Herbicide ZJ0273 Research Articles

A fusant of Amycolatopsis sp. M3-1 and Pseudomonas sp. Nai8 with high capacity of degrading novel pyrimidynyloxybenzoic herbicide ZJ0273 and naphthalene.

ZJ0273 (propyl 4-(2-(4, 6-demethoxy pyrimidin-2-yloxy) benzylamino) benzoate) is a novel pyrimidynyloxybenzoic-based herbicide developed in China for oilseed crop. This study was aimed to construct new strains capable of degrading naphthalene and ZJ0273 by protoplast fusion between Amycolatopsis sp. M3-1 and Pseudomonas sp. Nai8. Eight recombinant strains were successfully produced, and the strains could simultaneously utilize ZJ0273 and naphthalene as the sole carbon and energy source, respectively. One of recombinant strains, MN6 with higher degrading efficiency, was chosen for further study. Under the condition of pH 7.0, 30 °C, ZJ0273 and naphthalene degradation percent by the recombinant strain MN6 could reach 65.10% (20 days) and 88.46% (48 h), respectively. According to the identified six metabolites (M1-M6) by LC-MS/MS, biodegradation pathway of ZJ0273 was proposed. ZJ0273 biodegradation catalyzed by the recombinant strain MN6 involved continuous biocatalytic reactions such as de-estering, hydrolysis, acylation, C-N cleavage, de-methyl, and ether cleavage reactions.

Effects of the novel pyrimidynyloxybenzoic herbicide ZJ0273 on enzyme activities, microorganisms and its degradation in Chinese soils.

Enzyme activity and microbial population in soils have important roles in keeping soil fertility. ZJ0273 is a novel pyrimidynyloxybenzoic-based herbicide, which was recently developed in China. The effect of ZJ0273 on soil enzyme activity and microbial population in two different soils was investigated in this study for the first time. The protease activity was significantly inhibited by ZJ0273 and this inhibiting effect gradually weakened after 60-day incubation. The results also showed that ZJ0273 had different stimulating effects on the activities of dehydrogenase, urease, and catalase. Dehydrogenase was consistently stimulated by all the applied concentrations of ZJ0273. The stimulating effect on urease weakened after 60-day incubation. Catalase activity was subject to variations during the period of the experiments. The results of microbial population showed that the number of bacteria and actinomycetes increased in ZJ0273-treated soil compared with the control after 20days of incubation, while fungal number decreased after only 10days of incubation in soils. DT50 (half-life value) and k (degradation rate constant) of ZJ0273 in S1 (marine-fluvigenic yellow loamy soil) and S2 (Huangshi soil) were found 69.31 and 49.50days and 0.010 and 0.014day(-1), respectively.

The influence of new herbicide ZJ0273 on the total- and branched-chain amino acids in oilseed rape (Brassica napus L.) leaves as revealed by near-infrared spectroscopy

Propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy) benzylamino)benzoate (ZJ0273) is a new herbicide which inhibits acetolactate synthase (ALS). The ZJ0273 is considered as safe for the environment and exhibits a satisfactory effect on weed control in the rapeseed field. ALS is the key enzyme of reactions in the biosynthesis of total amino acids (TAAs) especially branched-chain amino acids (BCAAs). This study reports the effect of ZJ0273 on BCAAs and TAAs in rapeseed leaves using near-infrared spectroscopy (NIRS) techniques. A decrease in TAAs and BCAAs contents was observed as the herbicide dosages were increased along with leaf senescence. The wavelengths 2,416 and 1,340 nm were selected to develop the NIRS model for detecting BCAAs and TAAs, and correlation coefficients of model’s prediction set were 0.9823, 0.9764, 0.9831, and 0.9968 for valine, isoleucine, leucine, and TAAs, respectively. The results indicated that 100 mg/L ZJ0273 was a safe dosage for oilseed rape as it did not show a significant effect on the contents of amino acids compared to other higher dosages (500 and 1,000 mg/L).

Microbial degradation characteristics and kinetics of novel pyrimidynyloxybenzoic herbicide ZJ0273 by a newly isolated Bacillus sp. CY

ZJ0273 (propyl 4-(2-(4,6-demethoxy pyrimidin-2-yloxy)benzylamino)benzoate) is a novel herbicide developed in China for oilseed crop. Sixteen bacteria capable of utilizing ZJ0273 as the sole carbon source were isolated from soils. One of the isolates was designated as Bacillus sp. CY based on its physiological and biochemical characteristics and phylogenetic analysis of 16S rDNA sequences. The present study aimed to investigate the ZJ0273 degradation characteristics and kinetics by Bacillus sp. CY which has the ability to utilize ZJ0273 as the sole source of carbon and energy under aerobic conditions. The optimum biodegradation temperature, pH, and ZJ0273 initial concentration were 20-40 °C, 5.0-9.0, and 50-400 mg/l, respectively. Strain CY degraded 65 % of ZJ0273 (initial concentration of 50 mg/l) during 30 days of incubation in basal mineral medium at pH 8.0 and 35 °C. DT50 (half-life value), k (degradation rate constant of ZJ0273), and R (2) are 19.20 days, 0.0361 day(-1), and 0.9464, respectively.

Aerobic biodegradation kinetics and pathway of the novel herbicide ZJ0273 in soils

Degradation of ZJ0273, a recently developed pyrimidynyloxybenzoic‐based herbicide, was investigated in five different soils under aerobic conditions. ZJ0273 degradation rate was strongly affected by soil physico‐chemical characteristics and the inoculation of ZJ0273‐degrading bacteria. Greater organic matter (OM) content, neutral pH and inoculation of ZJ0273‐degrading bacteria can increase degradation rate and decrease the half‐life value (DT50). At 30°C the biodegradation rate of ZJ0273 reached 41–85% in natural (unsterilized) soils. It ranged from 69 to 96% at 90 days after treatment (DAT) in five different types of soils after re‐inoculation of Amycolatopsis sp. M3‐1 and DT50 decreased by 34 , 81, 16, 20 and 32 days, respectively, in soils S1, S2, S3, S4 and S5. Furthermore, using the six metabolites (M1–M6) identified six metabolites (M1–M6) by liquid chromatography‐mass spectrometry (LC‐MS) and their behaviour, a biodegradation pathway of ZJ0273 in soils was proposed. New metabolites, M5 and M6, were found in soils. Biodegradation of ZJ0273 involved continuous biocatalytic reactions, such as de‐estering, hydrolysis, acylation, C‐N cleavage, de‐methyl and ether cleavage reactions. Finally, ZJ0273 was bio‐transformed into M4 and M6, which could be degraded and oxidized into CO2 and H2O through the tricarboxylic acid (TCA) cycle.

The metabolism of a novel herbicide ZJ0273 in oilseed rape and crickweed

Experiments were conducted to investigate the metabolism of a novel herbicide ZJ0273 in the seedlings of the susceptible crickweed and tolerant oilseed rape. Six metabolites of ZJ0273 were identified in crickweed seedlings, compared to 8 metabolites in oilseed rape. Among these metabolites, the metabolite M7, with the chemical name of 2-(4,6-dimethoxypyrimidin-2-yloxy) benzoic acid, was found to be the most herbicidally effective molecule with significant inhibition to acetolactate synthase. The differences in the amount and rate of ZJ0273 transformation to M7, and its metabolic detoxification, contributed to the primary mechanism of herbicidal selectivity of ZJ0273.

Degradation of the novel herbicide ZJ0273 by Amycolatopsis sp. M3-1 isolated from soil

ZJ0273, propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy) benzylamino) benzoate, is a novel and broad-spectrum herbicide. In this study, 15 bacteria capable of utilizing ZJ0273 as the sole carbon source were isolated from soil. One of the isolates belonged to the family Amycolatopsis and was designated to Amycolatopsis sp. M3-1; at 30°C and pH 7.0, degradation rate of ZJ0273 could reach at 59.3% and 68.5% in 25days and 60days, respectively. Furthermore, six metabolites (M1-M6) during the degradation of ZJ0273 by Amycolatopsis sp. M3-1 were identified by a combination with multi-position (14)C-labeled compounds (B-ZJ0273 and C-ZJ0273), chromatography, liquid scintillation spectrometer, and LC-MS, a novel pathway of ZJ0273 degradation by Amycolatopsis sp. M3-1 was proposed based on the identified metabolites and their biodegradation courses. ZJ0273 was initially hydrolyzed into M1 (4-(2-(4,6-dimethoxypyrimidin-2-yloxy) benzylamino) benzoic acid), then further oxidized into M3 (2-(4,6-dimethoxypyrimidin-2-yloxy) benzoic acid). M1 also could undergo a carbonylation into M2 (4-(2-(4,6-dimethoxypyrimidin-2-yloxy) benzamido) benzoic acid), and then its C-N and C-O bonds were cleaved to yield M3 (2-(4,6-dimethoxypyrimidin-2-yloxy) benzoic acid) and M4 (4,6-dimethoxypyrimidin-2-ol), respectively. Moreover, another two new metabolites, M5 (2-(4-hydroxy, 6-methoxypyrimidin-2-yloxy) benzoic acid) and M6 (2, 4-dihydroxy-pyrimidine) were found. M5 was formed through de-methyl of M3 and then hydrolyzed into M6.

Estimation of Acetolactate Synthase Activity in Brassica napus under Herbicide Stress Using Near-Infrared Spectroscopy

The appropriateness of near-infrared spectroscopy was investigated for the fast and noninvasive estimation of acetolactate synthase (ALS) activity in oilseed rape (Brassica napus L.) leaves, which is important for studying the effects of the new herbicide propyl 4-(2-(4,6-dimethoxy-2-pyrimidinyloxy)benzylamino)benzoate (ZJ0273) on plant metabolism and physiological regulation. A total of 248 leaf samples were collected at different leaf positions and ZJ0273 concentrations. A complete comparison was performed on the preprocessing methods, which included smoothing, multiplicative scatter correction, standard normal variate, derivatives, detrending, and direct orthogonal signal correction (DOSC), as well as linear calibrations, i.e., multiple linear regression (MLR) and partial least squares (PLS), and nonlinear calibration (least squares-support vector machine, LS-SVM). In all linear (MLR and PLS) models, the optimal models were the successive projections algorithm (SPA)-MLR (DOSC) and SPA-PLS (DOSC) models, with the same prediction performance (rp = 0.938 and RMSEP = 6.587). The best prediction model was the nonlinear SPA-LS-SVM (MSC) model, with rp = 0.988 and RMSEP = 2.844. A direct linear function using raw and DOSC preprocessed spectra at 2094 nm was developed and obtained a good prediction performance. These results will be helpful for the design of ALS detection sensors and in-field monitoring systems for the growth status of oilseed rape. The results are also useful for study of the inhibitory effects, metabolic pathways, and environmental residues of the new herbicide ZJ0273 during the growth of oilseed rape.

Differential morphological and physiological responses of two oilseed Brassica species to a new herbicide ZJ0273 used in rapeseed fields

Weeds are considered as a major threat to the production of oilseed Brassica crops. The use of herbicides that are safe for crops and effective in controlling weeds is crucial for the agronomists and farmers. Propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy)benzylamino)benzoate (ZJ0273), a derivative of 2-pyrimidinyloxy- N-aryl benzoate, is a new herbicide used in the rapeseed field. To evaluate the tolerance of Brassica species against this new herbicide, two cultivars of rapeseed Brassica napus cv. ZS 758 and Brassica rapa cv. Xiaoyoucai were tested by a foliar spray of ZJ0273 at the rate of 100, 500 and 1000 mg/L and a currently used ALS (acetolactate synthase)-inhibiting herbicide bispyribac-sodium (BS) at the rate of 100 mg/L. The results showed that both the cultivars of Brassica were less affected by ZJ0273 as compared to BS. Increasing level of ZJ0273 herbicide from 100 to 1000 mg/L increased the stress for the plants of both the cultivars as indicated by enhanced accumulation of malondialdehyde content. The activities of ALS and antioxidant enzymes (superoxide dismutase and peroxidase), soluble protein and sugar contents, photosynthetic system (SPAD value, photosynthetic rate and chlorophyll fluorescence) as well as the agronomic characters also declined consistently with each successive increase in ZJ0273 concentration. In general, the plants treated with 100 mg/L ZJ0273 recovered from the herbicide stress after 28 days. B. napus showed more tolerance than B. rapa to the new herbicide. Nevertheless, BS application at 100 mg/L did not allow the plants of both the cultivars to recover from the herbicidal stress.

Plant availability and phytotoxicity of soil bound residues of herbicide ZJ0273, a novel acetolactate synthase potential inhibitor

The plant availability and phytotoxicity of soil bound residues (BR) of herbicide ZJ0273, a novel acetolactate synthase (ALS) potential inhibitor, to rice ( Oryza sativa L.) and corn ( Zea mays L.) was investigated in three different soils including a Fluvio-marine yellow loamy soil ( S 1), a Red clayey soil ( S 2), and a Coastal saline soil ( S 3), using 14C-labeling tracer and bioassay techniques. When soils were amended with BR at 0.6, 1.2 and 1.8 nmol g −1, dose-dependent and significant inhibition was observed for rice seedlings within 14 d after treatment, but no significant inhibition occurred to corn seedlings in the same treatment. Radioactive analysis of soil extracts following sequential extractions showed that the 14C labeled residues of ZJ0273 were released from the amended soil BR upon planting. For example, when amended with 1.8 nmol g −1, about 68.3%, 57.0%, and 61.1%, respectively, of the added BR were released in S 1, S 2, and S 3 planted with rice seedlings, whereas 38.9%, 32.7% and 32.6% became available for uptake in the corresponding soils planted with corn seedlings. The released compounds were identified as ZJ0273 and its degradation products M1 and M2, with M2 as the primary component. Bioassay on rice showed that concentration for 50% inhibition (IC 50) of ZJ0273, M1, and M2 were 33.16, 1.93 and 0.49 μM, respectively. Therefore, BR formed after application of ZJ0273 may become available for plant uptake during rice cultivation and lead to phytotoxic effects, and the phytotoxicity is mainly caused by the release of the biologically active metabolite M2. This knowledge is valuable for designing crop rotation practices so that crop injury and yield losses due to carry-over herbicide phytotoxicity may be avoided.

Spatial and temporal changes in acetolactate synthase activity as affected by new herbicide ZJ0273 in rapeseed, barley and water chickweed

Propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy)benzylamino)benzoate (ZJ0273), derived from a precursor compound pesticide (a derivative of 2-pyrimidinyloxy- N-aryl benzoate), is a novel herbicide used in the rapeseed field. This paper studied the differential tolerance or susceptibility of four plant species viz. Brassica napus L. (rapeseed), Hordeum vulgare L. (barley), Malachium aquaticum (L.) Fries (water chickweed), and Alopecurus aequalis Sobol. (equal alopecurus), to various concentrations of ZJ0273, based on the rate of inhibition of acetolactate synthase (ALS) activity. Brassica napus was observed as the most tolerant among four species in which ALS activity in vivo was hardly affected by ZJ0273 at 100 mg/L (recommended dose in the rapeseed field). In contrast, H. vulgare was found to be more susceptible which was reflected in the relative decline of ALS activity and lower biomass production even under at a lower rate of 10 mg/L of ZJ0273. The two weed species exhibited moderate susceptibility as compared to H. vulgare. The present study confirmed that ZJ0273 at 100 mg/L is the optimal dose of herbicide for rapeseed field. Seedling stage was critical in determining the tolerance of crops and susceptibility of weeds which indicated that over seven leaf stage was much safer for rapeseed whereas the two leaf stage juvenile water chickweed was more susceptible to the same rate of ZJ0273. Spatial and temporal variations in ALS activity as affected by 100 mg/L ZJ0273 in water chickweed, rapeseed and barley also suggested that the synchronized sampling based on the tissue position and seedling stage was necessary to examine precisely, the special difference in tolerance or susceptibility to ALS-inhibiting herbicide. The uppermost younger leaf seemed to be the most appropriate sample for evaluation. Furthermore, the variant tendency of ALS activity from different parts of plants between monocotyledons and dicotyledons to ZJ0273 treatments showed differential response which revealed some possible mechanism related to the altered absorption and translocation of the herbicide.

Fate characterization of a novel herbicide ZJ0273 in aerobic soils using multi-position 14C labeling

ZJ0273, propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy)benzylamino)benzoate, is a novel herbicide being developed for use on rape crops. The environmental behavior and fate of ZJ0273, however, are not well understood. In this study, multi-position 14C-labeled compounds (B-ZJ0273 and C-ZJ0273) were used to investigate the fate of ZJ0273 in aerobic soils, and compare their differences in extractable residues (ER), bound residues (BR), and mineralization. ER for both 14C labels followed a similar dissipation trend in the same soil, and results showed a strong effect of soil pH, with the ER level being significantly higher in acidic soils than neutral or alkaline soils. The disappearance of parent compound followed first-order dynamics for both labels in the same soils. No difference in BR was observed between the two 14C-labels. BR increased with time and reached 17.5–35.3% of applied 14C in the three soils after 100-d incubation, which is much smaller than the non-accumulative criteria (70%) as stated in the directives by the Commission of the European Communities. Mineralization to CO 2 also depended closely on the soil type, with the cumulative mineralized fraction being substantially greater in the neutral and alkaline soils than in the acidic soil. Results also showed that cumulative mineralization rates of the two labels in the acidic soil were much smaller than those in the neutral and alkaline soils. Mineralization of C-ZJ0273 (1.2–9.9%) was found to be more extensive than that of B-ZJ0273 (0.5–6.6%), suggesting that benzyl ring was more susceptible to cleavage than pyrimidine ring. The use of multi-position labeling proved to be valuable; it served the purpose of validating the rigorousness of experimental protocols, and provided insights into the behavior and fate of the different molecular fragments.

Determination of herbicide ZJ0273 residue in rapeseed by radioisotopic tracing method

Residue of herbicide ZJ0273 (propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy)benzylamino)benzoate) in rapeseed under simulated field conditions was evaluated by radioisotopic tracing method. Radio-HPLC and HPLC–MS analysis revealed that the parent ZJ0273 is the only radioactive residue in rapeseed. The residue of propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy)[ phenyl-U- 14C 6]benzylamino)benzoate (RCZJ) in postharvest rapeseed from the rape seedlings ( Brassica napus L.) in red clayey soil (S 1) and in fluvio marine yellow loamy soil (S 2) were 0.67 and 1.24 nmol/plant, respectively, based on the 14C radioactivity and the specific activity of the labelled ZJ0273. The RCZJs in the rapeseed (1 kg) from the seedlings in S 1 and in S 2 were 40.13 and 45.33 nmol, respectively, and the amounts of ZJ0273 in the rapeseed (1 kg) were 16.97 and 19.17 μg. The results suggested that only a trace level of residue of ZJ0273 presents in rapeseed due to the poor root absorption of ZJ0273 and the weak translocation of ZJ0273 from rape leaves.

Effects of 5-Aminolevulinic Acid on Oilseed Rape Seedling Growth under Herbicide Toxicity Stress

Weeds are one of the major constraints in oilseed Brassica production. Use of effective herbicides to control weeds in the fields is one of the major objectives of agronomists. To improve weed control efficacy and minimize the application costs, complex combinations of 5-aminolevulinic acid (ALA) and a new postemergence herbicide, propyl 4-(2-(4,6-dimethoxypyrimidin-2-yloxy)benzylamino)benzoate (ZJ0273), were used to investigate their combined effects in relation to seedling growth and development of oilseed rape (Brassica napus cv. ZS 758). Brassica seeds were treated with different concentrations of ZJ0273 [100 (normal dose for rape), 200, 500, and 1000 mg/L] and ALA (0.1, 1, 10, and 50 mg/L). ALA was applied as pre- and post-treatment alone and in combination with ZJ0273. We found that ZJ0273 stress imposed negative effects on rape seedling growth. Shoot fresh weight, shoot length, and root fresh weight were inhibited significantly under ZJ0273 stress, and the rate of decline increased consistently with increased ZJ0273 concentration. Root oxidizability was also inhibited significantly under ZJ0273 stress conditions, and the higher the concentration of the herbicide ZJ0273, the lower the oxidizability. Herbicide ZJ0273 treatment produced a gradual decrease in antioxidant enzymes (peroxidase, superoxide dismutase, and ascorbate peroxidase) and an increase in peroxidation substance (malondialdehyde accumulation). The increase and decrease were consistent with the ZJ0273 dosage. Our results indicated that pre- and post-treatments with a lower dosage of ALA (1 mg/L) improved rape seedling growth and root oxidizability parameters, whereas a higher concentration of ALA (50 mg/L) depressed growth. We also found that plants treated with 1 mg/L ALA produced the highest shoot fresh weights, shoot lengths, root fresh weights, and root oxidizability when the seeds were treated with different concentrations of ZJ0273. Lower dosages of ALA improved the activities of antioxidant enzymes, whereas the highest dosage of ALA increased the accumulation of peroxidation substance. These results indicate that ALA has promotive effects in the recovery of growth and development of rape seedlings under herbicide ZJ0273 toxicity stress.