Calculate the mass or volume of each component need to make the following buffer

Question

Calculate the mass or volume of each component need to make the following buffers. You must show calculations Triton Lysis Buffer (250 mL), pH 7.5 50 mM Tris 150 mM NaCl 0.5% Triton (v/v) Instructions for making buffer: Obtain an appropriate size beaker. Then add water to ~ 100 mL. Then, measure out glycerol and pour into the beaker. Start the solution stirring with magnetic stir bar on a stir plate. Weight out and add other dry components. To add in triton (which is very viscous): Use a 1000 ul Pipette with the end of the tip cut off. Carefully and slowly draw up triton. Pipette into beaker and pipette up and down into the buffer to wash the tip. Once solution is well mixed, add water to ~ 450 mL and pH to 7.5. Use a graduate cylinder to get the final volume to 500 mL. Transfer buffer to a 500 mL labeled bottle.

Explanation / Answer

0.5% (V/V) means 0.5mL triton is present in 100mL solution.

amount of triton present in 250mL = 0.5mL *250mL/100mL = 1.25 mL

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250 mL 150mM NaCl = 150mM * 0.25 L = 37.5 mmol = 0.0375 mol

Mass of NaCl = 0.0375 mol * 58.5 g/mol = 2.194 gm

Amount of NaCl required = 2.194 gm

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pH = pka + log [TrisH+/Tris]

7.5 = 8.1 + log [TrisH+/Tris]

or, [TrisH+/Tris] =0.25

calculate decimal fraction .

[TrisH+] = 0.25/1+0.25 = 0.25/1.25 = 0.2

[tris] = 1/1+ 0.25 = 0.8

Molarity of trisH+ = 0.2 * 0.05M = 0.01 M

moles of trisH+ = 0.01 M * 0.25 L = 2.5*10^-3 moles

mass of trisH+ = 2.5*10^-3 moles *157.60 g/mol =0.394 g

Molarity of tris=0.8 *0.05 M = 0.04 M

number of moles = 0.04 * 0.25 L =0.01 moles

,Mass of tris = 0.01 mol * 121.14g/mol = 1.21 gm

1.21 gm tris + 0.39 gm tris-HCl has to be added to 250mL water

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