Application

This kit can be used to measure the Flavonoids content in plant tissue samples. 

Detection significance

Flavonoids are common plant secondary metabolites, such as red, blue, and purple anthocyanins in plant tissues. Flavonoids can scavenge free radicals directly by hydrogen atoms. The ability of oxidation resistance of many flavonoids is higher than vitamin C and vitamin E.

Detection principle

In alkaline nitrite solution, flavonoids form red complex with aluminum ion. The flavonoid content of the sample can be calculated by measuring the absorptivity of the sample extract at 510 nm.

Experiment instruments

Micropipettor, Vortex mixer, Centrifuge, Vacuum dryer, Ultrasonic cell disruptor, Constant temperature shaking incubator, Spectrophotometer (510 nm).

The pretreatment of sample

1. Drying and crushing of plant tissues

Weigh 5-10 g fresh plant tissue and wash with distilled water, absorb moisture on the surface of tissue with filter paper, then put in a vacuum dryer and dry to constant weight at 80℃. Crush the sample and filter over 40 mesh screen, sealed at room temperature.

2. Extraction of Plant tissue

Accurately weigh 0.02 g sample in step 1, add 2 mL of 60% alcohol (self-prepared), then shake at 60℃ for 2 hours with constant temperature shaking incubator. Centrifuge at 1500 g for 10 min, then take the supernatant for detection. Or treat the sample with ultrasonic cell disruptor (power: 300W, 3 seconds/time, interval for 4 seconds, repeat for 30 min), then centrifuge at 10000 g for 10 min, then take the supernatant for detection.

Operation steps

1. The preparation of standard curve

Dilute the 1 mg/mL standard solution with double distilled water to a serial concentration. The recommended dilution gradient is as follows: 150, 120, 100, 80, 60, 20 mg/mL

2. The measurement of samples

1) Standard tube: Add 0.54 mL of standard solution with different concentrations into the 2 mL EP tubes.

Sample tube: Add 0.54 mL of Sample into a 2 mL EP tube.

Blank tube: Add 0.54 mL of double distilled water into a 2 mL EP tube.

2) Add 0.03 mL of Reagent 2 into each tube, oscillate fully and stand for 5 min at room temperature.

3) Add 0.03 mL of Reagent 3 into each tube, oscillate fully and stand for 5 min at room temperature.

4) Add 0.4 mL of Reagent 4 into each tube, oscillate fully and stand for 15 min at room temperature.

5) Set the spectrophotometer to zero with double-distilled water and measure the OD values of each tube at 510 nm with 0.5 cm diameter cuvette.

Note: It can be refer to the following operating table.

Technical parameter

1. The sensitivity of the kit is 0.315 μg/mL.

2. The intra-assay CV is 1.9 % and the inter-assay CV is 2.2 %.

3. The recovery of the kit is 98 %.

4. The detection range of the kit is 0.315-150 μg/mL.

Notes

1. The kit is for scientific research only.

2. Instructions should be followed strictly, changes of operation may result in unreliable results.

3. The valid of kit is 3 months.

4. Do not use components from different batches of kit.