Solarbio Total kapasitas antioksidan (T-AOC) Perangkat Pengujian

Kapasitas antioksidan total (T-AOC) Perangkat Pengujian

Catatan: Ambil dua atau tiga sampel berbeda untuk prediksi sebelum pengujian.

Peralatan Operasi: Spektrofotometer

Kucing No: BC1310

Ukuran：50T/48S

Komponen:

Extract solution: Cairan 50 mL×1. Penyimpanan pada suhu 4℃, precool before use.

Reagen I: Cairan 35 mL×1. Penyimpanan pada suhu 4℃.

Reagen II: Cairan 20 mL×1. Storage at 4℃ in shadow.

Reagen III: Cairan 5 mL×1. Storage at 4℃ in shadow.

Standar: Bedak×1, 10 mg of FeSO4·7H2O. Solusi kerja: menambahkan 0.9 mL of distilled water and 20μL of concentrated sulfuric acid (H2SO4) to forms 40 µmol/mL FeSO4 standard solution. Solution mixture (prepare when the solution will be used): Reagen I : Reagen II: Reagent III= 7:1:1, incubate at 37℃ before use.

Deskripsi Produk:

This kit is used to detect the total antioxidant levels of antioxidants and antioxidant enzymes in the samples. It is mainly used in the study of biological, medical and pharmaceutical studies to detect the total antioxidant capacity of antioxidant solutions.

In acid environment, Fe3+ -TPTZ are reduced to blue Fe2+ -TPTZ. The color reaction reflects the total antioxidant capacity.

Reagen dan Peralatan Diperlukan tetapi Tidak Disediakan:

Spektrofotometer, constant temperature water bath, low temperature centrifuge, 1 mL glass cuvette and distilled water.

Prosedur:

SAYA. Persiapan sampel:

1. Serum, plasma, saliva, or urine samples

Plasma (anticoagulation with heparin or sodium citrate, avoid using EDTA), sentrifugasi di 5000 rpm/min for 10 menit, take supernatant for test. Take serum, saliva or urine samples for direct determination. Juga, you can store at -80℃ and detect within 30 hari.

2. Cells or tissue sample

Mengambil 1-2 million cells or 0.1 g tisu, menambahkan 1.0 mL of Extract solution. Use homogenate or ultrasound to fully break up cells and release antioxidant, sentrifugasi di 10000 r/min and 4℃ for 5 menit, take supernatant for test. Measure the concentration of protein if needed.

II. Determination procedure:

1. Dilute 40 μmol/mL FeSO4 standard solution to 0.1, 0.05, 0.025, 0.0125, 0.00625, 0.003125 μmol/mL, mengambil 500 μL of standard solution (distilled water for blank control), add to 500 μL of Reagent II. Mix thoroughly for 10 menit, detect the absorbance in 593 nm, calculate ΔA=AS-AB. (SEBAGAI: standard solution tube, AB: blank control tube.) The final concentration of Fe2+ is 0.05、0.025、0.0125、0.00625、

0.003125、0.00156 μmol/mL.

2. Preheat the spectrophotometer 30 menit, sesuaikan panjang gelombang ke 593 nm and set zero with distilled
3. Tambahkan reagen dengan daftar berikut:

AKU AKU AKU. Perhitungan:

1. Buat kurva standar

Take the Fe2+ final concentration as X-axis, △A as Y-axis, create standard curve, get linear regression equation y=kx+ b, take ΔA’ into the equation to get x (μmol/mL).

2. Unit definition: the sample antioxidant capacity is indicated by the standard liquid ion concentration required for the same absorbance change(ΔA).

A. Protein concentration:

Kapasitas antioksidan total (μmol/mg prot) = x × Vrv÷ (Vs× Cpr) = 34× x ÷ Cpr

B. Sampel weight

Kapasitas antioksidan total (μmol/g weight) = x × Vrv÷ (Vs ÷ Vsv ×W) =34× x÷ W

C. Cell amount

Kapasitas antioksidan total (μmol/104cell) = x × Vrv÷ (Vs ×Vsv÷n) = 34× x÷ n

D. Solution volume

Kapasitas antioksidan total (μmol/mL) =x× Vrv÷ Vs =34×x

Tali: total reaction volume, 1.02 ml; Vs: sample volume, 0.03 ml;

Vsv: extraction volume, 1 ml; W: sample weight, G;

Cpr: sample protein concentration, mg/mL;

N: cell amount, unit based on 104 (ten thousand).

Catatan:

1. Reagent II is irritated to human body, please wear lab clothes and latex
2. The samples should not be appeared blue under acidic condition, or it will interference sample result of the
3. Detergent such as Tween, Triton, NP-40 and reductants such as DTT, mercapto ethanol should not be added in the
4. If the absorbance value determined by the sample is beyond the standard curve range, the sample should be diluted or concentrated properly before
5. The kit should be store at2-8℃.

Examples:

1. Add 0.1g shamrock to 1mL extract solution and grind thoroughly on ice, take supernatant, follow the determination procedure to operate, with 96-well flat-bottom plates to calculate: ΔA=A(T)-A(B)=0.909-0.148=0.761, standard curve: y=21.056x-0.0087, calculate x=0.037, according to mass of sample to calculate Total antioxidant capacity (μmol/g mass) =34×x÷W=34×0.037÷0.1=12.85 μmol/g mass.

Referensi：

[1] Pellegrini N, Serafini M, Salvatore S, dkk. Total antioxidant capacity of spices, dried fruits, nuts,

pulses, cereals, and sweets consumed in Italy assessed by three different in vitro assays[J]. Molecular nutrition & food research, 2006, 50(11): 1030-1038.

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