Folyl polyglutamate synthetase has been partially purified from mouse liver, and the general features of this enzyme have been characterized. The purification procedure utilized fractionation with ammonium sulfate, gel filtration, and affinity chromatography on ATP-agarose and resulted in a 350-fold increase in specific activity with 8-20% recovery of enzyme activity. Enzyme could be stabilized by glycerol or by ATP, but stability was not appreciably enhanced by folate. The enzymatic reaction was completely dependent on folate, ATP, and Mg2+ while partial reaction rates were observed in the absence of KCl or beta-mercaptoethanol. Highest reaction rates were observed at pH 8.2-9.5 at 37 degrees C. Chromatography of purified enzyme on calibrated gel filtration columns suggested a molecular weight of 65 000. Mouse liver folyl polyglutamate synthetase coupled [3H]glutamic acid to all of the naturally occurring folates studied. Analysis of the reaction products by high-performance liquid chromatography demonstrated that several folyl oligoglutamates were formed at low substrate concentrations but that only folyl diglutamate was formed at substrate concentrations approaching saturation. Dihydrofolate, tetrahydrofolate, 5,10-methylenetetrahydrofolate, 10-formyltetrahydrofolate, and 5-formyltetrahydrofolate were the best substrates. Folic acid and 5-methyltetrahydrofolate were also substrates for this reaction, but much higher concentrations of these compounds were required to saturate the enzyme. These data suggest that all of the tetrahydrofolyl compounds (except 5-methyltetrahydrofolate) are the monoglutamyl substrates for polyglutamation in vivo and that 5-methyltetrahydrofolate is not likely to be a direct precursor for folate polyglutamates in mouse liver.

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The hydrogenation of 1,3-hexadiene over a Pd/silica catalyst has been investigated using in situ FTIR spectroscopy. This study shows that the presence of an organic species on the surface of the catalyst can have a marked effect on the rate of hydrogenation of 1,3-hexadiene. Toluene-d 8 was coadsorbed on the surface of the catalyst and, depending on the order of introduction of the reactants to the system, has been shown to act as either a promoter or a poison for the hydrogenation of the diene. Introduction of the toluene-d 8 prior to the diene has a promoting effect with the toluene-d 8 acting as a H-transfer agent. When the toluene-d 8 is introduced after the diene it acts as a poison, blocking active sites and covering the surface. The ability of toluene-d 8 to act as a promoter or poison appears to be dependent on the availability of active sites at the time of its introduction to the reaction system. To act as a promoter, or H-transfer agent, the toluene-d 8 must first be able to form σ bonds to the surface by dissociation of the D atoms of the methyl group. When the site availability is poor, the toluene-d 8 retains its molecular identity upon adsorption, thus blocking active sites and reducing the rate of hydrogenation.

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