Improvement of multicatalytic properties of nitrilase from Paraburkholderia graminis for efficient biosynthesis of 2-chloronicotinic acid.

Nitrilases are promising biocatalysts to produce high-value-added carboxylic acids through hydrolysis of nitriles. However, since the enzymes always show low activity and sometimes with poor reaction specificity toward 2-chloronicotinonitrile (2-CN), very few robust nitrilases have been reported for efficient production of 2-chloronicotinic acid (2-CA) from 2-CN. Herein, a nitrilase from Paraburkholderia graminis (PgNIT) was engineered to improve its catalytic properties. We identified the beneficial residues via computational analysis and constructed the mutant library. The positive mutants were obtained and the activity of the "best" mutant F164G/I130L/N167Y/A55S/Q260C/T133I/R199Q toward 2-CN was increased from 0.14 × 10-3  to 4.22 U/mg. Its reaction specificity was improved with elimination of hydration activity. Molecular docking and molecular dynamics simulation revealed that the conformational flexibility, the nucleophilic attack distance, as well as the interaction forces between the enzyme and substrate were the main reason alternating the catalytic properties of PgNIT. With the best mutant as biocatalyst, 150 g/L 2-CN was completely converted, resulting in 2-CA accumulated to 169.7 g/L. When the substrate concentration was increased to 200 g/L, 203.1 g/L 2-CA was obtained with yield of 85.7%. The results laid the foundation for industrial production of 2-CA with the nitrilase-catalyzed route.

[Chemical Constituents from Dalbergia cochinchinensis].

OBJECTIVE: To investigate the chemical constituents from the heartwood of Dalbergia cochinchinensis. METHODS: Isolate and purify compounds by various column chromatographic methods. Spectral analysis were taken to identify the structures. RESULTS: Elev- en compounds were isolated and identified as dibutyl terephthalate (1), medicarpin (2), pterostilbene (3), 6-hydroxy-2-(2-hydroxy-4- methoxyphenyl)-benzofuran (4), pterocarpol (5), butyl isobutyl phthalate (6), pterolinus B (7), methyl 4-hydroxybenzoate (8), ethyl 4- hydroxybenzoate (9),2-(2'-methoxy-4'-hydroxy)-aryl-3-methyl-6-hydroxy-benzofuran (10) and 6α-hydroxycyclonerolidol (11). CONCLUSION: Compounds 1 and 6~10 are isolated from Dalbergia genus for the first time, and compounds 2, 4 and 11 are isolated from this plant for the first time.

[Phenylpropanoids constituents of Eucommia ulmoides leaves].

OBJECTIVE: To investigate phenylpropanoids constituents from Eucommia ulmoides leaves. METHODS: Various column chromatographic methods were used in isolation and purification. Physiochemical constant determination and spectral analysis were adopted to determine the chemical structures of phenylpropanoids. RESULTS: Nine phenylpropanoids were isolated and identified as caffeic acid (1), chlorogenic acid methylester (2), syringin (3), guaiacylglycerol (4), 5-methoxy-guaiacylglycerol (5), 5,9-dimethoxy-guaiacylglycerol (6), 9-n-butyl-guaiacylglycerol (7), 9-n-butyl-isoguaiacylglycerol (8), 8'-methoxy-olivil (9). CONCLUSION: Compounds 5 - 9 are isolated from this plant for the first time.

[Study on chemical constituents of iridoids from eucommiae folium].

OBJECTIVE: To investigate the chemical constituents of iridoids from Eucommiae Folium. METHODS: The compounds were isolated and purified by various column chromatography, the structures were identified on the basis of physiochemical properties and spectral analysis. RESULTS: Nine iridoids were isolated and identified as asperuloside (1), daphylloside (2), scandoside methyl ester (3), loganin (4), 8-epi-loganin (5), 7-epi-loganin (6), deacetyl asperulosidic acid methyl ester (7), geniposide (8) and geniposidic acid (9). CONCLUSION: Compounds 2 - 7 are isolated from this plant for the first time, compounds 4 - 6 are isolated from this genus for the first time.

[Study on chemical constituents in the roots and rhizome of Trillium tschonoskii (III)].

OBJECTIVE: To investigate the chemical constituents in the roots and rhizome of Trillium tschonoskii. METHODS: The roots and rhizome of Trillium tschonoskii were extracted with 70% ethanol and purified by polyamide, silica gel, RP-C18 and Sephadex LH-20 column chromatography. Chemical structures were identified by MS, 1D and 2D NMR experiments. RESULTS: Twelve compounds were isolated and identified from the ethyl acetate and n-butanol as 7,11-dimethyl-3-methylene-10, 11-dihdroxyl-1,6-dodecadien-diol(1),2, 6,10-trimethyl-2,10-dihdroxyl-6-dodecene-diol(2),3,7,11-trimethyl-3,9,11-trihydroxyl-1,6-dodecadiene-glycerol(3),p-hydroxymethyl-benzyl alcohol(4),20betaF, 22alphaF, 25alphaF-spirostan-5,13-ene-3beta,21alpha-diol(5), 2-methyl-3,4-dihydroxy-hexanedioic acid(6), alpha-Lrhamnopyranosyl(7), epitrillenogenin-24-O-acetate-1-O-[2,3,4,-tri-O-acetyl-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside] (8),epitrillenogenin-24-O-acetate-1-O-[2,4, -di-O-acetyl-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside](9), epitrillenogenin1-O-[2,3,4,-tri-O-acetyl-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside] (10), epitrillenogenin-1-O-[2,4, -di-O-acetyl-alpha-Lrhamnopyranosyl-(1-->2) -alpha-L-arabinopyranoside] (11) and epitrillenogenin-1-O-[4-O-acetyl-alpha-L-rhamnopyranosyl-(1-->2) -alpha-L-arabinopyranoside] (12). CONCLUSION: Compounds 4-6 are isolated from this genus for the first time, compounds 1-3, 11, 12 are isolated from this plant for the first time.

[Effects of biomass conditioner on soil nutrient and microbial community characteristics of alpine desertified grassland in northwest Sichuan, China]. / ç”Ÿç‰©è´¨è°ƒç†å‰‚å¯¹å·è¥¿åŒ—é«˜å¯’è‰åœ°æ²™åŒ–åœŸå£¤å »åˆ†å’Œå¾®ç”Ÿç‰©ç¾¤è½ç‰¹å¾çš„å½±å“.

Biomass conditioner made from agricultural and animal husbandry waste for resource disposal could be used to improve desertified soil, which is one of the effective ways of ecological management on desertified grasslands in northwest Sichuan. To clarify the effects of different raw material conditioners on alpine desertified grassland in northwest Sichuan, we analyzed the effects of three conditioners on soil nutrients and microbial community characteristics. With no conditioner as the control (CK), three different biomass conditioners were set up with an application rate of 12 t·hm-2, including mushroom dregs (JZ), straw (JG) and biochar (SWT). The results showed that all biomass conditioners could significantly increase soil available nutrients and active organic carbon by 23.0%-521.6%. Among the three conditioners, JG had the best effect, with an improvement range for soil nutrient and organic carbon of 65.1%-521.6%. Because biomass conditioner was only applied in the first year, soil available nutrients and active organic carbon in the second year decreased by 4.5%-92.3% compared with that of the first year, while soil organic carbon and microbial biomass carbon content of the second year increased by 5.6%-458.0%. The biomass conditioners changed the relative abundance of the dominant bacteria in the microbial community. JG significantly affected bacterial flora, while JG and JZ affected fungal flora. Compared with CK, JG significantly reduced the diversity of soil bacteria and fungi. The Shannon index was decreased by 2.9% and 31.8%, while the Simpson index was increased by 175.0% and 320.9%, respectively. Results of the redundancy analysis showed that the contents of soil available nutrients and active organic carbon were important factors affecting microbial community composition. The contents of soil nitrate and microbial biomass carbon had greater impacts on bacterial community composition, explaining 65.9% of community variations. The contents of soil available potassium and microbial biomass carbon had a greater impact on fungal community composition, explaining 83.2% of community variation. According to the comprehensive comparison, straw conditioner could significantly increase soil available nutrients and active organic carbon, and benefit the growth of beneficial bacteria and fungi, which could be used as a promotion measure to improve soil quality of alpine desertified grassland in northwest Sichuan.

[Effects of short-term deep vertically rotary tillage on topsoil structure of lime concretion black soil and wheat growth in Huang-Huai-Hai Plain, China]. / 短期深旋松对黄淮海沙姜黑土耕层结构及小麦生长的影响.

Annual rotary tillage can often create a compacted plough pan and shallow arable layer which hampers the high crop yield in Huang-Huai-Hai region. A brand new farming method named Vertically Rotary Tillage was introduced to solve this problem. One short-term field experiment was conducted to explore the effect of deep vertically rotary tillage on soil physical properties and photosynthetic characteristics at flowering stage of winter wheat. Two tillage treatments were designed including subsoiling tillage with 20 cm depth (SS20, CK) and deep vertically rotary tillage with 30 cm depth (DVR30). The result showed that compared with SS20 treatment, DVR30 treatment could thoroughly break the plow pan and loose the arable layer. The soil bulk density at 10-20 cm and 20-30 cm layers under DVR30 treatment was decreased by 9.5% and 11.2% respectively than that under SS20 treatment. Meanwhile, the penetration resistance at 20-30 cm layer under DVR30 treatment was also decreased by 42.3% than that under SS20 treatment. Moreover, water infiltration under DVR30 treatment and the soil water storage in the deep soil layers was then increased. The mass water content of soil increased significantly with the increase of soil depth. There was significant difference of mass water content of 30-40 cm 40-50 cm between SS20 and DVR30. The mass water content 30-40 cm and 40-50 cm layers under DVR30 treatment was increased by 16.9% and 10.6% compared with SS20 treatment, respectively. Furthermore, DVR30 treatment promoted the improvement of the photosynthetic capacity of wheat which could contribute to the dry matter accumulation of winter wheat. The net photosynthesis rate and SPAD at flowering stage of winter wheat leaves under DVR30 treatment were increased by 1.3% and 15.5% respectively than that under SS20 treatment, thereby the above and underground dry matter accumulation of winter wheat under DVR30 was increased significantly. Due to all the superiority of DVR30 treatment over SS20 treatment showed above, the winter wheat yield under DVR30 treatment was increased by 12.4% than that under SS20. It was concluded that deep vertically rotary tillage could provide a new and effective way to break up the compacted plough pan, build a reasonable soil structure and increase crop yield.

[Effects of different irrigation modes in winter wheat growth season on the grain yield and water use efficiency of winter wheat-summer maize].

Three irrigation modes in winter wheat growth season were carried out in Heilonggang basin of North China Plain to investigate their effects on the grain yield, water consumption, and water use efficiency (WUE) of winter wheat-summer maize. The three irrigation modes included irrigation before sowing (75 mm, W1), irrigation before sowing and at jointing stage (75 mm + 90 mm, W2), and irrigation before sowing, at jointing stage, and at filling stage (75 mm + 90 mm + 60 mm, W3). With the irrigation modes W2 and W3, the increment of the annual yield of winter wheat-summer maize was 8.7% and 12.5% higher than that with W1, respectively. The water consumption in winter wheat growth season decreased with increasing irrigation amount, while that in summer maize growth season increased with the increasing irrigation amount in winter wheat growth season. The WUE of winter wheat with the irrigation mode W2 was 11.1% higher than that with W3, but the WUE of summer maize had less difference between irrigation modes W2 and W3. The annual WUE (WUE(T)) of W2 and W1 was 21.28 and 21.60 kg(-1) x mm x hm(-2), being 7.8% and 9.4% higher than that of W3, respectively. Considering the annual yield, water consumption, and WUE, irrigation mode W2 could be the advisable mode for water-saving and high-yielding.