Fates of N′-Methylnicotinamide and Nicotinamide N-Oxide in Mice

This experiment was performed to investigate the possibility that N′ -methylnicotinamide (N′-methyl-3-pyridinecarboxamide) and nicotinamide N-oxide have niacin activity or not in animals. When 20 mg N′-methylnicotinamide per mouse was administered, urinary excretion of nicotinamide, N¹-methylnicotinamide (MNA), N¹-methyl-2-pyridone-5-carboxamide (2-Py), and N¹-methyl-4-pyridone-3-carboxamide (4-Py) increased 24-, 3-, 3-, and 3-fold, respectively, compared with the control values. The increased ratios of MNA, 2-Py, and 4-Py were almost the same as those when 20 mg nicotinamide was administered. Therefore, the relative activity of N′-methylnicotinamide to nicotinamide as niacin was considered to be about 1. When 20 mg nicotinamide N-oxide per mouse was administered, urinary excretion of nicotinamide, MNA, 2-Py, and 4-Py increased 6.4-, 1.8-, 1.6-, and 1.7-fold, respectively, compared with the control values. The increased ratios of MNA, 2-Py, and 4-Py were about 1/2 of those when 20 mg nicotinamide was administered, so the relative activity of nicotinamide N-oxide to nicotinamide as niacin is considered to be about 1/2. In conclusion, it was found the possibility that the reactions N′-methylnicotinamide → nicotinamide and nicotinamide N-oxide → nicotinamide occur, at least in mice, and that therefore N′-methylnicotinamide and nicotinamide N-oxide have niacin activity.     

Acid Protease Produced by Trametes sanguinea,a Wood-destroying Fungus

A wood-destroying fungus, Trametes sanguinea, produced a potent acid protease in a submerged culture. Maximum proteolytic activity of the culture was attained after 140-hours cultivation in a medium containing dextrin and corn steep liquor. The acid protease was obtained in crystalline form from the mycelium-free culture filtrate by the following successive treatments: acetone precipitation, ionexchange column chromatography, ammonium sulfate fractionation, dialysis, and crystallization by acetone. Throughout the over-all process, the acid protease was purified approximately 30-fold with about 8% recovery of the original activity.     

Simple Measurement of Blood NADP and Blood Levels of NAD and NADP in Humans

The tryptophan synthase genes, trpA and trpB, of Bacillus stearothermophilus IFO13737 were cloned by transformation of tryptophan auxotrophic mutations of the trp genes into Escherichia coli. The genes are located in the order of trpB and trp A, according to their coding orientation, in a 2.5 kb EcoRy-Hindlll DNA fragment. The complete nucleotide sequence of this DNA was determined. The trp A and trpB genes consist of 810bp (269 amino acid residues) and 1215bp (404 amino acid residues), respectively. The 5′-proximal portion of the trpB gene was found to overlap 20 nucleotides of the upstream coding region of the trpA gene. The homology of the amino acid sequences of the trp gene products of trp A and trpB of B. stearothermophilus is 35 and 50 %, respectively, to those of E. coli, and 55 and 70 %, respectively, to those of B. subtilis.     

Change of Freezing Resistance and Retention of Metabolic and Differentiation Potentials in Cultured Green Lavandula vera Cells Which Survived Repeated Freeze-thaw Procedures

Cryostorage is one suitable method to preserve various desired types of cells. However, all cells do not survive after storage in liquid nitrogen. This suggests the possibility that the properties of the cells which survive after the storage differ from those of the unfrozen original cells.

Therefore, we did the same freeze-thaw procedure of cultured green Lavandula vera cells over again and compared the metabolic and the differentiation potentials of the cells which survived after the repeated freeze-thaw procedures with those of the unfrozen original cells. The results we found were that the frequency of colony formation of the cells which survived after the procedures was high, but that the biosynthetic capability for biotin and the differentiation potentials such as chloroplast formation and plantlet formation of the cells were equal to those of the unfrozen original cells. Cryostorage of cells in liquid nitrogen is discussed in terms of the preservation of various desired types of cells.     

Studies on Erythorbic Acid Production by Fermentation

Screening was carried out for erythorbic acid (EA)-producing strains from about 5,000 newly isolated fungi and bacteria. Penicillium notatum FY 115 was screened out as most powerful EA producer. Only Penicillium, but no other genera, was obtained as EA producers from our screening program. Monospore selections and mutagenic treatments succeeded to elevate the yield of EA over 40% to glucose supplied. Various cultural conditions were studied, and pH change during fermentation process was proved to be most important for favorable EA production. Over 80% yield could be obtained when washed mycelium was used in dilute glucose solution.

Abundant accumulation of EA by the strain FY 115, Penicillium sp., in fermentation broth was studied, and EA, both free and Na-salt, was obtained as crystal in the yield of about 45% to glucose supplied, in the media of 8% glucose by jar fermentor, in considering the inhibitory effect of some metal ion.

Extraction processes were improved to elevate the yield and was developed the continuous multi-bed extraction system of anion-exchange resin, which resulted in the yield of 90.9% of EA from fermentation broth in sum total.     

Formation of Nicotinamide Ribose Diphosphate Ribose,a New Metabolite of NAD,by Aspergillus niger

Aspergillus niger (AKU 3302) degraded NAD to form Compound X. This compound was identified as nicotinamide ribose diphosphate ribose (NAmRDPR) by hydrolysis with alkaline or phosphodiesterase followed by chemical analysis of the products. NAmRDPR showed absorption maxima at 265~266 nm in 0.1 n HCI and 325 nm in 1.0 n KCN. Optimal pH for NAmRDPR formation by the enzyme preparation from this organism was around 4.0. Formation of NAmRDPR proceeded stoichiometrically with degradation of NAD. Some of other strains of A. niger formed NAmRDPR, but production of this compound was not demonstrated in other mold genera.     

Diet of Japanese eels Anguilla japonica in the Kojima Bay-Asahi River system, Japan

The diet of Japanese eels, Anguilla japonica, was investigated using stomach content and stable isotope analyses. Stable isotope enrichment of carbon and nitrogen (Δδ¹³C and Δδ¹⁵N) was first estimated for A. japonica by comparing the isotopic signatures (δ¹³C and δ¹⁵N) of reared eels to that of their food. The estimated isotope enrichment was then applied to the diet estimation of A. japonica in the Kojima Bay-Asahi River system, Japan, combined with conventional stomach content analysis. Stable isotope enrichment varied among tissues, from 0.2‰ to 0.8‰ for carbon and from 1.3‰ to 2.1‰ for nitrogen. Nitrogen isotope enrichment of A. japonica muscle estimated in this study was 2.1‰, which was different from the previously reported mean δ¹⁵N enrichment of several animals of 3.4‰. These results indicate that isotope-based diet estimations for A. japonica need to use species- and tissue-specific values of isotope enrichment. In the diet analysis, stomach contents and stable isotopes revealed that (1) A. japonica appear to usually feed on a single type of prey species in each feeding session, (2) principal prey species were mud shrimp, Upogebia major, in brackish Kojima Bay and crayfish, Procambarus clarkia, in the Asahi River, (3) A. japonica in Kojima Bay primarily depend on the pelagic food web as a carbon source due to mud shrimp being filter feeders and eels in the Asahi River primarily depend on the littoral food web. Based on these results and the recently reported eel movements between Kojima Bay and the Asahi River, it appears that A. japonica can adapt to various feeding environments as opportunists, but also utilize the food resources by targeting a single type of prey species during a single feeding session.