[vivian87]

I am doing an experiment on protein solublity of soya bean meal. The method I am using is 0.2% KOH. But I entitle some problem in calculation and determination of protein extracted from KOH. Please help me to have a look. Thank you very much.

Method:
Place 1.5g sample in 250ml beaker. Add 75ml of 0.2% KOH solution. Stir mixture for 20minutes with magnetic stirrer. At end of 20min, transfer 50mL of liquid to centrifuge tube. Then centrifuge at 6000rpm for 5min. Collect supernatant for Kjeldahl method.

And now here is my problem. How many supernatant should I use as sample in Kjeldahl? And how to calculate for the protein solubility? I read 1 calculation in internet, but I don't really understand.

Example:
Sample weight: 1.5072g
Liquid Kjeldahl value: 1.125 mg N/ml
Dry Kjeldahl value:47.47%

1.125 mg N/mL x 75mL (how much KOH added) = 84.4mg N / 1.5072g = 55.98mg N x 100g sample = 5.598g N
5.598g N * 6.25 = 34.99% CP
(34.99/47.47) x 100 = 73.68% solubility


Why 84.4mg N need to divide by sample weight? And why 55.98 mg N need to divide by 100g sample?

[Borek]

Why 84.4mg N need to divide by sample weight? And why 55.98 mg N need to divide by 100g sample?

84.4 mg divided by sample mass gives mass of nitrogen that goes into solution per 1g of substance, looks like a reasonable unit to me. Whatever happens later seems a little bit strange. There can be some logic behind, but I have troubles grasping it.

What is 6.25 (where does it come from) and what is CP (what does the acronym mean)?

[vivian87]

6.25 is the constant. CP is crude protein...

[Borek]

I bet it means that nitrogen is assumed to be 1/6.25 of the crude protein. If so, multiplication by 100g is a just a convoluted way of getting to %. 5.598g of nitrogen in 100g means 5.598% of nitrogen, multiply by 6.25 and the sample contains 35% CP.

[vivian87]

I see. Now i get a rough idea already.
The protocol said perform Kjeldahl on 15ml supernatant (mg N/ml setting; input 15 as the sample size). What does it mean?
How to set mg N/ml setting? For dry sample, the sample size is in g and the final result will be in %. Then how about for this liquid sample? Is it possible for me to do as dry sample? For dry sample, I will use 0.5g sample. Can I just weigh 0.5g liquid as my sample size so that the calculation is same as the dry sample? Or it is necessary to use mg N/ml for liquid sample?

[Borek]

To be honest I have problems following you.

Most of these calculations are based on mass conservation, dilution and mixing - see http://www.chembuddy.com/?left=concentration&right=dilution-mixing. If you take just a apart of the sample (say 15 mL of supernatant) and determine CONCENTRATION of nitrogen per mL in the supernatant - you can be sure the same concentration was in the original sample volume. If you determine AMOUNT of nitrogen in the 15 mL supernatant, it was fraction of the original amount - and what fraction it was, depends on the ratio of volumes of sample and supernatant.