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Reagent Lab Preparation For Biochemistry Practice

lab regaent

REAGENT LAB PREPARATION

1.Anthrone Reagent

Dissolve 2 g of anthrone in 1 liter of concentrated sulfuric acid. Store in a refrigerator. Prepare a fresh solution every 2 day

2. Benedict’s Qualitative Solution

Dissolve 173 g of sodium citrate dehydrate and 100 g sodium carbonate anhydrous in 800 mL DI water. Warm and stir to aid dissolution. Filter if necessary. In a separate container, dissolve 17.3 g copper (II) sulfate pentahydrate in 100 mL DI water. Slowly, while stirring constantly, add the copper sulfate solution to the first solution. Let cool and dilute to 1 L with DI water. (test for the presence of simple sugars)

3. Iodine Solution, Lugol’s

Dissolve 20 g of potassium iodide in 200 mL of DI water; add 10 g of iodine; stir to dissolve then dilute to 1 L. Store in a dark bottle. (general biological stain and vital stain stock solution, dilute 5:1 before use.)

3. Biuret Solution

Dissolve 2.3 g of copper (II) sulfate pentahydrate in 230 mL of DI water. Set a side. Dissolve 308 g sodium hydroxide in 770 mL of DI water (very exothermic; cool vessel in an ice water bath) and cool to room temperature. Add all the copper sulfate solution to the sodium hydroxide solution. Solution should be blue. (test for proteins)

4. Molisch Reagent

Dissolve 5 g 1-naphthol in 100 mL of 95% ethyl alcohol (test for aldehydes, sugars, and carbohydrates)

5. Hanus solution (for iodine number)

Dissolve 13.2 g of resublimed iodine in 1 L of glacial acetic acid which will pass the dichromate test for reducible matter. Add sufficient bromine to double the halogen content, determined by titration (3 mL is about the proper amount). The iodine may be dissolved by the aid of heat, but the solution should be cold when the bromine is added.

6. Bradford’s reagent

Coomassie Brilliant Blue G (Sigma 98%) ……………..102,4 mg

Ethanol 95%…………………………………………………………50 mL

Phosphoric acid 85% .(w/v)…………………………………….100 mL

Mili-Q water q.s.p………………………………………………..1000 mL

Note: Protect from light all steps.

Steps for Bradford’s reagent:

Step 1: Weight out Coomassie.

Step 2: Add alcohol to Coomassie Blue in a 100mL Becker, covered with aluminum paper.

Step 3: Agitate with a stir bar for 1 hour.

Step 4: Pour this solution in one 1000-mL Erlenmeyer (protected from light with aluminum paper).

Step 5: Add 100 mL of phosphoric acid.

Step 6: Complete volume to 1000 mL using Mili-Q water (or DD water).

Step 7: Stir manually.

Step 8: Filter twice with paper filter (in the dark). Store solution in a bottle covered with aluminum paper.

Reference: Handbook of reagent lab preparation

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