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- 保存条件：
保存在2-8℃，6个月。
- 保质期：
保存在2-8℃，6个月。
- 英文名：
Soil Alkaline Phosphatase(S-AKP/ALP)Activity Assay Kit
- 库存：
99
- 供应商：
北京索莱宝科技有限公司
- 规格：
50管/48样
测定意义：土壤磷酸酶是一类催化土壤有机磷化合物矿化的酶，其活性的高低直接影响着土壤中有机磷的分解转化及其生物有效性，是评价土壤磷素生物转化方向与强度的指标。土壤磷酸酶受到土壤碳、氮含量、有效磷含量和pH显著影响。通常按照其最适pH范围，分为碱性、中性和酸性三种类型磷酸酶。
测定原理：碱性环境中，S-AKP/ALP催化磷酸苯二钠水解生成苯-酚和磷酸氢二钠，通过测定酚的生成量即可计算出S-AKP/ALP活性。
溶液的配制：
1. 试剂二：临用前加50 mL 蒸馏水充分溶解，2-8℃保存8周。
2. 试剂四：临用前取1支加576 μL 无水乙醇(自备),24 μL 蒸馏水充分溶解。用不完的试剂2-8℃保存2 周(变褐色后不能再使用)
3. 标准品：0.5μmol/mL 苯-酚标准液。
产品说明：
土壤磷酸酶是一类催化土壤有机磷化合物矿化的酶，其活性的高低直接影响着土壤中有机磷的分解转化及其 生物有效性，是评价土壤磷素生物转化方向与强度的指标。土壤磷酸酶受到土壤碳、氮含量、有效磷含量和pH 显 著影响。通常按照其最适pH 范围，分为碱性、中性和酸性三种类型磷酸酶。
碱性环境中， S-AKP/ALP 催化磷酸苯二钠水解生成苯-酚和磷酸氢二钠，通过测定酚的生成量即可计算出S- AKP/ALP 活性。
注意：实验之前建议选择2-3个预期差异大的样本做预实验。
需自备的仪器和用品：
可见分光光度计、1mL玻璃比色皿、台式离心机、水浴锅/恒温培养箱、分析天平、可调式移液器、冰、研钵、 蒸馏水、30-50目筛、乙醇(>98%,AR) 和甲苯(>98%,AR)。
操作步骤：
一 、样本处理(可适当调整待测样本量，具体比例可以参考文献)
1. 新鲜土样自然风干或37℃烘箱风干，过30~50目筛。
2. 称取风干混匀土壤约0.1g, 加入0.05mL甲苯(自备),轻摇15min; 加0.4 mL 试剂一并且摇匀后，置于 37℃水浴锅/恒温培养箱，开始计时，催化反应24h; 到时后迅速加入1mL 试剂二充分混匀，以终止酶催化的反应。 10000rpm 室温离心10min, 取上清液置于冰上待测。
二 、测定步骤
1. 分光光度计预热30min 以上，调节波长到660nm, 蒸馏水调零。
2. 空白管：取1mL 玻璃比色皿，加入50μL 试剂一，100μ L 试剂三，20μL 试剂四，充分混匀，显色后再加蒸馏 水830μL, 混匀后室温静置30min, 于660nm 测定吸光度，记为A 空白管。(空白管只需要测定1-2次)
3. 标准管：取1mL 玻璃比色皿，加入50μL 标准液，100μL 试剂三，20 μL 试剂四，充分混匀，显色后再加蒸馏 水830μL, 混匀后室温静置30min, 于 6 6 0nm 测定吸光度，记为A 标准管。(标准管只需要测定1-2次)
4. 测定管：取1mL 玻璃比色皿，加入 50μ L 上清液，100μL 试剂三，20 μL 试剂四，充分混匀，显色后再加蒸馏 水830μL, 混匀后室温静置30min, 于 6 6 0nm 测定吸光度，记为A 测定管。
三、S-ALP/ALP 活性计算
活性单位定义：37℃中每克土壤每天释放1nmol 酚为1个酶活单位。
S-AKP/ALP 活 性 (U/g 土样)=[C 标准液×(A 测定管-A 空白管)÷(A 标准管-A空白管)]×V 总÷W÷T×1000 =725×(A 测定管-A 空白管)÷(A 标准管-A 空白管)÷W
C 标准液：0.5μmol/mL;V 总：催化体系总体积，1.45mL;W: 土壤样本质量，g;T: 催化反应时间，24h=1d; 1000: 单位换算系数，1μmol=1000nmol。
相关发表文献：
[1]Wang S,Song M,Wang C,Dou X,Wang X,Li X.Mechanisms underlying soil microbial regulation of available phosphorus in a temperate forest exposed to long-term nitrogen addition.Sci Total Environ.2023 Dec 15;904:166403. doi:10.1016/j.scitotenv.2023.166403.Epub 2023 Aug 18.PMID:37597553.
[2]Huang J,Ye J,Gao W,Liu C,Price GW,LiY,Wang Y.Tea biochar-immobilized Ralstonia Bcul-1 increases nitrate nitrogen content and reduces the bioavailability of cadmium and chromium in a fertilized vegetable soil.Sci Total Environ.2023 Mar 25;866:161381.doi:10.1016/j.scitotenv.2022.161381.Epub 2023 Jan 5.PMID:36621509.
[3]NiuT,Xie J,LiJ,Zhang J,Zhang X,MaH,Wang C.Response ofrhizosphere microbial community of Chinese chives under different fertilization treatments.Front Microbiol.2022 Nov 21;13:1031624.doi:10.3389/fmicb.2022.1031624. PMID:36478855;PMCID:PMC9719922.
[4]Chen W,Guo X,Guo Q,Tan X,Wang Z.Long-Term Chili Monoculture Alters Environmental Variables Affecting the Dominant Microbial Community in Rhizosphere Soil.Front Microbiol.2021 Jul 1;12:681953.doi: 10.3389/fmicb.2021.681953.PMID:34276615;PMCID:PMC8281244.
[5] Guan TK,Wang QY,Li JS,Yan HW,Chen QJ,Sun J,Liu CJ,Han YY,Zou YJ,Zhang GQ.Biochar immobilized plant growth-promoting rhizobacteria enhanced the physicochemical properties,agronomic characters and microbial communities during lettuce seedling.Front Microbiol.2023 Jul 5;14:1218205.doi:10.3389/fmicb.2023.1218205. PMID:37476665;PMCID:PMC10354297.
参考文献：
[1]Guan S.Soil Enzyme and Its Research Method [M].Beijing:Science Press,1982.
[2]Belfield A,Goldberg DM.Revised assay for serum phenyl phosphatase activity using 4-amino-antipyrine[J].Enzyme, 1971,12(5):561-573.
[3] Powell MEA,Smith MJH.The Determination of Serum Acid and Alkaline Phosphatase Activity with 4- Aminoantipyrine(A.A.P.)[J].Journal of Clinical Pathology,1954,7:245-248.
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