No abstract

Our results show that a net synthesis of thiamine triphosphate (TTP) can be demonstrated in vitro using rat brain extracts. The total homogenate was preincubated with thiamine or its diphosphate derivative (TDP), centrifuged, and washed twice. With TDP ( 1 mM) as substrate, a 10-fold increase in TTP content was observed in this fraction (nuclear fraction, membrane vesicles). A smaller, but significant, increase was observed in the P2 fraction (mitochondrial/synaptosomal fraction). In view of the low TTP content of our fractions, it was carefully assessed that authentic TTP was being formed. Incorporation of radioactivity from [P-32P]TDP and [T-~~PIATP in TTP suggests that these two compounds are its precursors. Furthermore, TTP synthesis was inhibited by ADP and relatively low concentrations of Zn2+. These results suggest that TTP synthesis is catalyzed by an ATP:TDP transphosphorylase rather than by the cytoplasmic adenylate kinase that may be present in the vesicles. After osmotic lysis of the vesicles at alkaline pH, TTP was recovered in protein-bound form. Concomitantly, a soluble thiamine triphosphatase, with aikaline pH optimum, was also released from the vesicles. No net synthesis could be obtained in the cytosolic fraction or in detergent-solubilized systems. Like TTP synthesis, chloride permeability of the vesicles was increased when the homogenate had been incubated with thiamine and particularly with TDP. Our results suggest a regulatory role of TTP on chloride permeability, but the target remains to be characterized. Key Words: Thiamine triphosphate-Thiamine diphosphateChloride permeability-Rat brain. Bettendorff L. et at. Metabolism of thiamine triphosphate in rat brain: Correlation with chloride permeability. J. Neurochem. 60, 423-434 (1993).

Peanut butter extracts and samples spiked with 5-40 μg anatoxin B1/kg were analyzed, together with naturally contaminated peanut products, by 3 extraction procedures: the official Dutch method (KB), he Liem et al. method (methanol), and the IUPAC method. The last procedure was selected as a reference method, since it has international application. KB extracts were separated on silica gel G plates with a mixture of chloroform-acetone (90 + 10), whereas IUPAC extracts were separated similarly on MN-G-HR plates. Methanol extracts were resolved on silica gel II plates, using chloroform-trichloroethylene-n-amyl alcohol-formic acid (80 + 15 + 4 + 1) as the developing solvent. After development, plates were scanned with a reflectance flying-spot densitometer. With such techniques, average recoveries for spiked peanut butter extracts ranged from 99 to 105%, with variation values of 11-12%. Recovery values of 69% (KB method) and 84% (methanol and IUPAC methods) were obtained for spiked peanut butter samples. Coefficients of variation ranged from 13 to 15% for fluorodensitometric measurements. Innaturally contaminated peanuts and peanut products , precision values were 13.6% for fluorodensitometric measurements compared to 36% for visual estimations . Both the methanol and IUPAC methods yield extracts suitable for densitometric analysis after spotting on TLC plates; the analytical results obtained are comparable. Extracts from the KB method contained more interfering fluorescent material than the other 2 methods