Spore germination, hyphal elongation and hyphal penetration of Alternaria kikuchiana were observed on alcohol-treated epidermal strips of onion bulb. I₅₀ of copper sulfate, 8-quinolinol and copper 8-quinolinolate on germination of A. kikuchiana were 10^-3M, 10^-4M and 10^-6M respectively, and those on penetration were 3×10^-4M, 3×10^-5M and 10^-6M respectively, indicating that copper ion and 8-quinolinol acted synergistically against A. kikuchiana. The synergism was more remarkable when 8-quinolinol existed more than twice the concentration of copper ion. On the contrary, the synergism was less remarkable when copper ion existed more than a half the concentration of 8-quinolinol. 8-Quinolinol at 10^-2M accelerated state 4 respiration of rat liver mitochondria to 2.5 times and showed uncoupling effect on oxidative phosphorylation at higher than 10^-3M. 8-Quinolinol also accelerated respiration of the mitochondrial fraction of A. kikuchiana to 1.4 times at 2×10^-3M. Thus a possibility was, suggested that the inhibitory effect of 8-quinolinol on the growth of A. kikuchiana is related to the uncoupling effect. Since mitochondrial membrane is generally permeable to uncouplers, the fungal cell membrane which is similar to mitochondrial membrane by nature may also be permeable to 8-quinolinol and permit copper ion to pass the barrier along with 8-quinolinol as a complex to afford the synergistic fungicidal action of the both chemicals. Further, there is a possibility that 8-quinolinol makes it possible for copper ion to pass through mitochondrial innerr membrane of A. kikuchiana and inhibit not only 6-phosphogluconate dehydrogenase in extramitochondrial cytoplasm, but also many dehydrogenases in mitochondria.

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Synthetic pyrethroids and DDT analogs increase the frequency of spontaneous discharges in the abdominal nerve cord excised from the crayfish Procambarus clarki drastically at concentrations of a certain range. This spontaneous neuroexcitatory activity is dependent upon the temperature during treatment. Most of these compounds also induce depolarization of the axonic membrane at rest in the circumesophageal connectives of crayfish, although at concentrations higher than those required for the activity on spontaneous discharges in the abdominal nerve cord. The spontaneous discharges caused by the insecticides continue for more than two hr after the start of the treatment, while discharges induced by acetylcholine subside immediately after a transient firing. Neuroexcitation by these insecticides is suppressed by either prior or simultaneous treatment with γ-aminobutyric acid.

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Sixty two kinds of monoterpenoids were tested by means of the test tube method and the beaker method as space repellents against the German cockroach (Blattella germanica L.). (-)-Linalool and nerol in lenear monoterpenoids, (-)-carvone, (+)-pulegol, (+)-pulegone and (+)-isopulegol in monocyclic monoterpenoids are the most effective space repellent found to date. However, thirty-two kinds of monoterpenoids, for example, (+)-linalool, (+)-limonene, linalyl acetate, bicyclic monoterpenoids and so on, are low or not effective in action. A correlationship of repellent potency to volatility was not demonstrated within the series. It has been suggested that repellent potency within this series is not dependent upon volatility and molecular size or optical rotation, but dependent primarily upon molecular configuration and the position of some functional groups.

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The growth inhibiting activity of 74 N-(1-methyl-1-phenylethyl) acylamides against bulrush, Scirpus juncoides was determined with the vial test. The structure-activity relationships were analyzed with the multiple regression technique using such physicochemical substituent parameters as hydrophobic π and steric E^c_s values of alkyl group of the acyl moiety. The variation in the activity was shown to be related parabolically with variation in both the hydrophobicity and steric bulkiness of the alkyl substituents. When the second largest subalkyl group at the α-position of the substituent is bulkier than methyl and the unsaturated bond in the substituent conjugates with the carbonyl group of the amide, the activity is to be lowered.

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The growth inhibiting activity of 27 derivatives of N-benzyl-2-bromo-3, 3-dimethylbutanamide having various substituents at the α-position and the benzene ring of the N-benzyl group against bulrush, Scirpus juncoides, was determined in the vial test. The structure-activity relationships were analyzed quantitatively by physicochemical substituent parameters and multiple regression analysis. The variation in the activity of N-(α-substituted benzyl) derivatives was shown to be related parabolically with variation in the steric dimension of the substituents. The greater the hydrophobicity of substituents is, the greater the activity is. The variation in the steric dimension of the benzene ring substituents was responsible for the activity of N-(1-methyl-1-substituted phenylethyl) derivatives.

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The herbicidal activity of 45 N-benzylbutanamides against Cyperus rotundas was determined by the pot test with pre-plant soil incorporation method. The variation in the activity of N-(1-methyl-1-phenylethyl)-substituted acetamides was shown by the Hansch analysis to be related with the variation in the steric bulkiness of the substituent. The haloalkyl substituent was detrimental to the activity. Among the N-(1-methyl-1-phenylethyl)-2-ethyl-3, 3-dimethylbutanamides having various substituents on the benzene ring, 4-methylphenyl derivative exhibited the highest activity.

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A microanalytical method of mildiomycin for residue analysis in cucumber fruits, tobacco leaves and soil was developed. Mildiomycin in cucumber fruits and tobacco leaves was extracted with water and cleaned up by CM-Sephadex column chromatography. Mildiomycin in soil was hardly extracted with water, but sufficiently with saturated sodium pyrophosphate solution. The extract was desalted and cleaned up by CG-50 column chromatography. Mildiomycin reacted at pH 8.0 with fluorescamine, a reagent for the fluorometric quantitation of primary amine, and was determined by ion-pair reversed-phase high performance liquid chromatography with fluorometric detection. The limits of detection in cucumber fruits, tobacco leaves and soil were 0.06ppm, 0.2ppm and 0.7ppm respectively. The recoveries of mildiomycin from cucumber fruits, tobacco leaves and soil at respective fortification levels of 1.2, 3 and 10ppm were over 70%.

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All dialkyl 1, 3-dithiolan-2-ylidenemalonates examined are considerably stable in rice plants, showing their t_1/2 of 2.0-3.8 days. A non-cyclic analog with the t_1/2 of 0.8 day, however, is rapidly metabolized probably through the oxidation at its thioether groups. Although the rat liver homogenate has both oxidation and hydrolysis activities, it shows predominantly the esterase activity in the absence of NADPH. In the rat liver homogenate, again, the non-cyclic analog is much more susceptible to oxidative metabolism in the presence of NADPH than, but as stable against the ester hydrolysis as the cyclic counterparts. The ester hydrolysis rates of the compounds well relate to the branching (s) at the α-position or the resulting electronic (σ*) or steric (E^c_s) effects of alcoholic alkyl groups.

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Chlobenthiazone (S-1901), 4-chloro-3-methyl-2(3H)-benzothiazolone, is a new fungicide for the control of rice blast. Greenhouse studies indicated that the fungicide had high preventive and residual activities at low concentrations against the disease. On the contrary, curative activity was not obtained even at a concentration as high as 500μg/ml. The fungicide was also effective in controlling the disease by submerged application, indicating systemic movements in plants. When ¹⁴C-labeled chlobenthiazone was applied to the leaves or roots of rice seedlings, radioactivity was found to be rapidly translocated to the untreated parts of the plants, confirming the result in pot trials. Furthermore, the rice seedlings exposed to the fungitoxic vapor were protected from the disease. It is suggested that both the systemic and the vapor phase activities must contribute to the effective disease control in fields. Some of the field trials demonstrated that chlobenthiazone controlled rice blast by foliar or submerged application.

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It was observed that the apparent degradation of dimethoate in a closed system is significantly greater than that in an open system, although their difference is not so large as in the case of fenitrothion. For this observation, an autocatalytic degradation mechanism is proposed which includes S-methyl isomerization reaction catalyzed by both carbamoyl group in the dimethoate molecule and dimethyl sulfide gas which is evolved during further decomposition of S-methyl-dimethoate, where the catalytic role of dimethyl sulfide is relatively small. Stabilization of dimethoate by the adduct formation with 4-methyl-2-t-butylphenol and by the inclusion into β-cyclodextrin were established, and their stabilization mechanism was discussed on the basis of their physicochemical properties and the proposed degradation pathway.

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Metabolism and organ distribution in mice has been investigated following intraperitoneal injection of the insecticide [methyl-¹⁴C] Gardona (tetrachlorvinphos). During the first 24hr, about 40% of the applied dose was eliminated in the expired air and urine. The ¹⁴CO₂ accounted for 14% of the administered dose. Three acidic ¹⁴C-metabolites in the urine were isolated and identified as monomethyl phosphate, dimethyl phosphate and desmethylgardona. Gardona is mainly metabolized by cleavage of the P-OCH₃ bond followed by hydrolysis at the vinyl-ester bond to give monomethyl phosphate. Minor pathways involve initial attack at the P-O-vinyl ester bond to yield dimethyl phosphate and oxidative cleavage of O-methyl group to yield formate via formaldehyde. The 2-chloro-1-(2, 4, 5-trichlorophenyl) vinyl moiety is excreted in the urine in different modifications. The contribution of the P-O-methyl cleavage to detoxification of the insecticide has been discussed.

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Monolayer cultures of human embryonic lung diploid cells (HEL 299) were exposed to an insecticidal chemical, Carbaryl (1-naphthyl N-methylcarbamate), and its effect on nucleic acid and protein syntheses was measured. No significant differences among the effects on DNA, RNA and protein syntheses were observed. The rates of the syntheses of the macromolecules were inhibited severely at a concentration of 10^-3M. At lower dose levels between 10^-7M and 10^-5M, the syntheses of the macromolecules were virtually unaffected.

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Spores of Pyricularia oryzae treated with tricyclazole [5-methyl-1, 2, 4-triazolo(3, 4-b)benzothiazole] at concentrations higher than 0.1μM produced hyaline appressoria on cellulose membrane. The appressoria lacked the ability to penetrate the membrane. The inhibitory effect of tricyclazole (0.2μM) was nullified completely by the addition of 3, 4-dihydroxyphenyl alanine (DOPA) (10μM) and partially by the addition of 1, 8-dihydroxynaphthalene (1, 8-DHN) (50μM) but not by that of scytalone. DOPA and 1, 8-DHN also colored hyaline appressoria of a melanin deficient mutant with no ability to penetrate cellulose membrane isolated from a wild type P. oryzae and the melanized appressoria recovered the ability to penetrate cellulose membrane. The results presented in this paper suggest that it is the lack of melanization in appressoria that caused failure of penetration from appressoria in P. oryzae treated with tricyclazole and in its melanin deficient mutant.

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[in Japanese]

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[in Japanese]

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The data on the 50% inhibitory concentration (IC₅₀) of various pesticides and related compounds to cultured animal and human cells as reported in the literature are compiled and they are compared with the LD₅₀ values. Although no correlation between LD₅₀ of chemicals for mice and rats and the concentrations toxic to cultured cells is found, this summary will serve as basic data for the development of new toxicity test methods using cell culture systems.

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A method was developed to test the inhibitory effects of chemicals on the growth of rice blast fungus, Pyricularia oryzae, by observing all the fungal growth stages in a single system. A drop of conidial suspension of the fungus was placed on epidermal strips of onion bulb and kept in a moist chamber at 25°C. Conidial germination began after 3hr and attained to a level of 90-100% within 5hr. Appressorial formation began immediately after conidial germination, reached 70-89% after 5hr, and nearly 100% 20hr after the inoculation. Granulation of epidermal cells occurred after 22hr, appressorial penetration into the epidermal cells started within 24hr and reached about 80-90% 27hr after inoculation. At the same time, mycelial growth was remarkable on epidermal cells. Sporulation was accelerated within 2 to 4 days after inoculation. Fifteen min after the water drops were blotted off the epidermal strips during the fungal growth, water was again dripped onto the same strips. The results showed that the fungal growth was not disturbed by this procedure, but rather promoted. We would like to propose that this method be termed the “onion epidermal strip method.”

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Pentachlorophenol (PCP) and its sodium salt have been widely used as agricultural and industrial pesticides, especially as herbicides in paddy fields during the decade from 1960 in Japan. Degradation of PCP in soils and metabolism of PCP by isolated soil microorganisms have been intensively studied. The rate of PCP dissipation from soils under moist conditions varied among soils, and half-life of PCP was 7-14 days. The dissipation of PCP was influenced by the presence of certain fungicides as well as autoclaving. Another studies clarified the dissipation of PCP was also reduced in submarged condition. Repeated application of PCP to soils resulted in acceleration of the PCP dissipation. The evolution of ¹⁴CO₂ from soil treated with ¹⁴C-PCP was observed. From these results it appeared that the dissipation of PCP depended on microbial decomposition. Pseudomonas sp. isolated from a soil degraded ¹⁴CPCP rapidly with release of approximately 50% of the ¹⁴C as ¹⁴CO₂ and incorporated the ¹⁴C into the bacterial cell constituents such as amino acids. Tetrachlorocatechol (TCC) and tetrachlorohydroquinone (TCHQ) were identified as degradation products. Metabolic pathways of ortho or para hydroxylation of PCP as initial degradation step followed by rapid ring cleavage were proposed. PCP was methylated to pentachloroanisole (PCA) by isolated Mycobacterium sp. The isolate simultaneously metabolized PCP to TCHQ followed by successive methylation to form their dimethyl ethers, tetrachloro-1, 2-dimethoxybenzene or tetrachloro-1, 4-dimethoxybenzene, via monomethyl ethers, tetrachloro-2-methoxyphenol or tetrachloro-4-methoxyphenol. Chlorothiophenols as well as chlorophenols were also methylated by this bacterium. When either methionine, ATP, and Mg⁺⁺ or S-adenosylmethionine was incubated with the cell-free extracts of Mycobacterium sp. in phosphate buffer containing PCP, PCA was produced. Incubation of methyl-¹⁴C labeled S-adenosylmethionine with PCP by cell-free extracts resulted in transfer of the radioactivity to PCA. These results revealed that the methylation of PCP by isolated Mycobacterium sp. was attributable to the enzyme(s) concerned with the transferance of methyl group to PCP.

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On the basis of the natural product models and biochemical designs, attempts were made to find leads for the development of new pesticide chemicals. Antilipidemic agents of several different chemical classes and cis-7-chloro-3a, 8b-dihydro-3a-methylfuro[3, 4-b]benzofuran-3 (1H)-one, which inhibited HMG-CoA reductase activity in rats in vivo, showed ovicidal activity against the eggs of azuki bean weevils, Callosobruchus chinensis, but failed to produce clear anti-juvenile hormone (JH) activity. A number of imidazole compounds possessing a JH-like terpene chain, designed to be potential inhibitors of JH biosynthesis, were synthesized and bioassayed on the silkworm, Bombyx mori larvae. New compounds, 1-geranyl-5-phenylimidazole (A-1) and the 1-citronellyl analog (A-2), showed remarkable anti-JH activity. When treated to the third instar larvae, miniature pupae were formed after molting to the fourth instar larvae. The fifth instars were also omitted by treatment of the fourth instar larvae. The presence of a phenyl group at the 5 position of the imidazole ring is essential for anti-JH activity. The introduction of a methoxy group in the 7 position of the terpene chain of A-1 and A-2 eliminated or decreased the activity. The anti-JH effects of A-1 and A-2 on B. mori larvae were completely blocked by simultaneous application of such a JH mimic as methoprene. Benzimidazoles having a terpenoid moiety, 1-(3, 7-dimethyl-7-methoxy-2-octenyl)-2-methylbenzimidazole (B-1) and the 7-ethoxy analog (B-2), showed high insecticidal activity against houseflies and green rice leafhoppers, Nephotettix cinticeps. They inhibited also larval molting of B. mori at low doses when applied topically to the larvae 1-2 days before erdysis. 8-Isopropyl-6-oxabicyclo[3.2.1]octan-7-one (trans-lactone) with a partial skeleton of picrotoxinin, which is an antagonist of γ-aminobutyric acid, and some related lactones were synthesized and their insecticidal activities were assayed on houseflies. In the insecticidal activity of the lactone compounds the configuration of the isopropyl group at C₈-position played an important role. Thus trans-lactone had the highest activity. Ninety s-triazine derivatives were prepared by the reaction of amino acids and peptides with substituted biguanides and bioassayed on plant growth. Among the tested compounds the s-triazine derived from leucylleucine exhibited the highest growth inhibitory activity on rice.

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