the study of isotopic fractionation induced by the isotope effects of fermentation and distillation. For both processes, an experimental model has been established to describe and explain the observed fractionation phenomena. It is shown that reproducible results can be obtained when appropriate analytical conditions are used. Moreover, the ability of ethanol to act as a reliable indicator of the (18)O/(16)O ratio of sugars in orange juice (and therefore to be used as an internal reference for detecting water addition) is demonstrated both in theory and in practice.     

Antioxidant capacity of oat (Avena sativa L.) extracts. 1. Inhibition of low-density lipoprotein oxidation and oxygen radical absorbance capacity

Milled oat groat pearlings, trichomes, flour, and bran were extracted with methanol and the fractions tested in vitro for antioxidant capacity against low-density lipoprotein (LDL) oxidation and R-phycoerythrin protein oxidation in the oxygen radical absorbance capacity (ORAC) assay. The oxidative reactions were generated by 2,2'-azobis(2-amidinopropane) HCl (AAPH) or Cu(2+) in the LDL assay and by AAPH or Cu(2+) + H(2)O(2) in the ORAC assay and calibrated against a Trolox standard to calculate Trolox equivalents (1 Trolox equivalent = 1 TE = activity of 1 micromol of Trolox). The antioxidant capacity of the oat fractions was generally consistent with a potency rank of pearlings (2.89-8.58 TE/g) > flour (1.00-3.54 TE/g) > trichome (1.74 TE/g) = bran (1.02-1.62 TE/g) in both LDL and ORAC assays regardless of the free radical generator employed. A portion of the oat antioxidant constituents may be heat labile as the greatest activity was found among non-steam-treated pearlings. The contribution of oat tocols from the fractions accounted for <5% of the measured antioxidant capacity. AAPH-initiated oxidation of LDL was inhibited by the oat fractions in a dose-dependent manner, although complete suppression was not achieved with the highest doses tested. In contrast, Cu(2+)-initiated oxidation of LDL stimulated peroxide formation with low oat concentrations but completely inhibited oxidation with higher doses. Thus, oats possess antioxidant capacity most of which is likely derived from polar phenolic compounds in the aleurone.     

Identification of 5,7,3',4'-tetramethoxyflavone metabolites in rat urine by the isotope-labeling method and ultrahigh-performance liquid chromatography-electrospray ionization-mass spectrometry

5,7,3',4'-Tetramethoxyflavone (TMF), one of the major polymethoxyflavones (PMFs) isolated from Kaempferia parviflor , has been reported possessing various bioactivities, including antifungal, antimalarial, antimycobacterial, and anti-inflammatory activities. Although several studies on the TMF have been reported, the information about the metabolism of TMF and the structures of TMF metabolites is still not yet clear. In this study, an isotope-labeling method was developed for the identification of TMF metabolites. Three isotope-labeled TMFs (5,7,3',4'-tetramethoxy[3'-D(3)]flavone, 5,7,3',4'-tetramethoxy[4'-D(3)]flavone, and 5,7,3',4'-tetramethoxy[5,4'-D(6)]flavone) were synthesized and administered to rats. The urine samples were collected, and the main metabolites were monitored by ultrahigh-performance liquid chromatography-electrospray ionization-mass spectrometry. Five TMF metabolites were unambiguously identified as 3'-hydroxy-5,7,4'-trimethoxyflavone, 7-hydroxy-5,3',4'-trimethoxyflavone sulfate, 7-hydroxy-5,3',4'-trimethoxyflavone, 4'-hydroxy-5,7,3'-trimethoxyflavone, and 5-hydroxy-7,3',4'-trimethoxyflavone.     

Role of aldehyde oxidase in the hepatic in vitro metabolism of 3-methylindole in pigs

The metabolism of 3-hydroxy-3-methylindolenine (HMI), a recently discovered metabolite of 3-methylindole (3MI, skatole) produced by porcine liver microsomes, was investigated in vitro using porcine liver cytosol. HMI was rapidly metabolized to a single product, 3-hydroxy-3-methyloxindole (HMOI), by porcine cytosol. By the use of the selective inhibitors menadione and quinacrine, it was shown that the enzyme responsible for the oxidation of HMI into HMOI was aldehyde oxidase (AO; aldehyde:oxygen oxidoreductase, EC 1.2.3.1). The activity of AO in the conversion of HMI to HMOI was measured in a population of pigs (n = 30) with a wide range of 3MI levels in back fat (0.07-0.30 mg/kg). AO activity was found to be negatively correlated (r = -0.70; P < 0.001) with the level of 3MI in fat. The results of the present study suggest that AO plays an important role in the metabolism of 3MI in the pig and that its catalytic activity is related to an adequate 3MI clearance.