Some studies of the active intermediates formed in the microsomal metabolism of cyclophosphamide and isophosphamide.
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The effects of polymeric polyphosphonate and of ethane hydroxydiphosphonate on the development of ectopic bone were compared at different stages of bone development. The diphosphonate affected bone development at different stages, as reflected by alkaline phosphatase and aryl sulphatase activities, as well as by the creation of calcium ions by bone and on its readiness to undergo isotopic exchange. The polymeric polyphosphonate (mol. wt., 3500-8000), on the other hand, did not exhibit any of these effects, although it did inhibit the activity of the enzymes in vitro to an extent comparable with that of the diphosphonate. The results corroborate the assumption that the effects of polymeric polyphosphonates on hard tissues are confined to the extracellular space while the effects of diphosphonates on bone development are due to intracellular activity and not to sequestering of extracellular calcium.
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We have refined a method for measuring erythrocyte volume using the 31P-NMR spectrum of a probe molecule, dimethyl methylphosphonate. This compound, when added to an erythrocyte suspension, gives rise to two 31P-NMR resonances, and the frequency separation between them is linearly dependent on the intracellular haemoglobin concentration. If, for a given cell sample (under standard conditions), the separation of the two dimethyl methylphosphonate peaks has been measured and an independent estimation of the mean cell haemoglobin content and concentration has been obtained, then changes in the mean cell volume due to altered experimental conditions may be estimated from the peak separation measured under the new conditions. Although the peak separation was independent of extracellular pH, it did vary with (i) a range of extracellular suspension media, (ii) temperature, (iii) dimethyl methylphosphonate concentration, (iv) haemoglobin ligand state and (v) different blood donors.
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Collagenase-isolated pancreatic islets of non-inbred ob/ob mice, containing more than 90% beta-cells, were labelled with radioactive orthophosphate (32P or 33P) and then subjected to non-recirculating perifusion. The basal D-glucose concentration in the perifusion medium was 2.8 mM. When the concentration was suddenly raised to 5.6, 8.3 or 16.7 mM, D-glucose promptly elicited a transient and dose-dependent release of radiophosphate. In the presence of 2.8 mM D-glucose, 0.1 mM of the poorly permeating sulphydryl blocker, chloromercuribenzene-p-sulphonic acid, also evoked a phosphate flush resembling the one induced by D-glucose. The basal radiophosphate release was partially inhibited by 1 mM 4-acetamido-4-'-isothiocyanostilbene-2,2'-disulphonic acid. However, the phosphate flush induced by 16.7 mM D-glucose was not noticeably inhibited by 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid. It is concluded that the phosphate flush emanates from beta-cells and that membrane sulphydryl groups may participate in its regulation. Although at least the basal phosphate release may in part represent transmembrane transport through 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid-sensitive anion channels, other mechanisms are also likely to participate in the glucose-induced phosphate flush.
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It was previously reported that struvite crystals could be formed in the aerobic composting reaction provided that Mg and P salts are added [Bioresource Technology 79 (2001) 129]. The formation of struvite crystals significantly reduced gaseous loss of ammonia and resulted in substantial increase in the ammonia content in the compost, attaining 1.5%. In this context, the present study was conducted to determine the optimal doses of Mg and P salts for struvite crystallization. It was found that cumulative ammonia production was about 33-36% of the initial total nitrogen in the aerobic composting reaction, irrespective of the amounts of Mg and P salts added. The theoretical doses for complete conversion of ammonia into struvite crystals seemed to be about 33-36% of the initial nitrogen. The addition of Mg and P salts at this level, however, caused adverse effects on the degradation of organic materials. Therefore, it was concluded that the optimal doses of Mg and P salts should be about 20% of the initial nitrogen in the compost mixture not to cause any harmful effects on the composting reaction.
We present a specific method for the determination of disodium clodronate in human plasma and urine using a gas-chromatographic system with nitrogen phosphorus detector (NPD). The compound was extracted from plasma and urine samples by an anion-exchange resin and derivatizated with bistrimethylsilyltrifluoroacetamide (BSTFA). Sodium bromobisphosphonate was used as internal standard. The calibration curves were linear in both plasma and urine, with a regression coefficient r > 0.9975 in plasma and r > 0.9977 in urine. The limit of quantitation was 0.3 microg/ml in plasma and 0.5 microg/ml in urine. The method was validated by intra-day assays at three concentration levels. During the study we carried out inter-day assays to confirm the feasibility of the method. The precision in plasma at 0.5, 15, and 45 microg/ml was 12.4, 0.2, and 6.5% (n = 40), respectively; in urine at 0.8, 8, and 40 microg/ml it was 8.6, 6.4, and 9.3% (n = 40), respectively. The method was accurate and reproducible, and was successfully applied to determine the pharmacokinetic parameters of clodronate in healthy volunteers after intravenous infusion and intramuscular injection of 200 mg of the compound. The Cmax after intravenous infusion and intramuscular injection was 16.1 and 12.8 microg/ml, respectively. AUC(0-48 h) after infusion administration and intramuscular injection was 44.2 +/- 18.0 and 47.5 +/- 12.4 h microg/ml, respectively. The elimination half-life in both administrations was 6.31 +/- 2.7 h.
The water quality of the Pozuelos-Murillo lagoon system in southern Mexico was evaluated during three periods between March and October 2002, with particular emphasis on the detection of organochlorine and organophosphorous pesticide residues in water and sediments. Physicochemical and microbiological parameters of water were also measured and integrated in a water quality index (WQI). Multivariate analysis was used to regionalise the lagoon system as a function of the behaviour of all measured parameters. Solid phase micro-extraction followed by gas chromatography (SPME-GC) was used for pesticide analysis. The concentration of phosphorous was found to be higher than that of nitrogenous compounds. This, besides a deficiency in dissolved oxygen and a high organic matter concentration (as COD), reflects eutrophication processes in some areas of the system. Measured levels of faecal coliforms and oils and greases were above the limits established by Mexican law and comparable to the concentrations reported for other highly polluted systems in Mexico. Residues of DDD (2.0 microg L(-1)) in water and DDE (247 ng g(-1)) and endosulfan I (814 ng g(-1)) sediments were detected by SPME-GC. The spatial distribution of these contaminants implies major potential risks because the most polluted sites were found to be those with the highest fishing activity. Although in general the WQI is on acceptable levels (65-80%), some contamination problems are evident.
With through space and through bond experiments in two-dimensional NMR we analyze the transformation from the thorium phosphate-hydrogen phosphate hydrate (TPHPH) to the beta form of the thorium phosphate diphosphate (beta-TPD) in relation with the phosphorus networks. These techniques are complementary: the through space coupling gives an insight on the dipolar phosphorus networks while the through bond coupling is particularly efficient in the detection of the P2O7 groups. With these experiments we show that in a first step, by heating the precursor TPHPH above 250 degrees C, it transforms into an alpha form of TPD. This transformation is due to the complete condensation of hydrogen phosphate groups HPO4 into P2O7 entities. By heating alpha-TPD above 950 degrees C it transforms into its well-known beta form. The alpha form is characterized by a hygroscopic behavior: some water molecules are present near the P2O7 groups that makes non-equivalent their phosphorus nuclei. PO4 dipolar networks are always present in the alpha form. The main effect of these PO4 and P2O7 units is to give the system a channel structure and the water enters in them.
The crystal structure of chorismate synthase (CS) from Streptococcus pneumoniae has been solved to 2.0 A resolution in the presence of flavin mononucleotide (FMN) and the substrate 5-enolpyruvyl-3-shikimate phosphate (EPSP). CS catalyses the final step of the shikimate pathway and is a potential therapeutic target for the rational design of novel antibacterials, antifungals, antiprotozoals, and herbicides. CS is a tetramer with the monomer possessing a novel beta-alpha-beta fold. The interactions between the enzyme, cofactor, and substrate reveal the structural reasons underlying the unique catalytic mechanism and identify the amino acids involved. This structure provides the essential initial information necessary for the generation of novel anti-infective compounds by a structure-guided medicinal chemistry approach.