BioReagent 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
2-8°C储存,避光,室温。低温运输 。请查阅批次 COA 获取详细规格。
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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。
植物体内的碳素营养状况以及农产品的品质形状，常以糖含量作为重要指标，单糖和某些寡糖（如麦芽糖）含有游离的醛基或酮基，具有还原性，属于还原糖；多糖和蔗糖等属于非还原糖，可以利用非还原糖能被酸水解为单糖的特性通过测定水解后的单糖含量对总糖进行测定。
检测原理：还原糖在碱性加热条件下被氧化成糖酸，3,5-二硝基水杨酸被还原为棕红色的氨基化合物，在一定范围内还原糖的量与棕红色产物的颜色深浅程度呈一定比例关系，在540nm处用酶标仪测定棕红色物质的吸光度，该吸光度值与还原糖含量呈线性关系，利用标准曲线计算样品中的还原糖和总糖的含量。该试剂盒仅用于科研领域，不适用于临床诊断或其他用途。
| P1501777 | Component | 100T | 300T | Storage |
| P1501777A | Glu标准 (1mg/mL) | 1 mL | 1 mL | 2-8℃ |
| P1501777B | DNS检测液 | 10 mL | 30 mL | RT. Store in the dark. |
| P1501777C | 显色液 (总糖使用) | 5 mL | 10 mL | RT. Store in the dark. |
自备仪器和试剂
1、蒸馏水、盐酸溶液、氢氧化钠溶液
2、50mL离心管、1mL离心管、离心机、水浴锅或恒温箱、酶标仪、96 孔板、水浴锅
实验流程
1、还原糖的提取
1.1 称取植物样品0.5~3g，剪碎，加入蒸馏水约3mL匀浆，转移至烧杯或三角瓶中，用12mL蒸馏水冲洗研磨器2~3次，洗出液也转移至该容器。
1.2 50℃水浴30min，并不时搅拌，以便还原糖彻底浸出。
1.3 将沉淀和浸出液转移至50mL离心管，4000g离心5min。
1.4 留取上清液，向沉淀中加入20mL蒸馏水，混匀，再次4000g离心5min。
1.5 留取上清液，将2次获得的上清液合并，用蒸馏水定容至100mL（提取液），混匀，作为还原糖待测液。
2、总糖的水解和提取
2.1 称取植物样品0.5~3g，剪碎，加入蒸馏水约3mL匀浆，转移至烧杯或三角瓶中，用12mL蒸馏水冲洗研磨器2~3次，洗出液也转移至该容器。
2.2 向容器中加入10mL 6M盐酸溶液，搅拌均匀，然后置于沸水浴中加热水解30min，并不时搅拌。
2.3 取2滴滴加于载玻片上，滴加1滴显色液（约50μL），检查水解是否完全，如已经水解完全，则不显示蓝色。
2.4 水解完毕后，冷却至室温，加入6M 氢氧化钠溶液，使溶液pH至7.4，用蒸馏水定容至100mL，混匀，4000g离心5min或过滤。
2.5 取上清或滤液10mL，用蒸馏水定容至100mL，成稀释10倍的总糖水解液（提取液），取50μL总糖水解液，测定其还原糖的含量。
3、葡萄糖标准品配制
取干净离心管或试管，按下表操作，依次获得系列浓度的Glu标准。
| 标准品工作液 | Glu标准 (1mg/mL)（mL） | 蒸馏水 （mL） | 浓度 （mg/mL） |
| 1 | 0.01 | 0.04 | 0.2 |
| 2 | 0.02 | 0.03 | 0.4 |
| 3 | 0.03 | 0.02 | 0.6 |
| 4 | 0.04 | 0.01 | 0.8 |
| 5 | 0.05 | 0 | 1.0 |
4、加样
取1mL离心管，按照下表设置空白孔、标准孔、测定孔，溶液应按照顺序依次加入，并注意避免产生气泡，小心混匀。如果样品中的糖浓度过高，可以减少样品用量或适当稀释后再进行测定，样品的检测最好能设置2~3平行孔，求平均值。
| 试剂（μL） | 空白孔 | 标准孔 | 测定孔 |
| 蒸馏水 | 50 | / | / |
| Glu标准品（1-5） | / | 50 | / |
| 提取液 | / | / | 50 |
| DNS检测液 | 100 | 100 | 100 |
沸水浴中准确煮沸5min，取出，自来水冷却至室温，补加蒸馏水250μL。
5、还原糖测定
混匀，依次抽取300uL转移至相应96孔板中，以空白孔调零测定540nm处标准孔、测定孔的吸光度。
6、结果计算
6.1 标准曲线的绘制
以Glu标准品（1-5），即标准葡萄糖浓度（mg/mL）为横坐标，以相应的吸光度为纵坐标，作图得标准曲线，根据提取液得吸光度在标准曲线上查出相应得葡萄糖浓度。
6.2 含量计算
还原糖的百分含量：
100g样品中还原糖含量（g）=(c×V T )/(m×1000)×100= (c×V T )/(m×10)
总糖的百分含量：
100g样品中总糖含量（g）=(c×N×V T )/(m×1000)×100×0.9=(c×N×V T )/(m×10)×0.9
参数说明
c：从标准曲线查的糖量（mg/mL）
V T ：提取液的总体积，100ml
m：植物样品的质量，g
N：总糖水解液的稀释倍数，10
7、试剂开封后请尽快使用， 以防影响后续实验效果。
The carbon nutritional status in plants and the quality characteristics of agricultural products are often evaluated using sugar content as an important indicator. Monosaccharides and some oligosaccharides (such as maltose) contain free aldehyde or ketone groups, possess reducibility, and are classified as reducing sugars. Polysaccharides and sucrose are non-reducing sugars. The total sugar content can be determined by measuring the monosaccharide content after hydrolysis, utilizing the property that non-reducing sugars can be hydrolyzed to monosaccharides by acid.
Detection Principle: Reducing sugars are oxidized to sugar acids under alkaline heating conditions, while 3,5-dinitrosalicylic acid (DNS) is reduced to a brownish-red amino compound. Within a certain range, the amount of reducing sugar is proportional to the color intensity of the brownish-red product. The absorbance of this brownish-red substance is measured at 540 nm using a microplate reader. This absorbance value has a linear relationship with the reducing sugar content. The reducing sugar and total sugar content in the sample are calculated using a standard curve.
This kit is for scientific research use only and is not intended for clinical diagnosis or other purposes.
| P1501777 | Component | 100T | 300T | Storage |
| P1501777A | Glu Standard (1 mg/mL) | 1 mL | 1 mL | 2-8℃ |
| P1501777B | DNS Detection Solution | 10 mL | 30 mL | RT. Store in the dark. |
| P1501777C | Color Solution (for Total Sugar) | 5 mL | 10 mL | RT. Store in the dark. |
User-Prepared Instruments and Reagents
1. Distilled water, Hydrochloric acid solution, Sodium hydroxide solution
2. 50 mL centrifuge tubes, 1 mL centrifuge tubes, Centrifuge, Water bath or incubator, Microplate reader, 96-well plate, Water bath
Experimental Procedure
1. Extraction of Reducing Sugars
1.1 Weigh 0.5-3 g of plant sample, cut into pieces, add about 3 mL of distilled water and homogenize. Transfer to a beaker or conical flask. Rinse the grinder 2-3 times with 12 mL of distilled water and transfer the rinsate to the same container.
1.2 Incubate in a 50°C water bath for 30 min, stirring occasionally to ensure thorough extraction of reducing sugars.
1.3 Transfer the precipitate and extract to a 50 mL centrifuge tube. Centrifuge at 4000 g for 5 min.
1.4 Collect the supernatant. Add 20 mL of distilled water to the precipitate, mix well, and centrifuge again at 4000 g for 5 min.
1.5 Collect the supernatant. Combine the supernatants from the two steps. Dilute to 100 mL with distilled water (this is the extract). Mix well. This serves as the test solution for reducing sugars.
2. Hydrolysis and Extraction of Total Sugars
2.1 Weigh 0.5-3 g of plant sample, cut into pieces, add about 3 mL of distilled water and homogenize. Transfer to a beaker or conical flask. Rinse the grinder 2-3 times with 12 mL of distilled water and transfer the rinsate to the same container.
2.2 Add 10 mL of 6 M hydrochloric acid solution to the container, mix well, then heat in a boiling water bath for 30 min for hydrolysis, stirring occasionally.
2.3 Take 2 drops and place on a glass slide, add 1 drop of Color Solution (about 50 µL) to check if hydrolysis is complete. If hydrolysis is complete, no blue color should develop.
2.4 After hydrolysis, cool to room temperature. Add 6 M sodium hydroxide solution to adjust the pH to 7.4. Dilute to 100 mL with distilled water, mix well. Centrifuge at 4000 g for 5 min or filter.
2.5 Take 10 mL of the supernatant or filtrate and dilute to 100 mL with distilled water, creating a 10-fold diluted total sugar hydrolysate (extract). Take 50 µL of this total sugar hydrolysate to measure its reducing sugar content.
3. Glucose Standard Preparation
Take clean centrifuge tubes or test tubes and prepare a series of Glu standards according to the table below.
| Standard Working Solution | Glu Standard (1 mg/mL) (mL) | Distilled Water (mL) | Concentration (mg/mL) |
| 1 | 0.01 | 0.04 | 0.2 |
| 2 | 0.02 | 0.03 | 0.4 |
| 3 | 0.03 | 0.02 | 0.6 |
| 4 | 0.04 | 0.01 | 0.8 |
| 5 | 0.05 | 0 | 1.0 |
4. Assay Setup
Take 1 mL centrifuge tubes. Set up Blank, Standard, and Test wells according to the table below. Add solutions sequentially, avoiding bubbles. Mix carefully. If the sugar concentration in the sample is too high, reduce the sample volume or dilute appropriately before assay. It is best to set up 2-3 replicate wells for samples and take the average.
| Reagent (µL) | Blank Well | Standard Well | Test Well |
| Distilled Water | 50 | / | / |
| Glu Standard (1-5) | / | 50 | / |
| Extract | / | / | 50 |
| DNS Detection Solution | 100 | 100 | 100 |
Heat accurately in a boiling water bath for 5 min. Remove, cool to room temperature with tap water. Add 250 µL distilled water.
5. Reducing Sugar Measurement
Mix well. Transfer 300 µL sequentially to the corresponding wells of a 96-well plate. Measure the absorbance of Standard and Test wells at 540 nm, using the Blank well to zero the instrument.
6. Result Calculation
6.1 Standard Curve Plotting
Using the Glu standards (1-5), i.e., the standard glucose concentrations (mg/mL) as the x-axis and the corresponding absorbance values as the y-axis, plot the standard curve. Find the corresponding glucose concentration on the standard curve based on the absorbance of the extract.
6.2 Content Calculation
Percentage Content of Reducing Sugars:
Reducing sugar content per 100 g sample (g) = (c × V T ) / (m × 1000) × 100 = (c × V T ) / (m × 10)
Percentage Content of Total Sugars:
Total sugar content per 100 g sample (g) = (c × N × V T ) / (m × 1000) × 100 × 0.9 = (c × N × V T ) / (m × 10) × 0.9
Parameter Description
c: Sugar amount found from the standard curve (mg/mL)
V T : Total volume of the extract, 100 mL
m: Mass of the plant sample, g
N: Dilution factor of the total sugar hydrolysate, 10
7. Use reagents promptly after opening to avoid affecting subsequent experimental results.
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| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 I2614533 |
分析证书 | P1501777 |
| H2631425 |
分析证书 | P1501777 |
| H2621212 |
分析证书 | P1501777 |
| H2620245 |
分析证书 | P1501777 |
| ZJ26F0838403 |
分析证书 | P1501777 |
| G2630347 |
分析证书 | P1501777 |
| G2624347 |
分析证书 | P1501777 |
| F2624401 |
分析证书 | P1501777 |
| E2628509 |
分析证书 | P1501777 |
| E2627310 |
分析证书 | P1501777 |
| ZJ26F0535518 |
分析证书 | P1501777 |
| ZJ26F0535517 |
分析证书 | P1501777 |
| D2621590 |
分析证书 | P1501777 |
| C2630218 |
分析证书 | P1501777 |
| A2621164 |
分析证书 | P1501777 |
| L2522112 |
分析证书 | P1501777 |
| K2518545 |
分析证书 | P1501777 |
| ZJ25F1129378 |
分析证书 | P1501777 |
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