Cobalamin deficiency results in severe metabolic disorder of serine and threonine in rats.

Dietary cobalamin (vitamin B12; Cbl) deficiency caused significant increases in plasma serine, threonine, glycine, alanine, tyrosine, lysine and histidine levels in rats. In particular, the serine and threonine levels were over five and eight times, respectively, higher in the Cbl-deficient rats than those in the sufficient controls. In addition, some amino acids, including serine and threonine, were excreted into urine at significantly higher levels in the deficient rats. When Cbl was supplemented into the deficient rats for 2 weeks, in coincidence with the disappearance of the urinary excretion of methylmalonic acid (an index of Cbl deficiency), the plasma serine and threonine levels were normalized. These results indicate that Cbl deficiency results in metabolic disorder of certain amino acids, including serine and threonine. The expression level of hepatic serine dehydratase (SDH), which catalyzes the conversion of serine and threonine to pyruvate and 2-oxobutyrate, respectively, was significantly lowered by Cbl deficiency, even though Cbl does not participate directly in the enzyme reaction. The SDH activity in the deficient rats was less than 20% of that in the sufficient controls, and was normalized 2 weeks after the Cbl supplementation. It is thus suggested that the decrease of the SDH expression relates closely with the abnormalities in the plasma and urinary levels of serine and threonine in the Cbl-deficient rats.

[Follow-up survey of dioxins in the blood of Yusho patients (in 1998-1999)].

Follow-up survey of the blood concentration of PCDDs, PCDFs and non-ortho-chlorine substituted PCBs in Yusho patients is very important for their health control. We determined the blood concentration of these dioxin isomers in 119 blood samples collected in 1998 and 1999 using by a high-resolution gas chromatograph/high-resolution mass spectrometry (HRGC/HRMS). Thirty years had passed since the Yusho occurrence, and the total blood concentrations of PCDFs in the blood of Yusho patients were still as high as ever, except the patients with the PCB pattern C. Among the different groups of PCB patterns, the concentration of 2,3,4,7,8-PeCDF, 1,2,3,4,7,8-HxCDF and 1,2,3,6,7,8-HxCDF showed significant differences, excluding the group BC because of the small number of cases. The findings indicate that these isomers are not decomposed in human body since they had highly contaminated the causal rice oil at the onset. In typical Yusho patients (Group A of PCB pattern), the mean TEQ concentrations of PCDDs, PCDFs and non-ortho-chlorine substituted PCBs in the blood collected in 1998 were 21, 206, and 14 pg-TEQ/g lipid, respectively, and in 1999, 30, 308 and 14 pg-TEQ/g lipid, respectively. The toxic contribution rate of PCDFs TEQ was still at high levels for total TEQ in both years. The patients of the group C, however, the mean blood levels of the dioxins TEQ in 1998 and 1999 did not differ from those of the normal controls determined in 1996 although some patients in this group showed a high toxic contribution rate of PCDFs TEQ for total TEQ. We will try the PCB isomer specific analysis of the Yusho patients blood, and will investigate the difference from normal controls blood.

Characterization of a vitamin B12 compound from unicellular coccolithophorid alga (Pleurochrysis carterae).

A unicellular coccolithophorid alga, Pleurochrysis carterae, contained 125.4 +/- 1.2 microg of vitamin B12 per 100 g dry cell weight of the lyophilized algal cells. A vitamin B12 compound was purified from the lyophilized algal cells and partially characterized. The silica gel 60 TLC and reversed-phase HPLC patterns of the purified pink-colored compound were identical to those of authentic vitamin B12, but not those of vitamin B12 analogues inactive for humans. When 22-week-old B12-deficient rats which excreted substantial amounts of methylmalonic acid (75.5 +/- 12.3 mg/day) in urine were fed the P. carterae (10 g per kg diet)-supplemented diet for 12 d, urinary methylmalonic acid excretion (as an index of vitamin B12 deficiency) of the rats became undetectable and hepatic vitamin B12 level of the rats was significantly increased.

Feeding dried purple laver (nori) to vitamin B12-deficient rats significantly improves vitamin B12 status.

To clarify the bioavailability of vitamin B12 in lyophylized purple laver (nori; Porphyra yezoensis), total vitamin B12 and vitamin B12 analogue contents in the laver were determined, and the effects of feeding the laver to vitamin B12-deficient rats were investigated. The amount of total vitamin B12 in the dried purple laver was estimated to be 54.5 and 58.6 (se 5.3 and 7.5 respectively) microg/100 g dry weight by Lactobacillus bioassay and chemiluminescent assay with hog intrinsic factor respectively. The purple laver contained five types of biologically active vitamin B12 compounds (cyano-, hydroxo-, sulfito-, adenosyl- and methylcobalamin), in which the vitamin B12 coezymes (adenosyl- and methylcobalamin) comprised about 60 % of the total vitamin B12. When 9-week-old vitamin B12-deficient rats, which excreted substantial amounts of methylmalonic acid (71.7(se 20.2) micromol/d) in urine, were fed the diet supplemented with dried purple laver (10 microg/kg diet) for 20 d, urinary methylmalonic acid excretion (as an index of vitamin B12 deficiency) became undetectable and hepatic vitamin B12 (especially adenosylcobalamin) levels were significantly increased. These results indicate that vitamin B12 in dried purple laver is bioavailable to rats.

DNA sensing on a DNA probe-modified electrode using ferrocenylnaphthalene diimide as the electrochemically active ligand.

Naphthalene diimide derivative 1 carrying ferrocenyl moieties at the termini of imide substituents binds intact calf thymus DNA 4 times more strongly than the denatured DNA, and its complex with the intact DNA dissociates 80 times more slowly than that with the denatured DNA. On the basis of these observations, ligand 1 was applied to a probe of electrochemical DNA sensing. A thiol-linked single-stranded DNA probe was immobilized through the S-Au bonding to 20-30 pmol/mm2 on a gold electrode. Following hybridization with the complementary DNA, the electrode was soaked in a solution containing 1 (intercalation step) and then washed with buffer for 5 s. The cyclic voltammogram and differential pulse voltammogram for this electrode gave an electrochemical signal due to the redox reaction of 1 that was bound to the double-stranded DNA on the electrode. Thus, dA20 and the yeast choline transport gene were quantitated at the subpicomole level. The sensitivity of DNA detection was improved to 10 zmol by reducing the amount of immobilized DNA probe and protecting the uncovered surface of the electrode with 2-mercaptoethanol.

[Therapeutic trial for promotion of fecal excretion of PCDFs and PCBs by the administration of cholestyramine in Yusho patients].

Any effective therapy for elimination of causal agents remaining in Yusho patients was not found until now. To know the profile of fecal excretion of polychlorinated dibenzofurans (PCDFs) and polychlorinated biphenyls (PCBs), the amounts of PCDFs and PCBs in the stool of six Yusho patients with the typical symptoms were determined. The stool samples of Yusho patients were collected in 1989. PCDFs, i.e., 2,3,7,8-tetrachlorodibenzofuran (TCDF), 2,3,4,7,8-pentachlorodibenzofuran (PnCDF), 1,2,3,4,7,8- and 1,2,3,6,7,8-hexachlorodibenzofurans (HxCDFs), 1,2,3,4,6,7,8-heptachlorodibenzofuran (HpCDF) and octachlorodibenzofuran (OCDF) were detected in all of the samples. PCDFs found in the stool samples were mostly PnCDF and HxCDFs. Of PCDFs detected, PnCDF and HxCDFs contributed to 42 +/- 4.7% and 43 +/- 5.5% as mean +/- SE, respectively. The fecal excretion of PnCDF and HxCDFs in Yusho patients was 720 +/- 490 pg/day and 790 +/- 620 pg/day as mean +/- SE, respectively. On the other hand, the fecal excretion of PnCDF and HxCDFs in normal controls was 32 +/- 13 pg/day and 47 +/- 5.2 pg/day as mean +/- SE, respectively. The fecal excretion of PnCDF and HxCDFs in Yusho patients was about 23 times and 17 times each higher than that in normal controls. The fecal excretion of PCBs in Yusho patients and normal controls was 400 +/- 430 ng/day and 150 +/- 39 ng/day, respectively, as mean +/- SE. In order to promote the excretion of these toxic chemicals in the stool of Yusho patients, the patients were continuously administered with cholestyramine, an anion exchange resin, at a dose of 4 g, 3 times a day, for 6 months.(ABSTRACT TRUNCATED AT 250 WORDS)

[Statistical analysis of subjects with type A, BC and C pattern of blood PCB on GC].

Various combinations of measured data on PCBs, PCQs, and PCDFs in adipose tissue or blood were tested on its discriminating power between normal persons and persons poisoned by yusho oil. The most discriminative single parameter was concentration of PCDF in adipose tissue. Furthermore, discriminative power increased by using additional parameters.

[Polychlorinated dibenzofurans (PCDFs) in the subcutaneous adipose tissue of yusho patients and normal controls].

Polychlorinated dibenzofurans (PCDFs) are well known to be the toxic chemicals in both animal experiments and human studies. It is, therefore, important to determine the level of PCDFs still retained in patients for understanding relationship between the concentration of PCDFs and present symptoms of the disease. In this study, the abdominal subcutaneous adipose tissue of 18 yusho patients and those of 11 normal controls who were all of volunteers were collected, and their levels were determined by high resolution gas chromatography/high resolution mass spectrometry. Results obtained were as follows: The principal compounds detected in the adipose tissue of yusho patients were 2,3,4,7,8-pentachlorodibenzofuran (PenCDF), 1,2,3,4,7,8- and 1,2,3,6,7,8-hexachlorodibenzofuran (HCDF). The concentration of the compounds in 7 patients, wearing typical symptoms, from 160 to 3,000 ppt for 2,3,4,7,8-PenCDF, from 51 to 1,000 ppt for 1,2,3,4,7,8-HCDF, and from 16 to 220 ppt for 1,2,3,6,7,8-HCDF. In normal controls, 2,3,4,7,8-PenCDF was detected only in five samples at the low level of from 16 to 38 ppt. On an average, PCDF levels in in the typical 7 yusho patients and 11 normal controls were 1,900 ppt and 16 ppt, respectively. On the basis of the results, the concentrations of PCDF congeners in the adipose tissues of the typical 7 patients was 100 times higher than that of the normal controls. Hence, we consider that the present levels of PCDFs in the patients probably play an important role for the symptoms of the yusho.