BioReagent 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
避光,-20°C储存。超低温运输 。请查阅批次 COA 获取详细规格。
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
β-GC（EC 3.2.1.21）广泛存在于动物、植物、微生物和培养细胞中，催化 β-糖苷键水解，具有多方面生理作用：在纤维素的糖化作用中，β-GC 负责进一步水解纤维素二糖和纤维素寡糖生成葡萄糖；β-GC 水解萜烯类香气前驱体，使糖苷键合态变成游离态。从而产生香味；β-GC 能够水解植物体内野黑樱苷，释放 HCN，从而防止昆虫取食。
测定原理：
β-GC 分解对-硝基苯-β-D-吡喃葡萄糖苷生成对-硝基苯酚，后者在 400nm 有最大吸收峰，通过测定吸光值升高速率来计算 β-GC 活性。
|
注：
① 试剂一：粉剂×2瓶，-20℃ 保存；临用前每瓶加入 6mL 蒸馏水，充分溶解备用；用不完的试剂分装为合适用量 -20℃ 避光保存，4 周有效，避免反复冻融。
② 标准品（选做）：5mmol/L 对-硝基苯酚，可用于自行绘制标准曲线。
自备仪器及试剂
可见分光光度计、台式离心机、水浴锅、可调式移液器、1mL 玻璃比色皿、研钵、冰和蒸馏水。
样本处理：
1、细菌或培养细胞：先收集细菌或细胞到离心管内，离心后弃上清；按照细菌或细胞数量 (10⁴ 个)：提取液体积 (mL) 为 500~1000: 1的比例（建议 500 万细菌或细胞加入 1mL 提取液），超声波破碎细菌或细胞（冰浴，功率 20％ 或 200W，超声 3s，间隔 10s，重复 30 次）；15000g 4℃ 离心 10min，取上清，置冰上待测。
2、组织：按照组织质量 (g)：提取液体积 (mL) 为1：5~10 的比例（建议称取约 0.1g 组织，加入 1mL 提取液），进行冰浴匀浆。15000g 4℃ 离心 10min，取上清，置冰上待测。
3、培养液、血清 (浆) 等液体样本：直接检测。
测定步骤：
1、分光光度计预热 30min 以上，调节波长至 400nm，蒸馏水调零。
|
注：
① 加入试剂二后迅速混匀，放入 37℃ 准确水浴 30min 后，立即放入 95℃ 水浴 5min (盖紧，以防止水分散失)，流水冷却后充分混匀 (以保证浓度不变)，8000g，4℃，离心 5min，取上清液；
② 加入试剂三后充分混匀，室温静置 2min 后，取 1mL 于400nm 处测定吸光值 A，计算 ΔA=A测定管-A对照管。每个测定管需设一个对照管。
酶活性计算公式：
配制不同浓度梯度对-硝基苯酚，取500μL加入1000μL试剂三混匀后检测得到的，回归方程为y=0.006387x-0.006024，R2=0.9994；x为标准品浓度 (nmol/mL), y为吸光值。
(1) 按液体体积计算
酶活定义：每 mL 样本每分钟产生 1nmol 对 - 硝基苯酚定义为一个酶活性单位。
Cpr：样本蛋白质浓度，mg/mL
W：样本质量，g
N：细胞或细菌总数，以 10⁴ 计
T：反应时间，30min
注意事项：
1.ΔA测定检测范围为0.02~1.5，如果ΔA测定小于 0.02，可延长37℃反应时间，如果ΔA测定大于1.5可将样本用提取液稀释后测定，注意同步修改计算公式。
2.提供的标准曲线及公式仅供参考，若有需要，可用提供的标准品自制标曲（计算公式及线性范围根据自制标准曲线需作对应修改），但需要注意，受仪器及个人操作影响，客户自制标准曲线可能与说明书中提供的标准曲线有差别。若无另外的实验要求，也可直接使用说明书所给标曲进行计算。
实验实例：
|
2、β-葡萄糖苷酶酶活检测：标记酶活≥30U/mg，使用提取液溶解不同浓度酶液，检测结果如下：
|
|
|
β-GC (EC 3.2.1.21) is widely distributed in animals, plants, microorganisms and cultured cells. It catalyzes the hydrolysis of β-glycosidic bonds and exerts diverse physiological functions: During cellulose saccharification, β-GC further hydrolyzes cellobiose and cellooligosaccharides to produce glucose; β-GC hydrolyzes terpenoid aroma precursors to convert bound glycosidic fragrances into free aromatic substances, thus releasing characteristic flavor; β-GC can decompose prunasin in plants to release hydrogen cyanide (HCN), deterring insect feeding.
Assay Principle：
β-GC catalyzes the decomposition of p-nitrophenyl-β-D-glucopyranoside to generate p-nitrophenol, which has a maximum absorption peak at 400 nm. The activity of β-GC is calculated by measuring the rising rate of absorbance value.
|
Notes：
① Reagent 1, Powder×2 bottles, store at -20℃. Before use, add 6 mL distilled water to each bottle and dissolve completely for later use. Divide unused reagent into appropriate aliquots, store at -20℃ away from light, valid for 4 weeks. Avoid repeated freeze-thaw cycles.
② Standard (optional): 5 mmol/L p-nitrophenol, available for self-prepared standard curve plotting.
Instruments and Reagents to Be Prepared by Users
Visible spectrophotometer, tabletop centrifuge, water bath, adjustable pipettes, 1 mL glass cuvettes, mortar, ice, distilled water.
Sample Pretreatment：
1. Bacteria / Cultured Cells
Collect bacteria or cells into centrifuge tubes and discard supernatant after centrifugation. Mix with extraction solution at the ratio of 500–1000 ×10⁴cells : 1 mL extraction solution (recommended: 5 million cells per 1 mL extraction solution). Lyse cells by sonication on ice: power 20% / 200 W, sonicate for 3 s, pause 10 s, repeat 30 times. Centrifuge at 15000 g, 4℃ for 10 min, transfer supernatant and keep on ice for testing.
2. Tissue
Homogenize tissue on ice at the mass-to-volume ratio of 1 g tissue : 5–10 mL extraction solution (recommended: approx. 0.1 g tissue with 1 mL extraction solution). Centrifuge at 15000 g, 4℃ for 10 min, transfer supernatant and keep on ice for testing.
3. Liquid samples (culture medium, serum, plasma, etc.): Test directly.
Assay Procedures：
1. Preheat spectrophotometer for no less than 30 min, set wavelength to 400 nm, and zero with distilled water.
|
Notes
① Mix thoroughly immediately after adding Reagent 2. Incubate accurately in a 37 °C water bath for 30 min, then transfer to a 95 °C water bath for 5 min at once (keep the tube tightly capped to prevent water evaporation). Cool the mixture under running tap water and mix well to maintain constant concentration. Centrifuge at 8000 g and 4 °C for 5 min, then collect the supernatant.
② Mix thoroughly after adding Reagent 3, and let the mixture stand at room temperature for 2 min. Transfer 1 mL of the solution and measure the absorbance value A at 400 nm. Calculate ΔA = Atest − Acontrol. One control tube shall be prepared for each test tube.
Enzyme Activity Calculation Formula
Prepare serial diluted concentrations of p-nitrophenol. Take 500 μL standard solution and mix with 1000 μL Reagent 3 for detection. The linear regression equation is y = 0.006387x − 0.006024, R² = 0.9994.；x = concentration of standard substance (nmol/mL)；y = absorbance value
(1)Calculation based on liquid volume
Enzyme activity definition: One enzyme activity unit is defined as the amount of enzyme that produces 1 nmol p-nitrophenol per minute per mL sample.
Precautions
1. The valid detection range of measured ΔA is 0.02–1.5. If the measured ΔA is lower than 0.02, extend the reaction time at 37°C. If the measured ΔA exceeds 1.5, dilute the sample with extraction solution before testing, and remember to modify the calculation formula accordingly.
2. The attached standard curve and formula are for reference only. If necessary, users may generate a self-made standard curve using the supplied standard substance (the calculation formula and linear range shall be adjusted correspondingly based on the self-calibrated curve). Please note that due to differences in instruments and manual operation, customer-generated standard curves may deviate from the one listed in this manual. If no additional experimental requirements are specified, the standard curve provided in the instructions can be directly adopted for calculation.
Experimental Example
|
2. Activity Determination of β-Glucosidase: The labeled specific activity is ≥30 U/mg. The enzyme powder was dissolved with extraction solution to prepare enzyme solutions of different concentrations for testing, and the detection results are as follows:
|
Frequently Asked Questions and Solutions：
1、Poor repeatability of sample readings
|
2、Low or abnormally high readings of samples and standards
|
| 组件 | 名称 | 物理外观 | 50T | 储存 |
| G1521822A | 提取液 | 液体 | 50mL | 2-8℃ |
| G1521822B | 试剂一 | 粉剂 | 2瓶 | -20℃, 避光 |
| G1521822C | 试剂二 | 液体 | 25mL | 2-8℃ |
| G1521822D | 试剂三 | 液体 | 50mL | 2-8℃ |
| G1521822E | 标准品 | 液体 | 1mL | 2-8℃, 避光 |
通过匹配包装上的批号来查找并下载产品的 COA，每批产品都进行了严格的验证，您可放心使用！
| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0939374 |
分析证书 | G1521822 |
危险品化学品经营许可证（带存储）