β-amylase(β-AL) Activity Assay Kit, 50 Assays

The β-Amylase Activity Assay Kit is a spectrophotometric assay designed for the quantitative determination of β-amylase (EC 3.2.1.2) activity in plant or biological samples. β-Amylase catalyzes the hydrolysis of α-1,4-glycosidic linkages at the non-reducing ends of starch to release maltose and other reducing sugars. This kit is based on the colorimetric measurement of reducing sugars, which reduce 3,5-dinitrosalicylic acid to produce a red-brown compound detectable at 540 nm.

The kit is ideal for comparative studies, enzyme kinetics, and screening experiments in plant physiology, agricultural science, and biochemical research.

Key Features

• ✅ Sensitive and Quantitative: Detects β-amylase activity via reducing sugar production using a colorimetric method.
• ✅ Distinction Between α- and β-Amylase: By heat-inactivating α-amylase or β-amylase selectively, the kit allows for specific quantification of each enzyme's activity.
• ✅ High Reproducibility: Standard curve-based analysis ensures consistency across replicates.
• ✅ Comprehensive Components: Includes all necessary reagents and standards for 50 tests.

Kit Components

Required Equipment (User Supplied)

• Spectrophotometer (540 nm)
• Water baths (70°C and 40°C)
• Centrifuge (6000 ×g)
• Mortar/homogenizer
• Transfer pipettes
• 1 mL cuvettes
• Distilled water

Procedure Summary

1. Sample Extraction
   Homogenize 0.1 g sample in 0.8 mL water, extract at room temp (15 min), centrifuge, and dilute to 10 mL for amylase stock solution. Further dilute 1:5 for total amylase measurement.
2. Standard Curve Preparation
   Dilute 10 mg/mL glucose solution to prepare six standards (0.00625–0.2 mg/mL).
3. Reaction Setup
   Perform tests with α-amylase, total amylase, and standards using a combination of heat treatments and reagent additions. Incubate, cool, and read absorbance at 540 nm.
4. Calculation
   • Generate a standard curve (Abs vs. glucose concentration)
   • Calculate α- and total amylase activity
   • Derive β-amylase activity by subtracting α from total activity
   • Express results as U/g fresh weight or U/mg protein

Calculation Formulas

• α-Amylase (U/g) = (2 × x₁) ÷ W
• Total Amylase (U/g) = (10 × x₂) ÷ W
• β-Amylase (U/g) = [(10 × x₂) − (2 × x₁)] ÷ W
  Where:
• x₁, x₂ = glucose concentration from standard curve
• W = sample weight in grams
• Cpr = protein concentration (if using protein normalization)

Storage & Stability

• Store all components at 4°C upon receipt.
• Reconstituted reagents should be used promptly or stored as specified.

References

• Dziedzoave NT et al., Food Control, 2010: “Influence of variety and growth environment on β-amylase activity of flour from sweet potato.”