Data Science for Transportation

HOSHI A., YOSHIDA M., IIGO M., TOKUZEN R., FUKUKAWA K., UEDA T.

Neplanocin A is a cyclopentenyl analog of adenosine which has been isolated from the culture filtrate of Ampullariella reqularis. Antitumor activity of twenty three derivatives of neplanocin A was examined against L1210, sarcoma 180 and L5178Y. Neplanocin A showed a marked inhibition of growth of L1210 in vivo. Other derivatives which had a 2' or 3'-substituted cyclopentene group showed weak cytotoxicity against L5178Y cells in vitro. Neplanocin A inhibited the biosynthesis of ribonucleic acid (RNA) and protein, while 6-chloroneplanocin A, a new active derivative, showed a specific inhibition of only RNA synthesis. The two hydroxy groups in the cyclopentene moiety with a ribose type structure are important for marked antitumor activity.

IRIE T., OTAGIRI M., SUNADA M., UEKAMA K., OHTANI Y., YAMADA Y., SUGIYAMA Y.

Cyclodextrins (CyDs) at higher concentrations were found to cause hemolysis of human erythrocytes in the order of beta- greater than alpha- greater than gamma-CyD in isotonic solution. Biphasic effects of CyDs were observed for the osmotic and heat-induced hemolysis; i.e. the protection at relatively low CyD concentrations and stimulation at higher CyD concentrations. From the scanning electron microscopic observations, CyDs induced shape changes of membrane internalization type on erythrocytes. CyDs caused the release of cholesterol from erythrocyte membrane in the order of beta- greater than gamma- greater than alpha-CyD. These results clearly indicate that CyD-induced hemolysis is probably a secondary event resulting from the membrane disruption which elicited the removal of membrane components from erythrocytes.

SONODA Y., SATO Y.

7-Oxo-24,25-dihydrolanosterol (7-oxo-DHL), an inhibitor of cholesterol biosynthesis from lanosterol in vitro, lowered the serum total-cholesterol and triglyceride at a level of 0.1% in a diet on male Wistar rats. This effect was slightly lower than that of clofibrate (CF) which was examined as a reference. Further, the reduction in serum total-cholesterol levels were associated with a slight reduction in the levels of high density lipoprotein (HDL) cholesterol, resulting in a decrease in the atherogenic index. The rate of decrease of liver total-cholesterol by 7-oxo-DHL was higher than that of CF soon after taking off the diet. Although CF produced a marked increase of liver size, no significant increase was observed in the rats fed with 7-oxo-DHL. The effects of 7-oxo-DHL in the rats fed with 3% cholesterol in a diet were examined and compared with that of control. These results strongly suggest that 7-oxo-DHL is considerably hypolipidemic in rats.

NAGATA K., MATSUNAGA T., BUPPODOM P., ISHIMATSU M., YAMATO H., YOSHIHARA S., YOSHIMURA H.

Pretreatment of male Wistar rats with 3,4,5,3',4'-pentachlorobiphenyl (PenCB), a potent 3-methylcholanthrene-type inducer, increased selectively 7 alpha-but strongly repressed 2 alpha-, 6 beta- and 16 alpha-hydroxylations of testosterone in the liver microsomes. To understand this unique phenomenon, the testosterone metabolism by three isozymes (P-452, P-448 L and P-448 H) of cytochrome P-450 purified from Wistar rats treated with PenCB was studied in a reconstituted system. For comparison 4 other isozymes (P-451 I, P-451 II, P-450 II and P-450 III) of cytochrome P-450 from untreated and phenobarbital-treated rats were also studied. In addition, the contribution of cytochrome P-450's to testosterone hydroxylation was examined by an immune complex inhibition of the activity and by a determination of the individual cytochrome P-450 in microsomes using antibodies. In the reconstituted system, 7 alpha-hydroxylation of testosterone was catalyzed almost exclusively by P-452, with the exception of P-451 I. On the other hand, the 6 beta-hydroxylation was catalyzed by most of the cytochrome P-450's tested at considerably lower rates. Among the seven forms, P-451 II was the most effective catalyst for 2 alpha- and 16 alpha-hydroxylations, with equally high turnover numbers, while other forms showed only low or no activity for either or both hydroxylations. The microsomal activity of 7 alpha-hydroxylation in PenCB-treated rats was inhibited almost completely by anti-P-452. A partial inhibition of the 16 alpha-hydroxylation was achieved by anti-P-451 II and anti-P-452 while the 6 beta-hydroxylation was insensitive to anti-P-451 II but slightly sensitive to anti-P-452. The activity of 2 alpha-hydroxylation was not detected in the liver microsomes from PenCB-treated rats. Immunochemical quantitation showed that in the microsomes from PenCB-treated rats, P-451 II, P-452, P-448 L and P-448 H accounted for 4.1, 3.6, 31.6 and 62.8%, respectively, of the total cytochrome P-450 (2.69 nmol/mg protein). On the other hand, the microsomal cytochrome P-450 in untreated rat livers (0.83 nmol/mg protein) consisted of 62.7% as P-451 II, less than 1% as P-452 and the remainder as P-451 I and other unknown forms.(ABSTRACT TRUNCATED AT 400 WORDS)

NISHIGAKI T., NAKAMURA K., KINOSHITA T., KUWANO H., TANAKA M.

Metabolites of -[(4-amino-2-methyl-5-pyrimidinyl) methyl]-1-(2-chloroethyl)-1-nitrosourea hydrochloride (ACNU) in rat urine were investigated. After intravenous administration of 14C-ACNU into rats, four major radioactive metabolites and two minor ones were detected in the urine by two-dimensional thin-layer chromatograhic analysis. The main metablite was identified to be an imidazolidinone compound, 1-[(4-amino-2-methyl-5-pyrimidinyl) methyl]-5-hydroxy-2-imidazolidinone (M-D). One of the other major metabolites was identified to be a nitrosated compound of the main metabolite i.e., 1-[(4-amino-2-methyl-5-pyrimidinyl) methyl]-5-hydroxy-3-nitroso-2-imidazolidinone (M-C). These were new types of metabolites which have not been reported in the metabolic study of other chloroethylnitrosourea derivatieves. Compared with authentic compounds, two metabolites were identified to be a denitrosated derivative of ACNU i.e., 1-[(4-amino-2-methyl-5-pyrimidinyl) methyl]-3-(2-chloroethyl) urea (M-B), and a cyclized pyrimidopyrimidine compound which lacks the ethylene moiety of ACNU, i.e., 3, 4-dihydro-7-methylpyrimido [4, 5-d] pyrimidin-2-(1 H)-one (M-A). The two minor metabolites were supposed to be compounds derived from M-A. Discussions were made on mechanism of formation of these metabolites in vivo.

NISHIGAKI T., NAKAMURA K., TANAKA M.

In vitro decomposition of ACNU, 3-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-1-(2-chloroethyl)-1-nitros ourea hydrochloride, in various conditions was studied with the use of the 14C-labeled compound. Metabolite A, 3,4-dihydro-7-methylpyrimido[4,5-d]pyrimidin-2(1H)-one (an intramolecular cyclized product), was formed spontaneously in the phosphate buffer (pH 7.4) with simultaneous liberation of the alkylating moiety. With rat liver enzyme preparations, formation of three metabolites was observed. Those were metabolite B, 1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)urea (a denitrosated product), metabolite C, 1-[(4-amino-2-methyl-5-pyrimidinyl) methyl]-5-hydroxy-3-nitroso-2-imidazolidinone (a product via oxidative dechlorination), and metabolite D, 1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-5-hydroxy-2-imidazolidinone (a denitrosated product of metabolite C). Formation of metabolite B was catalyzed with both cytosolic and microsomal enzymes, not inhibited with SKF-525A, and partly dependent on nicotinamide adenine dinucleotide phosphate (NADPH). These results suggest that at least two enzymatic steps would be involved in the formation of this product. Metabolites C and D were produced by the microsomal preparation, being dependent on O2 and NADPH, inhibited by CO and SKF-525A, and enhanced by phenobarbital pretreatment. When metabolite C was incubated with cytosolic and microsomal preparations, more efficient formation of metabolite D with the former than the latter was observed. From these results, it was assumed that oxidative dechlorination of ACNU to metabolite C would be catalyzed with the microsomal mixed function oxidase, and metabolite D would be produced via denitrosation process of metabolite C.

YAMAOKA K., NAKAGAWA T.

A nonlinear least squares program based on first-order simultaneous differential equations, MULTI (RUNGE), for personal computers was developed. Four algorithms, i.e. the classical Gauss-Newton method, the damping Gauss-Newton method, the modified Marquardt method and the simplex method can be used for the nonlinear curve fitting in MULTI (RUNGE). The differential equations and the initial conditions which are voluntarily defined by the user are numerically solved by the Runge-Kutta-Gill method and the numerically integrated solutions are fitted to the observed data points by the iterative least squares algorithms. MULTI (RUNGE) can be executed on many personal computers without any modification, because this program is written in the Microsoft minimum BASIC commands alone. The dimensions of differential equations, the number of parameters to evaluate and the number of experimental data points are restricted only by the available memory in computers and by the computing time.

NAGANUMA A., URANO T., IMURA N.

A rapid and simple method for preparation of 203Hg-labeled methylmercuric chloride (CH3(203)HgCl) from 203HgCl2 and methylcobalamin is described. More than 99% of 203Hg was methylated with methylcobalaminin 0.01 N HCl during 1 h. CH3(203)HgCl formed during the reaction was rapidly and completely separated from cobalamins and unreacted 203HgCl2 by CM-Sephadex C-25 minicolumn. The low cost procedure for preparation of CH3(203)HgCl can be completed within 2 h and yields an inorganic 203Hg-free CH3(203)HgCl which is useful in methylmercury toxicology.

MAEDA M., ABIKO N., SASAKI T.

cis-Diamminoplatinum (II) complexes with selenoguanine, thioguanine, 6-thioxanthine, or 6-mercaptopurine were synthesized by the reaction of stoichiometric amounts of selenopurine or thiopurine with aquated cis-dichlorodimmineplatinum (II) in slightly acidic medium, and their antitumor activity was studied against L1210 cells in mice. These compounds exhibited a medium antitumor activity with very low toxicity. The antitumor activity was dependent on the nature of the purine ligand. These complexes were very stable in various aqueous solvents at 37 degrees C for 10 d but not in the presence of mouse serum. The mechanism of the action effected by the complex is not clear. However, the slow release of an antitumor active purine from the complex, SeG-Pt (NH3)2, was observed.

TORII H., YOSHIDA K., KOBAYASHI T., TSUKAMOTO T., TANAYAMA S.

After oral administration of 14C-labeled idebenone (14C-CV-2619) to rats, the plasma 14C level reached a plateau at 15 min, which persisted till 8 h and then decreased with a half-life of 4.5 h. In dogs, after oral dosing, the plasma 14C peaked at 15 min, followed by biophysical decline with half-lives of 2.2 and 15.4 h. The plasma of both animals contained mostly metabolites, with a small amount of unchanged CV-2619, which was greater than 90% protein-bound. In rats given 14C-CV-2619 orally or intravenously, 14C was distributed widely in tissues, with relatively high concns. in the gut, liver and kidney. CV-2619 readily entered the rat brain to undergo subcellular distribution with a significant amount localized in mitochondria. The concn. of 14C in rat fetus was low, as was that in the milk. Oral 14C-CV-2619 was eliminated by rats and dogs mostly as metabolites within 48 h. In rats, more was excreted in urine than in feces, whereas in dogs excretion by these two routes was almost equal. Enterohepatic cycling of biliary 14C occurred in rats. Repeated oral ingestions of 14C-CV-2619 to rats resulted in no accumulation of 14C. The metabolites found in rats and dogs were QS-10, QS-8, QS-6 and QS-4 formed by oxidative shortening of the side chain of CV-2619, and desmethylated CV-2619 and QS-4. Glucuronides and sulfates of the dihydro (quinol) derivatives of the above metabolites were also detected. Dihydro QS-4 sulfate was the major metabolite in plasma and urine of both animals, while dihydro QS-10 glucuronide was predominant in rat bile.

YAMAZAKI M., ARAI A.

The administration of matrine, which has antiulcerogenic activity against stress ulcer, exhibited clear contraction on the preparation of the fundus strip of rats at high concentration (from 2 X 10(-4) g/ml). The contractile response of the fundus strip to matrine was not inhibited by treatment with tetrodotoxin, and was not modified with atropine at 10(-4) g/ml, while pretreatment of the fundus strip with antihistamines abolished or reduced the contractile response. The maximum contractile response of the fundus strip was not altered significantly by pretreatment of the rat with indomethacin.

SASAKI T., UCHIDA N.A., UCHIDA H., TAKASUKA N., KAMIYA H., ENDO Y., TANAKA M., HAYASHI T., SHIMIZU Y.

Aqueous extracts (macromolecule fractions in particular) of 25 species of well-known marine animals were tested for their antitumor activity against transplanted sarcoma 180 solid form in ICR mice. All of the macromolecule fractions from the extracts of animals in the shellfish category except that of Mytilus edulis inhibited the growth of sarcoma 180. The use of aqueous extracts from Strongylocentrotus nudus, Halocynthia hilgendorfi f. ritteri, Styela plicata, Ecteinascidia turbinata, and Megabalanus rosa also revealed relatively high inhibition ratios of over 60%.

YOKOHAMA S., YOSHIOKA T., KITAMORI N., SHIMAMOTO T., KAMAD A.

The absorption mechanisms of γ-butyrolactone-γ-carbonyl-L-histidyl-L-prolinamide citrate (DN-1417) and thyrotropin-releasing hormone (TRH) were studied in the rat. In situ absorption experiments were carried out by radioimmunoassay, and experiments using everted sacs of small intestine were by radioactivity measurements with 14C-labeled DN-1417 or 3H-labeled TRH in the low concentration range of drug and by a high pressure liquid chromatography in the rather high concentration range of drug. The site specificity of absorption in the small intestine of rats could not be found with DN-1417 whereas TRH-T was absorbed from only the upper part of small intestine. Dose-proportional absorption of of DN-1417 was observed in experiments of in situ as well as in vitro. Dose-proportional transfer of DN-1417 through the everted small intestine was also found within the concentration range from 120 ng/ml to 27 mg/ml, whereas the transfer ratio of TRH decreased with increase in the concentration of TRH. DN-1417 transfer from mucosal to serosal fluid was not inhibited by the replacement of medium Na ions by K ions, pretreatment of intestinal mucosa with HgCl2, the existence of an oligopeptide, or the existence of β-lactam antibiotics which had been reported to be absorbed by active transport or carrier-mediated transport systems. While, TRH transfer was inhibited by the replacement of medium Na ions by K ions, pretreatment of intestinal mucosa with HgCl2, the existence of an oligopeptide, and the existence of β-lactam antibiotics. From these results, it was concluded that DN-1417 transfer through the small intestine was mainly due to the simple diffusion, and the contribution of the carrier-mediated transport to TRH transfer through the small intestine was not disregarded

UEKAMA K., OTAGIRI M., UEMURA Y., FUJINAGA T., ARIMORI K., MATSUO N., TASAKI K., SUGII A.

Inclusion complex of prednisolone with beta-cyclodextrin (beta-CyD) in 1:2 molar ratio was prepared and its dissolution, membrane permeation and oral absorption behaviors were examined. The rates of dissolution and permeation through a cellophane membrane in water were significantly increased by beta-CyD complexation. A crossover bioavailability study was performed using human subjects with lower doses of prednisolone tablets, where the plasma levels of the drug were determined by radioimmunoassay. The enhanced bioavailability of prednisolone by beta-CyD complexation suggested the possibility of smaller doses and fewer side effects in prednisolone therapy.

ISHIDA Y., SASAKI Y., KIMURA Y., WATANABE K.

Selectivities were examined between alpha 1 and alpha 2-adrenolytic activities of two apogalanthamine analogs (dibenzazocine derivatives), 6-methyl-5,6,7,8-tetrahydrodibenz[c,e]-azocine (DA-VIII-Me) and its N-allyl analog (DA-VIII-allyl), and two dibenzazepine derivatives, azapetine (DA-VII-allyl) and its N-methyl analog (DA-VII-Me). The alpha-adrenolytic activities were evaluated as inhibitory effects against the response to norepinephrine of isolated vas deferens and anococcygeus muscle of rats. The pA2 values for DA-VIII-Me, DA-VIII-allyl, DA-VII-Me and azapetine on the isolated rat vas deferens were 7.32, 7.76. 6.61 and 7.78 respectively, which were similar to those on the anococcygeus muscle. The alpha 2-adrenolytic activities of these compounds against the twitch-inhibitory response to clonidine in transmurally stimulated rat vas deferens and guinea-pig ileum were less potent than those against the above alpha 1-adrenoceptors. In addition, their inhibitory activities on the aggregation of human platelets induced by norepinephrine were weaker than those of phentolamine and yohimbine. These results indicate that the apogalanthamine and azapetine analogs tested are more selective in blocking alpha 1-adrenoceptors than alpha 2-adrenoceptors.