2,3-Dimethoxy-10-oxostrychnidinium 2-(2,4,6-trinitroanilino)benzoate monohydrate : a 1:1 proton-transfer salt of brucine with o-picraminobenzoic acid

In the structure of the 1:1 proton-transfer compound of brucine with 2-(2,4,6-trinitroanilino)benzoic acid C23H27N2O4+ . C13H7N4O8- . H~2~O, the brucinium cations form the classic undulating ribbon substructures through overlapping head-to-tail interactions while the anions and the three related partial water molecules of solvation (having occupancies of 0.73, 0.17 and 0.10) occupy the interstitial regions of the structure. The cations are linked to the anions directly through N-H...O(carboxyl) hydrogen bonds and indirectly by the three water molecules which form similar conjoint cyclic bridging units [graph set R2/4(8)] through O-H...O(carbonyl) and O(carboxyl) hydrogen bonds, giving a two-dimensional layered structure. Within the anion, intramolecular N-H...O(carboxyl) and N H...O(nitro) hydrogen bonds result in the benzoate and picrate rings being rotated slightly out of coplanarity inter-ring dihedral angle 32.50(14)\%]. This work provides another example of the molecular selectivity of brucine in forming stable crystal structures and also represents the first reported structure of any form of the guest compound 2-(2,4,6-trinitroanilino)benzoic acid.

The collision induced loss of carbon monoxide from deprotonated benzyl benzoate in the gas phase. An anionic 1,2-Wittig type rearrangement

The ion PhCO2--CHPh, upon collision activation, undergoes competitive losses of CO and CO2 of which the former process produces the base peak of the spectrum. Product ion and substituent effect (Hammett) studies indicate that PhCO2--CHPh cyclises to a deprotonated hydroxydiphenyloxirane which ring opens to PhCOCH(O-)Ph. This anion then undergoes an anionic 1,2-Wittig type rearrangement {through [PhCO- (PhCHO)]} to form Ph2CHO- and CO. The mechanism of the 1,2-rearrangement has been probed by an ab initio study [at MP4(SDTQ)/6-31++G(d,p) level] of the model system HCOCH2O- →; MeO- + CO The analogous system RCO2--CHPh (R = alkyl) similarly loses CO, and the migratory aptitudes of the alkyl R groups in this reaction are Bu′ > Me > Et ∼Pri). This trend correlates with the order of anion basicities (i.e. the order of ΔG○acid values of RH), supporting the operation of an anion migration process. The loss of CO2 from PhCO2--CHPh yields Ph2CH- as the anionic product: several mechanistic scenarios are possible, one of which involves an initial ipso nucleophilic substitution.

The loss of CO from the ortho, meta and para forms of deprotonated methyl benzoate in the gas phase

The ortho, meta and para anions of methyl benzoate may be made in the source of a mass spectrometer by the S(N)2(Si) reactions between HO- and methyl (o-, m-, and p-trimethylsilyl)benzoate respectively. All three anions lose CO upon collisional activation to form the ortho anion of anisole in the ratio ortho>>meta > para. The rearrangement process is charge directed through the ortho anion. Theoretical calculations at the B3LYP/6-311++G(d,p)//HF/6-31+G(d) level of theory indicate that the conversion of the meta and para anions to the ortho anion prior to loss of CO involve 1,2-H transfer(s), rather than carbon scrambling of the methoxycarbonylphenyl anion. There are two mechanisms which can account for this rearrangement, viz. (A) cyclisation of the ortho anion centre to the carbonyl group of the ester to give a cyclic carbonyl system in which the incipient methoxide anion substitutes at one of the two equivalent ring carbons of the three membered ring to yield an intermediate which loses CO to give the ortho anion of anisole, and (B) an elimination reaction to give an intermediate benzyne-methoxycarbonyl anion complex in which the MeOCO- species acts as a MeO- donor, which then adds to benzyne to yield the ortho anion of anisole. Calculations at the B3LYP/6-311++G(d,p)//HF/6-31+G(d) level of theory indicate that (i) the barrier in the first step (the rate determining step) of process A is 87 kJ mol(-1) less than that for the synchronous benzyne process B, and (ii) there are more low frequency vibrations in the transition state for benzyne process B than for the corresponding transition state for process A. Stepwise process A has the lower barrier for the rate determining step, and the lower Arrhenius factor: we cannot differentiate between these two mechanisms on available evidence.

Time- and Temperature-Dependent Study in the Three-Component Zinc-Triazolate-Oxybis(benzoate) System: Stabilization of New Topologies

Three new three-dimensional zinc-triazolate-oxybis(benzoate) compounds. [{Zn-3(H2O)(2)}{C12H8O(COO)(2)}(2)-{C2H2N3}(2)]center dot 2H(2)O(I), [Zn-7{C12H8O(COO)(2)}(4){C2H2N3}(6)]center dot H2O, (II), and[{Zn-5(OH)(2)}{C12H8O(COO)(2)}(3){C2H2N3}(2)] (III), synthesized by a hydrothermal reaction of a mixture of Zn(OAc)(2)center dot 2H(2)O, 4,4'-oxybis(benzoic acid), 1,2,4-triazole, NaOH, and water. Compound I has an interpenetrated diamond structure and II and III have pillared-layer related structures. The formation of a hydrated phase (I) at low temperature and a completely dehydrated phase (III) at high temperature suggests the importance of thermodynamic factors in the formation of three compounds. Transformation studies of I in the presence of water shows the formation of a simple Zn-OBA compound, [Zn(OBA)(H2O)] (IV), at 150 and 180 degrees C and compound III at 200 degrees C. The compounds have been characterized by single-crystal X-ray diffraction, powder X-ray diffraction. thermogravimetric analysis, IR, and photoluminescence studies.

Follicle stimulating hormone secretion and dominant follicle growth during treatment of Bos indicus heifers with intra-vaginal progesterone releasing devices, oestradiol benzoate, equine chorionic gonadotrophin and prostaglandin F-2 alpha

The aim of this study was to investigate the effects on follicle stimulating hormone (FSH) secretion and dominant follicle (OF) growth, of treatment of Bos indicus heifers with different combinations of intra-vaginal progesterone releasing devices (IPRD), oestradiol benzoate (ODB), PGF(2 alpha), and eCG. Two-year-old Brahman (BN; n=30) and Brahman-cross (BNX; n=34) heifers were randomly allocated to three IPRD-treatments: (i) standard-dose IPRD [CM 1.56 g; 1.56 g progesterone (P-4); n = 17]; (ii) half-dose IPRD (CM 0.78 g; 0.78 g p(4); n=15); (iii) half-dose IPRD + 300 IU eCG at IPRD removal (CM 0.78 g+G; n=14); and, (iv) non-IPRD control (2 x PGF(2 alpha); n=18) 500 mu g cloprostenol on Days -16 and -2. IPRD-treated heifers received 250 mu g PGF(2 alpha) at IPRD insertion (Day 10) and IPRD removal (Day -2) and 1 mg ODB on Day -10 and Day -1. Follicular dynamics were monitored daily by trans-rectal ultrasonography from Day -10 to Day 1. Blood samples for determination of P-4 were collected daily and samples for FSH determination were collected at 12 h intervals from Day -9 to Day -2. A significant surge in concentrations of FSH was observed in the 2 x PGF(2 alpha), treatment 12 h prior and 48 h after follicular wave emergence, but not in the IPRD-treated heifers. Estimated mean concentrations of total plasma P-4 during the 8 days of IPRD insertion was greater (P<0.001) in the CM 1.56 g P-4 treated heifers compared to the CM 0.78 g P-4 treated heifers (18.38 ng/ml compared with 11.09 ng/ml, respectively). A treatment by genotype interaction (P=0.036) was observed in the mean plasma P4 concentration in heifers with no CL during IPRD insertion, whereby BN heifers in the CM 1.56 g treatment had greater plasma P-4 than the BNX heifers on Days-9, -7, -6, -5, and -4. However, there was no genotype effect in the CM 0.78 g +/- G or the 2 x PGF(2 alpha) treatment. Treatment had no effect on the DF growth from either day of wave emergence (P=0.378) or day of IPRD removal (P=0.780) to ovulation. This study demonstrates that FSH secretion in B. indicus heifers treated with a combination of IPRD's and ODB to synchronise ovulation was suppressed during the period of IPRD insertion but no significant effect on growth of the DF was observed. (C) 2013 Elsevier B.V. All rights reserved.

Animal-Level Factors Affecting Ovarian Function in Bos indicus Heifers Treated to Synchronize Ovulation with Intravaginal Progesterone-Releasing Devices and Oestradiol Benzoate

The primary objective of this study was to investigate the impact of animal-level factors including energy balance and environmental/management stress, on the ovarian function of Bos indicus heifers treated to synchronize ovulation. Two-year-old Brahman (BN) (n = 30) and BN-cross (n = 34) heifers were randomly allocated to three intravaginal progesterone-releasing device (IPRD) treatment groups: (i) standard-dose IPRD [Cue-Mate (R) (CM) 1.56 g; n = 17]; (ii) half-dose IPRD [0.78 g progesterone (P4); CM 0.78 g; n = 15]; (iii) half-dose IPRD + 300 IU equine chorionic gonadotrophin at IPRD removal (CM 0.78 g + G; n = 14); (iv) and a control group, 2x PGF2a [500 mu g prostaglandin F2a (PGF2a)] on Day -16 and -2 (n = 18). Intravaginal progesterone-releasing device-treated heifers received 250 mu g PGF2a at IPRD insertion (Day -10) and IPRD removal (Day -2) and 1 mg oestradiol benzoate on Day -10 and -1. Heifers were managed in a small feedlot and fed a defined ration. Ovarian function was evaluated by ultrasonography and plasma P4 throughout the synchronized and return cycles. Energy balance was evaluated using plasma insulin-like growth factor 1 (IGF-I) and glucose concentrations. The impact of environmental stressors was evaluated using plasma cortisol concentration. Heifers that had normal ovarian function had significantly higher IGF-I concentrations at commencement of the experiment (p = 0.008) and significantly higher plasma glucose concentrations at Day -2 (p = 0.040) and Day 4 (p = 0.043), than heifers with abnormal ovarian function. There was no difference between the mean pre-ovulatory cortisol concentrations of heifers that ovulated or did not ovulate. However, heifers that ovulated had higher cortisol concentrations at Day 4 (p = 0.056) and 6 (p = 0.026) after ovulation than heifers that did not ovulate.

A field investigation of a modified intravaginal progesterone releasing device and oestradiol benzoate based ovulation synchronisation protocol designed for fixed-time artificial insemination of Brahman heifers

Pregnancy rates (PR) to fixed-time AI (FTAI) in Brahman heifers were compared after treatment with a traditional oestradiol-based protocol (OPO-8) or a modified protocol (OPO-6) where the duration of intravaginal progesterone releasing device (IPRD) was reduced from 8 to 6 days, and the interval from IPRD removal to oestradiol benzoate (ODB) was increased from 24 to 36 h. Rising 2 yo heifers on Farm A: (n = 238 and n = 215; two consecutive days AI); B (n = 271); and C (n = 393) were allocated to OPO-8 or OPO-6. An IPRD was inserted and 1 mg ODB i.m. on Day 0 for OPO-8 heifers and Day 2 for OPO-6 heifers. On Day 8, the IPRD was removed and 500 μg cloprostenol i.m. At 24 h, for OPO-8 heifers, and 36 h, for OPO-6 heifers, post IPRD removal all heifers received 1 mg ODB i.m. FTAI was conducted at 54 and 72 h post IPRD removal for OPO-8 and OPO-6 heifers. At Farm A, OPO-6 heifers, AI on the second day, the PR was 52.4 to FTAI (P = 0.024) compared to 36.8 for OPO-8 heifers. However, no differences were found between OPO-8 and OPO-6 protocols at Farm A (first day of AI) (39.9 vs. 35.7), or Farms B (26.2 vs. 35.4) and C (43.2 vs. 40.3). Presence of a corpus luteum at IPRD insertion affected PR to FTAI (43.9 vs. 28.8; P < 0.001). This study has shown that the modified ovulation synchronisation protocol OPO-6 may be a viable alternative to the OPO-8 protocol for FTAI in B. indicus heifers.

Purification and properties of benzoate-4-hydroxylase from a soil pseudomonad

Benzoate-4-hydroxylase from a soil pseudomonad was isolated and purified about 50-fold. Polyacrylamide gel electrophoresis of this enzyme preparation showed one major band and one minor band. The approximate molecular weight of the enzyme was found to be 120,000. Benzoate-4-hydroxylase was most active around pH 7.2. The enzyme showed requirements for tetrahydropteridine as the cofactor and molecular oxygen as the electron acceptor. NADPH, NADH, dithiothreitol, β-mercaptoethanol, and ascorbic acid when added alone to the reaction mixture did not support the hydroxylation reaction to any significant extent. However, when these compounds were added together with tetrahydropteridine, they stimulated the hydroxylation. This stimulation is probably due to the reduction of the oxidized pteridine back to the reduced form. This enzyme was activated by Fe2+ and benzoate. It was observed that benzoate-4-hydroxylase could catalyze the oxidation of NADPH in the presence of benzoate,p-aminobenzoate, p-nitrobenzoate, p-chlorobenzoate, and p-methylbenzoate, with only benzoate showing maximum hydroxylation. Inhibition studies with substrate analogs and their kinetic analysis revealed that the carboxyl group is involved in binding the substrate to the enzyme at the active center. The enzyme catalyzed the conversion of 1 mol of benzoate to 1 mol of p-hydroxybenzoate with the consumption of slightly more than 1 mol of NADPH and oxygen.

Purification, properties and induction of a specific benzoate-4-hydroxylase from Aspergillus niger (UBC 814)

An inducible benzoate-4-hydroxylase has been partially purified from crude extracts of the mycelial felts of Aspergillus niger. This enzyme catalyzes the transformation of benzoate to p-hydroxybenzoate with equimolar consumption of NADPH and O2. It requires tetrahydropteridine as a prosthetic group. The optimum activity was found at pH 6.2 with a Km value at 30°C of 1.6 · 10−4 M for NADPH and 1.3 · 10−4 M for benzoate. Fe2+ (iron) is required for the enzyme activity. The enzyme is stabilized by the inclusion of benzoate, EDTA and glutathione in the extracting buffer. The enzyme is specific for benzoate as substrate. Sulfhydryl group(s) are essential for enzyme activity as indicated by p-chloromercuri-benzoate and N-ethylmaleimide inactivation. Benzoate-4-hydroxylase activity is decreased in the mycelial felts of Aspergillus niger grown in the presence of higher concentrations of benzoate. Maximum activity of the enzyme was observed at 36 h after inoculation.

Detoxication of benzoate by glycine conjugation in the silkworm, Bombyx mori L

The role of hippuric acid formation as a mechanism for detoxication of benzoic acid in the silkworm has been investigated. Benzoate is inhibitory to the growth of the silkworm and excreted as hippuric acid, which is not toxic. Hippuric acid is not a normal constituent of excreta. Synthesis of hippuric acid has been shown to occur in the intestines of the silkworm. Hippuricase activity is present in the fat body and silk-gland tissue.

Chemoenzymatic synthesis of the carbasugars carba-beta-L-galactopyranose, carba-beta-L-talopyranose and carba-alpha-L-talopyranose from methyl benzoate

The cis-dihydrodiol metabolite from methyl benzoate has been used as a synthetic precursor of carba-beta-L-galactopyranose, carba-beta-L-talopyranose and carba-alpha-L-talopyranose. The structures and absolute configurations of these carbasugars were determined by a combination of NMR spectroscopy, stereochemical correlation and X-ray crystallography.

Isotopic composition of inorganic carbon as an indicator of benzoate degradation by Pseudomonas putida: temperature, growth rate and pH effects.

Degradation experiments of benzoate by Pseudomonas putida resulted in enzymatic carbon isotope fractionations. However, isotopic temperature effects between experiments at 20 and 30 °C were minor. Averages of the last three values of the CO2 isotopic composition (δ13CCO2(g)) were more negative than the initial benzoate δ13C value (−26.2‰ Vienna Pee Dee Belenite (VPDB)) by 3.8, 3.4 and 3.2‰ at 20, 25 and 30 °C, respectively. Although the maximum isotopic temperature difference found was only 0.6‰, more extreme temperature variations may cause larger isotope effects. In order to understand the isotope effects on the total inorganic carbon (TIC), a better measure is to calculate the proportions of the inorganic carbon species (CO2(g), CO2(aq) and HCO3−) and to determine their cumulative δ13CTIC. In all three experiments δ13CTIC was more positive than the initial isotopic composition of the benzoate at a pH of 7. This suggests an uptake of 12C in the biomass in order to match the carbon balance of these closed system experiments.

Studies on the persistence of estradiol benzoate and nortestosterone decanoate in hair of cattle following treatment with growth promoters, determined by ultra-high-performance liquid chromatography-tandem mass spectrometry

Measurement of steroid esters in bovine hair samples, using sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS), provides a powerful tool for identifying animals treated illicitly with growth promoters. The successful application of such testing requires appropriate sampling of hair from treated animals. This paper describes the results of hair analysis by LC-MS/MS for two animal studies in which animals were treated with estradiol-3-benzoate and nortestosterone decanoate. The results from the first animal study indicate that animals treated with these anabolic steroids may not always be identified from analysis of hair samples; positive test results occur sporadically and only for some of the treated animals. The results from the second animal study identify conditions attaching to positive hair samples, such as, that concentrations of steroid esters in hair are related to distance of sampling from point of injection and to time post-treatment, that concentrations of steroid esters in hair are related to dose given to the animal but that this relationship may vary over time post-treatment, and that steroid esters may be measured in regrowth hair taken some weeks after treatment. Steroid esters are determined along the length of the hair, confirming that accumulation of steroid esters into hair occurs from various sources, including blood, sweat and sebum. The reported research provides some useful insights into the mechanisms governing the persistence of steroid esters in bovine hair following illicit treatment with growth promoters. (C) 2009 Elsevier B.V. All rights reserved.

Variation in stable carbon isotope fractionation during aerobic degradation of phenol and benzoate by contaminant degrading bacteria

Variation in the natural abundance stable carbon isotope composition of respired CO2 and biomass has been measured for two types of aerobic bacteria found in contaminated land sites. Pseudomonas putida strain NCIMB 10015 was cultured on phenol and benzoate and Rhodococcus sp. I-1 was cultured on phenol. Results indicate that aerobic isotope fractionations of differing magnitudes occur during aerobic biodegradation of these substrates with an isotopic depletion in the CO2 (Delta(13)C(phenol-CO2)) as much as 3.7 parts per thousand and 5.6 parts per thousand for Pseudomonas putida and Rhodococcus sp. I-1 respectively. This observation has significant implications for the use of a stable isotope mass balance approach in monitoring degradation processes that rely on indigenous bacterial populations. The effects of the metabolic pathway utilised in degradation and inter-species variation on the magnitude of isotope fractionation are discussed. Possible explanations for the observed isotope fractionation include differences in the metabolic pathways utilised by the organisms and differences in specific growth rates and physiology. (C) 1999 Elsevier Science Ltd. All rights reserved.