As the transketolase requires thiamine diphosphate as cofactor, the transketolase activity of erythrocyte hemolysate can be used as an index for thiamine deficiency. In the present report, the effects of thiamine and cyclocarbothiamine (CCT), each 100μg/rat/day orally, on the transketolase activity and urinary excretion of thiamine in thiamine deficient rats were discussed. Urinary excretion of thiamine was greatly reduced after 3 days feeding with thiamine deficient diet, then increased as twice as control if thiamine or CCT was given for 3 days. After that, daily thiamine excretion was about the same amount until 20th day in thiamine administered group, but gradually increased in CCT group. Transketolase activity was significantly depressed after 10 days in thiamine deficient rats, and much more depression was noticed after 20 days. Thiamine and CCT were effective to prevent the depression of the enzyme activity, but there was no significant difference between these two groups.
Thiamine deficient status was estimated by determining the transketolase activity in redcell hemolysate in normal children and in several sick status in childhood. Transketolase activity was expressed by TDP-effect which is the per cent increase of hexose after the addition of thiamine diphosphate into the hemolysate. The mean value of TDP-effect was 3.8 ± 1.1% in normal infants of artificial feeding, 8.8 ± 0.7% in early childhood, and 12.6 ± 0.7% in prepubescence, and their daily thiamine intakes were 0.78. 0.42 and 0.99 mg, respectively. There was slight thiamine deficiency in prepubescent stage. In orthostatic dysregulation and acetonemic vomiting, the activity was depressed. In bronchial asthma, recurrent abdominal pain, noctural enuresis, diarrhae and convulsion, there were no significant differences from the normal group. During hypothermic operation of children with heart diseases, the enzyme activity was not changed significantly in contrast to the preoperative stage.
Pyridoxine in a neutral solution was irradiated with an ultraviolet lamp (characteristic wavelength, 2537Å under aerobic and anaerobic conditions, respectively. Photolysis products, detected on paper chromatograms and paper ionophoregrams, were isolated by Dowex-1 (formate form) column chromatography and identified by their spectroscopic behaviors and other characteristics. The sequence of the photolytic route was determined by successive photolysis of the intermediates. Under aerobic conditions, pyridoxine was oxidized through two different pathways : The one is PIN→PAL→4-PiA (4-PiA lactone)→4,5-di-PiA, and the other is PIN→iso-PAL→5-PiA (4-PiA-lactone)→4,5-di-PiA. 4,5-di-PiA was thought to be further decomposed. On the other hand, pyridoxine was converted to its dimer when exposed to light under anaerobic conditions.
A simple method for microbiological assay of pyridoxal was investigated by using Sacch. carlsbergensis. When the growth-stimulating activity of pyridoxal was compared with those of pyridoxal derivatives, the activity of pyridoxal-oxime was found to be negligibly low on assay ; that was only 1/100 the potency of pyridoxal. From the results, it was suggested that the pyridoxal content in vitamin B_6 solution can be calculated by the subtraction of activity of hydroxylamine-treated vitamin B_6 solution from the total activity. The formation of oxime from pyridoxal was carried out by the addition of 1 mg of hydroxylamine to the solution containing each 10μg of pyridoxal, pyridoxol and pyridoxamine, and by boiling for 10 min. at pH 6.5. When the pyridoxal contents in dried yeasts, rat livers and kidneys were determined by this method, the recovery values were 103-114%.
Pyridoxine 5-octanoate was synthesized and its chemical and physical properties were studied. Molar extinction coefficient of its ethanol solution at 286.5mμ was 6.5 × 10^3 cm^<-1>mole^<-1>. It was soluble in various organic solvents such as benzene, chloroform, and ethanol, but its solubility in water was 0.273μmole/ml. Its solvent affinity was intermediate between pyridoxine and pyridoxine 3,4-dioctanoate. Pyridoxine 5-octanoate and pyridoxine 5-benzoate showed a weak vitamin B_6 activity for Saccharomyces carlsbergensis. Pyridoxine 5-octanoate was hydrolyzed completely in 0.055N H_2SO_4 at 126.5℃ for one hour.
Absorption of fatty acid esters of pyridoxine (I) through everted sac of the rat intestine was studied. Pyridoxine 5-monoctanoate (II) and pyridoxine 3,4-dioctanoate (III) were hydrolyzed completely in the mucosal side, but pyridoxine 3,4-dilaurate (IV) was hydrolyzed only 5.5 % and pyridoxine 3,4-dipalmitate (V) was not. In the serosal side, the esters penetrated through the intestine were found to be (I) alone and no esters. The concentration of (I) in solution of the serosal side was the following order : (I)=(II)=(III)>(IV) and none for (V). There was a good correlationship between penetration through the everted sac of rat intestine, hydrolysis by homogenate of the rat intestine and solubility in a Krebs-Ringer's solution containing 10 % propylene glycol at pH 7.4. The penetration of esters of (I) was concluded that the esters were hydrolyzed in the intestinal fluid and (or) tissue and the resulted (I) penetrated through the intestine.
Gastro-intestinal absorption of fatty acid esters of pyridoxine was studied in human by the oral administration of 243.3 μmoles. It was found that the urinary excretion of pyridoxine 5-octanoate and 3,4-dioctanoate in 48 houre was as large as pyridoxine, but those of pyridoxine 3,4-dilaurate and -dipalmitate were small. By the study of thin-layer chromatography, the main metabolite of the esters was 4-pyridoxic acid, i.e., the same metabolite of pyridoxine. From the present study, it was found that the esters were hydrolyzed to pyridoxine in the human body. There was a good correlationship between the intestinal absorption of the esters in human and their penetration through everted sac of the rat intestine. The differences of absorption and penetration between the esters were closely correlated with the differences of degree of hydrolysis by homogenates of the mouse intestine.
The 3,3', 5'-trihydroxy-5', 6'-dihydro-β-carotene-5,6-monoepoxide structure proposed by A.L. Curl and N.I. Krinsky for neoxanthin was not coincident with natural neoxanthin obtained from spinach. The specimen having the above structure was synthesized from zeaxanthin isolated from Chinese lantern plant. As model experiments, the LiAlH_4 reduction of the 5,6-mono-epoxide, 5,8-epoxide, or 5,6,5', 6'-diepoxide of β-carotene, and the epoxidation or dehydration of 5- and 6-hydroxy-5,6-dihydro-β-carotene were fully studied. The spectral characterization of the epoxides of xanthophylls was also discussed.