Question

Rewrite the procedure in third person & in past tense:

Procedure

You will be provided with post-mitochondrial supernatant fractions from rat liver of control rats and phenobarbitone-treated rats. Carry out the assay for drug metabolism as follows.

A

1. Take 4 ml incubation medium (containing substrate aminopyrine) into each test flask.
2. Take 4 ml incubation medium (containing no substrate into one flask (reagent blank)).
3. Add 700 ml 0.25 M sucrose solution to all flasks.
4. Pre-incubate all flasks at 37 °C for five minutes.

B

1. Add 300 ml liver homogenate supernatant to all flasks.
2. Shake unstoppered at 37 °C for 30 minutes.

C

1. Add 2 ml ZnSO₄.7H₂O (8.9%) solution to all flasks. Mix.
2. Remove flask from bath and leave for five minutes.
3. Add 2 ml saturated Ba(OH), 8H₂O solution. Mix well.
4. Transfer to 10 ml test tube and centrifuge at 200rpm for 10 minutes.

D

1. Prepare formaldehyde standards. Pipette 2.5 ml portions of each concentration into a test tube (LPS).  Use 2.5 ml water as a blank.
2. Add 1.5.ml Nash reagent to each tube and vortex.

E

1. Place 2.5 ml supernatant (from tubes treated under section C) into test tube.
2. Add 1.5 .ml Nash reagent. Vortex
3. Incubate all tubes (including standards) at 60 °C for 30 minutes.
4. Leave to cool.  Read absorbency at 412 nm against a blank reagent.
5. Read formaldehyde standards against a reagent blank.
6. Express results as ~mmol formaldehyde formed/h/g/liver (homogenate).

Answer 1

Procedure:

4mL incubation medium (containing substrate) was taken into each flask. 4mL of incubation medium (containing no substrate) was taken in one flask. 700mL of 0.25M sucrose solution was added to each flask. All flasks were pre-incubated at 37 degree Celsius for 37 minutes.

300 mL of liver supernatant was added to all flasks. The contents were shaken at 37 degrees for 30 minutes.

2mL of Zinc sulphate solution was added to each flask and mixed. The flasks were left from water bath and left for 5 minutes. 2mL of saturated barium hydroxide was added and mixed well. The contents of flasks were transferred to a 10mL test tube and centrifuged at 200rpm for 10 minutes.

Next, formaldehyde standards were prepared. 2.5 mL portions of each concentration were pipette out into a test tube. 2.5 mL water was used as blank. 1.5 mL of NaSH reagent was added to each tube and vortexed.

2.5 mL of supernatant from step C was placed in test tube. 1.5 mL of NaSH reagent was added. All tubes were incubated at 60 degrees for 30 minutes. It was left to cool. The absorbance values were recorded at 412 nm against a blank.

Formaldehyde standards' value were also recorded against blank solution.

The results were expressed as mmol formaldehyde formed/h/g/liver.