Cartap concentrations in rice plant treated with soluble powder, dust and granular formulations were measured, and the toxic effect to the rice leafroller on rice plant examined. The initial concentrations of cartap in the leaf blade of the plant treated with soluble powder and dust were 49 and 5.7ppm respectively; only one tenth of these initial concentrations remained 7 days after application. In submerged application of the granule, cartap concentration in the leaf blade reached a maximum of 1.5ppm 3 days after application, and then slowly decreased. LD₅₀ values for cartap applied topically decreased from 61μg/g to 19μg/g from 2nd larval instar stage to 5th one. In the leaf dipping method, however, LC₅₀ values for 1st to 5th instar larvae ranged from 0.45 to 4.4ppm. In the root dipping method, LC₅₀ values were 0.51ppm for 1st instar, and 8.0ppm for 5th instar larvae. Terms of exceeding the LC₅₀ levels for 1st and 5th instar larvae in cartap concentration in the leaf blade were 29 and 8 days for soluble powder, 19 and 1 days for dust, and 18 and 0 days for granule application, respectively. The feeding amount of each instar larva without cartap ranged from 0.18 to 0.24cm²/mg/day. When larvae ate leaf blade which had been dipped in cartap solution of several concentrations, there was a high negative correlation between mortality and feeding amount rate.

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The effectiveness of pentachlorobenzyl alcohol (PCBA) in controlling rice blast was increased progressively by reduction of the particle size in formulations. For good biological performance, the particle size had to be smaller than 5μm in diameter. To achieve such fine particle size, wet-grinding with an agitator mill, Attritor^®, proved superior to dry-grinding with an air mill, Jet-O-Mizer^®. Technical grade PCBA was easily ground with an Attritor to particles less than 2μm as median diameter; most of the particles were less than 5μm in diameter. PCBA suspension concentrates were prepared by wet-grinding with an Attritor. The controlled flocculation of particles was necessary to prevent the hard caking of PCBA particles caused by free sedimentation in suspension concentrates. Flocculation was controlled by adjusting both particle size and the amount of sodium lignosulfonate added as a dispersant. Such a formulation was physically stable for at least 5 years.

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Five samples of 30% binapacryl suspension with different particle size distributions were tested against adult females of the citrus red mite, Panonychus citri McGregor. Three of them, having 17.7, 7.4 and 1.5μm as weight mean diameter (D_w), were prepared by repeated wet-sedimentation in the particle size range of 20-15, 10-5 and <3μm; two others with D_w of 2.9 and 1.7μm were prepared merely by wet-grinding with an agitator mill, Attritor^®. In laboratory tests, acaricidal activity at room temperature increased as the particle size of binapacryl was reduced. In greenhouse tests, residual activity also increased to some extent at 15°C, but apparently decreased at 30-35°C with the reduction of particle size. In a field test in the spring, the acaricidal effect was found to increase as the particle size decreased. However, in a summer field test, the medium-fine suspensions (2.9 and 7.4μm as D_w) were superior in effectiveness to the finest one (D_w of 1.7μm) and the coarsest one (D_w of 17.7μm).

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After single oral administration of near-lethal and sublethal doses of fenitrothion (5, 25, 250mg/kg), diazinon (300mg/kg) and methyl parathion (10mg/kg), and repeated administration of fenitrothion (5, 25mg/kg for 12 weeks), effects on mitochondrial respiratory functions and microsomal drug-metabolizing enzyme activities in the liver of rats were examined. Single oral administration of these thiophosphoryl compounds hardly affected mitochondrial ADP/O ratio and ATPase activity in the liver, whereas a single dose of 250mg/kg fenitrothion caused a slight decrease in the mitochondrial respiratory control ratio in male rats 24hr after administration. Single oral administration of near-lethal doses of the thiophosphoryl compounds produced a marked decrease in hepatic P-450 content, aniline hydroxylase and aminopyrine N-demethylase activities in male and female rats 1 to 72hr after administration. However, these changes recovered 72hr to 1 week after administration. Repeated administration of 5 and 25mg/kg/day fenitrothion for 12 weeks hardly affected microsomal P-450 content and the drug-metabolizing enzyme activities in male and female rats. Therefore, it is reasonable to consider that the effects on liver mitochondria and microsomes of near-lethal doses of these thiophosphoryl compounds are reversible.

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On single oral administration of each of the ¹⁴C-acid, -alcohol and -CN-labeled preparations of fenvalerate [(RS)-α-cyano-3-phenoxybenzyl-(RS)-2-(4-chlorophenyl) isovalerate] and the [2S, αS]-isomer at 4.2 to 30mg/kg, the radiocarbon from the acid and alcohol moieties was almost completely eliminated from the body of rats and mice, and the ¹⁴C tissue residues were generally very low. On the other hand, radiocarbon from the CN-labeled preparations was somewhat slowly excreted and higher ¹⁴C residues were found in the hair, skin and stomach contents. There were no apparent differences in the total ¹⁴C recovery, biological half-life and ¹⁴C tissue residues in the two compounds or sexes. Two weeks pre-feeding of 500ppm unlabeled fenvalerate resulted in more complete elimination of the radiocarbon from the body and lower ¹⁴C tissue residues in rats and mice given a single dose of CN-labeled fenvalerate, as compared with the non-feeding treatment. Both fenvalerate and the [2S, αS]-isomer were similarly metabolized mainly by oxidation at the 2′- and 4′-phenoxy positions of the alcohol and at the C-2 and C-3 positions of the acid moiety, cleavage of the ester linkage, conversion of the CN group to SCN- and CO₂, and conjugation of the resultant carboxylic acids and phenols with glucuronic acid, sulfuric acid and/or amino acids. Although no significant differences in the nature or amount of metabolites were seen between the sexes, dose levels or isomers, the following remarkable species differences were found: 1) taurine conjugate of 3-phenoxybenzoic acid was found in mice, but not in rats, 2) 4′-hydroxylation of the alcohol moiety and sulfate conjugate of 3-(4′-hydroxyphenoxy)benzoic acid occurred to greater extents in rats than in mice and, 3) a greater amount of thiocyanate was excreted in mice than in rats.

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ICR mice and Wistar rats of both sexes were fed a diet containing ZnSO₄ at 0, 300, 3, 000 and 30, 000ppm for 13 weeks. Animals in the 30, 000ppm group showed retarded growth along with low food intake, abnormal values in a few hematological parameters and regressive changes of the pancreatic exocrine gland. In addition, mice had decreased water intake and significant deviations in biochemical parameters, toxic lesions appeared in the stomach, intestine and spleen of both sexes and in the kidney of females. Four males and one female mouse were found dead or moribund during the study. The maximum no-effect level of ZnSO₄ was determined to be 3, 000ppm, which is approximately equivalent to the following milligram doses: mice; male 458mg/kg/day, female 479mg/kg/day, rats; male 234mg/kg/day, female 243mg/kg/day.

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In the previous paper (Part 1), it was revealed that the inhibitory effect of basic copper sulfate on the growth of Diaporthe citri depends on Cu²⁺ ion concentrations but not on basic copper sulfate concentrations. In order to further clarify the correlation between Cu²⁺ ion absorption and its inhibitory effect, we observed the amount of Cu²⁺ ion absorption in each organ of D. citri with an electronprobe X-ray microanalyzer. Germination-inhibited pycnospores contained about 1.6 times as much copper as germinating ones. Penetration-inhibited hyphae contained about 4.3 times as much copper as penetrating hyphae. Thus, a positive correlation between Cu²⁺ ion absorption in each organ of D. citri and its inhibitory effect on pycnospore germination and hyphal penetration was found. Furthermore, germination-inhibited pycnospores contained more copper than penetration-inhibited hyphae, which coincides with the previous observation that inorganic copper inhibits hyphal penetration of D. citri (50% inhibition at pH 6; Cu²⁺=1.2×10^-5M) at lower concentrations than it does pycnospore germination (50% inhibition at pH 6; Cu²⁺=4.2×10^-5M).

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Effectiveness of several insecticides for the control of tobacco cutworm, Spodoptera litura (F.), was evaluated in field and laboratory tests. Liquid and dust formulations of these insecticides were applied to soybean cultured in fields alternating with rice paddys at the time of the scattering of the insect's newly hatched and full-grown larvae. In early stage application, most of the test insecticides were effective in the indexes of corrected density of survivals, while in later stage application, methomyl, acephate, EPN, and chlorpyrifos methyl were effective in the indexes and the observation of injurious leaves of the plant. LD₅₀ of these insecticides against 3rd, 4th, and last instar larvae reared on an artificial diet were determined by a topical application method. The values for 3rd instar larvae were in the range of 0.16-4.6μg/insect, while those for last instar larvae were in the range of 2.8-66.8μg/ insect. Less than 5μg/insect of LD₅₀ for last instar larvae, which seemed to be the level of direct deposition on larval body, were demonstrated by methomyl, chlorpyrifos methyl and profenofos. LC₅₀ of test insecticides determined by the leaf dipping method were in the range of 12.7-121ppm for 3rd instar larvae and 44.3-711ppm for last instar larvae.

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Rice plants of the 3 to 4 leaf stage were cultivated in a nutrient solution containing ¹⁴C-mepronil at 10ppm and the metabolism of mepronil in the rice plants 5 and 10 days after application was studied. Radioactive component unextracted with methanol-water (2:1) increased gradually, reaching 20.1% (5 days) and 35.1% (10 days) of the total absorbed radioactivity. This bound fraction gave 3-isopropoxyaniline and 3-anisidine after acid hydrolysis. In the methanol-water soluble fraction mepronil represented the major component; 68.0% (5 days) and 51.8% (10 days) of the absorbed radioactivity. Eleven metabolites were found as the free or conjugated form. Among them, 3′-hydroxy-2-methylbenzanilide, 3′-methoxy-2-methylbenzanilide, o-toluic acid, 3′-isopropoxy-2-hydroxy-methylbenzanilide and 3′-(1-hydroxymethyl)ethoxy-2-methylbenzanilide were identified. Accordingly, mepronil seems to be hydroxylated at three positions and also cleaved at the amide linkage. Leaf treatment and stem injection by mepronil gave similar results. Mepronil translocated gradually upwards from the application sites and no distinct downward translocation was observed.

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[in Japanese]

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Two types of metabolic activation of triaryl phosphate to cause delayed neurotoxicity were disclosed: 1) tri-o-tolyl phosphate (TOCP) was converted into a highly active saligenin cyclic phosphate, 2-(o-tolyoxy)-4H-1, 3, 2-benzodioxaphosphorin 2-oxide, by the microsomal hydroxylation of a methyl group followed by cyclization, which was accelerated by plasma albumin; 2) tri-p-ethylphenyl phosphate (TPEP) was activated by similar hydroxylation at α-carbon atom followed by dehydrogenation to form α-oxo metabolites. Studies on structure-activity relationships by modifying the structure of the neurotoxic metabolites showed that the selective toxicity is governed greatly by the size of non-leaving groups (selectophore) and that the combination of the selectophore and phosphorylating activity, which is due to the cyclic ester structure in TOCP series and to the electron-withdrawing leaving group in TPEP series, decides the final biological activity of phosphorus compounds. Thus, the metabolite homologs carrying small alkyl group (s) as the non-leaving group in both series did not cause ataxia in hens, but exerted insecticidal activity. Salithion (2-methoxy-4H-1, 3, 2-benzodioxaphosphorin 2-sulfide) was therefore developed into a commercial insecticide. The phosphorylating property of saligenin cyclic phosphate esters was also utilized for the syntheses of biologically interesting natural phosphate esters by developing the thiolate isomer of salithion, MTBO. This has the advantage of oxidatively activatable thiolate ester group, in addition to the highly reactive cyclic ester structure of saligenin which also acts as a removable protective group. Some potent synergists of malathion were also found in the series of cyclic phosphorus esters related to the TOCP-metabolite. They were the potent inhibitors of carboxyesterase. Another synergist found was a thioquinol phosphate, which was obtained by modifying the metabolite of triphenyl phosphate, a malathion synergist. Another thiolate-type synergist IBP inhibited the degradation of malathion by inhibiting carboxyesterase and also demethylation in vivo. Finding L-leucine to be a neuroactive substance in the blood of silkworm larvae, we combined it and related amino acids with an extremely reactive five-membered cyclic phosphate and obtained novel insecticidal 4-isobutyl-2-methoxy-1, 3, 2-oxazaphospholidine 2-sulfide and the 4-isopropyl homolog. These were particularly effective to organophosphate resistant strains of houseflies. The structuretoxicity relationship of bicyclic phosphate esters, 4-substituted 2, 6, 7-trioxa-1-phospha-bicyclo [2.2.2] octanes, was characterized by the importance of the steric and hydrophobic properties of the bridge-head substituent for the high toxicity, which appeared to be due to their anti-GABA activity.

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Studies were begun to determine the effectiveness of N-aryl-3, 4, 5, 6-tetrachlorophthalamic acids against bacterial leaf blight of rice, caused by Xanthomonas campestris pv. oryzae. The structure-activity relationship of many derivatives indicated that the presence of both the carboxyl radical and four chlorine atoms at phthalic acid moiety was structurally essential for effectiveness against the disease, and optimization of activity was obtained in a derivative (techlofthalam) having 2, 3-dichlorophenyl as the aryl group. Techlofthalam showed no activity in vitro against the causal bacterium, when estimated by a turbidimetric method. Soil application with techlofthalam was found to be considerably effective at 4kg per hectare both in the greenhouse and the field. With foliar application, however, extremely different efficacies were shown with two inoculation methods; excellent control against lesion development was realized by spray inoculation, but only poor control was obtained by needle prick inoculation. Intensive field evaluation proved that techlofthalam spray at 60 to 120g per hectare showed a stable and excellent effect. In order to know the mode of action of techlofthalam, the change in population and aggressiveness of the bacterium in leaves sprayed with a 100ppm solution were examined. There was a definite trend of change in bacterial populations at all spray times, 1 day before and 3 days after inoculation, and after lesion formation; the bacterial population started to decrease a few days after spraying, and then was stable at much lower levels. The bacterium isolated from a sprayed leaf had lower aggressiveness than one from an unsprayed leaf. In consideration of generation time (approximately 10 hours in the exponential growth phase) of the bacterium in rice leaf, these results suggested techlofthalam's direct inhibitory effect on the multiplication of bacterium on the leaf. Retardation of the increase in viable bacterial population was found by a static culture using Suwa's synthetic medium containing techlofthalam at 0.1ppm; this was practically the same concentration as that gaschromatographically determined in guttation droplets on rice leaves a few days after spraying. Some other results also gave support to the conclusion that techlofthalam displays bacteriostatic action in rice leaf. Techlofthalam has been practically used in Korea since 1979.

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The physiological and biochemical metabolic fate of 1, 2-dibromo-3-chloropropane (DBCP) was studied in relation to the mammalian toxicity of the chemical. On oral administration of ¹⁴C-DBCP to male rats at 20 to 400mg/kg which was the lethal dose causing death in 24hr with a severe necrotic lesion in the liver and kidney (inner cortex), the ¹⁴C at all dose levels maldistributed in these target organs (and the target organ site). Radioactive material covalently bound to proteins was the predominant radioactive component in these target organs and dose-dependently increased with a concomitant severe depression of hepatic glutathione (GSH). Formation of ¹⁴C-urea which implies a de novo synthesis of ¹⁴C-labeled proteins and nucleic acids via the C₁-pool was not detected. Alkylating potential of DBCP was evidenced by in vitro binding experiments in which ¹⁴C-incorporation into the liver-9, 000g supernatant and microsomal (ms) proteins required an oxidative metabolic transformation of DBCP by ms-cyt P₄₅₀ enzyme system, but this was not dependent on the protein synthesis. Low molecular-weight SH compounds such as GSH prevented the alkylation of the macromolecules; however, no prooxidant action indicating involvement of a DBCP-free radical was detected. A low concentration of 1, 1, 1-trichloro-propane-2, 3-oxide, an inhibitor of ms-epoxide hydrase, stimulated the alkylation of the macromolecules. Thus, DBCP was concluded to be oxidatively converted into a reactive epoxide (s) and covalently bound with the nucleophilic sites of the cellular macromolecules in vitro and in vivo. Investigations on the toxicological and metabolic alterations after pretreatment (ip) of rats with phenobarbital, SKF-525A, and GSH demonstrated that the severity of the necrotic lesion in the centrilobular hepatocytes induced by 400mg/kg of DBCP was closely dependent on the alkylation of the hepatic macromolecules.

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