Residual amounts of 2, 2-bis-(p-chlorophenyl), 1, 1-dichloroethylene (p, p' DDE) and the concentrations of total lipids and triglyceride were studied with respect to the internal organs of rats which had been dosed with p, p' DDE and fed on the diet free from vitamin A or containing excess of vitamin A. Either the absence or excess of vitamin A was found to increase triglyceride content and p, p' DDE storage level in the liver. However, the ratio of p, p' DDE storage level in the liver to content of liver triglyceride was almost constant independently of the vitamin A treatment. It is suggested that the dose of vitamin A has some influences upon the metabolic processes of liver triglyceride in rats and changes the storage level of p, p' DDE in the organ.
The basic properties of microsomal thiamine diphosphatase (TDPase) in rat brain were examined. It was mainly localized in the microsomal fraction. Microsomal TDPase showed an absolute requirement for divalent cation which was best satisfied by Ca++. Theophylline (1mM), NADP (0.1mM) and NADPH (0.1mM) significantly inhibited its activity. However, caffeine, theobromine, NAD and various putative neurotransmitters did not affect the enzyme activity. UDP was inhibitory and IDP was a competitive inhibitor, but ATP had no effect on the enzyme activity.
The purine precursor in the riboflavin biosynthetic pathway in Eremothecium ashbyii was examined using a guanine analogue, 8-azaguanine, with non-growing cell systems. 1. Riboflavin formation in the culture filtrate was determined at 0, 5, 10 and 20 hr after start of the incubation of the non-growing cells in the presence of xanthine or 8-azaguanine (1mM, respectively). At 20 hr of incubation, the addition of xanthine stimulated riboflavin formation by 36% and the addition of 8-azaguanine inhibited the formation by 57%. 2. Acid soluble nucleotide pools in the cells were followed at 0, 5, 10 and 20 hr of the incubation period in the presence of xanthine or 8-azaguanine by means of anion exchange column chromatography. The result showed that the GTP pool changed markedly despite the fact that the adenosine nucleotide pool was almost constant irrespective of the presence or absence of these purines till 10 hr of incubation. Put, the decrease of the former was overcome in part by the addition of flavinogenic xanthine. Furthermore, the total amounts of GTP and guanosine accumulated in cells in the presence of 8-azaguanine reached the maximum already at 5 hr, attaining a level twice as much as the GTP contents of the control. 3. The role of guanosine nucleotide pool in riboflavin formation was further examined using 8-azaguanine. In this experiment the drug was added to the suspension of non-growing cells at 3 hr or 6 hr after the incubation was started and the reaction was continued till the 12th hr. A more clear-cut correlationship between riboflavin formation and guanosine nucleotide pool was observed by this experiment. The guanosine nucleotide pool (consisting of GMP, GDP and GTP) increased simultaneously with the inhibition of riboflavin formation. Of the guanosine nucleotides pools, the GMP pool increased 2.7 times above normal upon the addition of 8-azaguanine during the incubation for 6 hr and 5.3 fold for 9 hr. While, the GTP pool increased 1.9 fold above normal for 6 hrs' incubation in the supplementation of this drug but decreased to one-half of the normal at the incubation period of 9 hr. In these cases, the decreased amounts of GTP were equal to the increased amounts of GMP during the incubation periods of 6 hr and 9 hr in the presence of added 8-azaguanine. 4. The above results suggest strongly that GTP is an immediate precursor of riboflavin in the form of nucleotide.
Studies to elucidate the chemical structures of new flavins formed by Schizophyllum commune, namely schizoflavin (SF), were carried out. It was found by an enzymatic study that riboflavin (FR) was apparently converted via SF2 to SF1, and that SF2 was a direct precursor of SF1. SF1 was crystallized from water and repeated recrystallization produced yellow needles. The carboxylic group in SF1 and the aldehyde group in SF2 were detected by spot tests. Elemental analyses and the spectroscopic analyses of the crystalline SF1 supported the existence of carboxylic group in the terminal of its ribityl moiety. From these experimental results, schizoflavins were revealed to be oxidation products of FR that C-5' site of its ribityl moiety is oxidized. Consequently, SF1 and SF2 were identified as 7, 8-dimethyl-10-(2, 3, 4-trihydroxy-4-carboxybutyl) isoalloxazine and 7, 8-dimethyl-10-(2, 3, 4-trihydroxy-4-formylbutyl) isoal-loxazine, respectively.
It has been shown that the determination of small amounts of tocopherol in tissue lipids by methods that use, in sequence, saponification, extraction, TLC and colorimetric assay, is greatly influenced by the concentration of soap in the saponification mixture. Soaps appear to interfere with the recovery of tocopherols during the subsequent extraction step. Based on this knowledge a method is described that was used to measure as little as 15 to 20ng of tocopherol in 3 to 4mg of lipid obtained from rat retinas. Presumably the method would be applicable to other tissues as well. The results show that the rat retina is readily depleted of tocopherol if none is included in the diet.
A wheat grain was divided into three portions, endosperm, germ and bran, and the protein profile of each was examined comparatively after sequential extraction with isopropyl alcohol, sodium chloride, lactic acid and KOH solutions. Recovery of the protein in the extracts was 93-96%. The relative protein concentrations of the four soluble fractions of endosperm were very similar to those of bran, but germ showed a distinct distribution of the soluble proteins. Soluble protein fractions of endosperm, germ and bran, particularly their NaCl soluble fractions, exhibited distinct individuality upon examination by polyacrylamide gel electrophoresis and gel filtration. Indeed, it was suggested that the gel electrophoretic profile could serve as a criterion for the quality evaluation of wheat flour. On the other hand, the KOH soluble proteins of endosperm, germ and bran, which gave indiscrete electrophoretic patterns, existed predominantly in highly aggregated forms which disintegrated upon exposure to 1% SDS or by reduction with 2-mercaptoethanol.
Autoxidized LA is classified into four groups, LA, LAHPO, SP and FP. Lysozyme is inactivated by these products in the increasing order as follows: FP<LA<LAHPO<SP. The effects of these products on the amino acid composition of lysozyme is examined. All kinds of amino acid residues were not damaged until lysozyme was incubated with LA and LAHPO at 45°C for 100 days. The susceptible amino acid residues attacked by the autoxidized products are tryptophan, lysine and histidine. The specific loss of methionine by SP occurs during acidhydrolysis. The effect of SP was the strongest among the autoxidized products. FP was almost noneffective. The destructive actions of BP, MA and PA were compared with those of autoxidized products. Effects of these compounds did not resemble those of autoxidized products. It was concluded that tryptophan, lysine and histidine residues were specifically attacked by SP.