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Why is there a need for pH adjustment below ? Can pH go below 4.3 after adding acetic acid to PBS ?

“First prepare stock solution consisting of 50 ml of 10× PBS (pH 7.4), 12.5 ml of 80% glycerol, and 5 ml of glacial acetic acid. Adjust the pH to 4.3 with NaOH and then fill to 300 ml with distilled water. …”

Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5407559/

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    $\begingroup$ pH computations are not accurate. Authoritative is the fitting to the official practical pH scale defined by set of pH buffers. Unless particular solution is previously measured and precisely reproduced, it has to adjusted by measuring it pH..// Some pH buffers used for a given pH range are explicitly defined by adjusting with acid/base to the target pH. And yes, the pH is expected to go below 4.3, probably a lot. $\endgroup$
    – Poutnik
    Commented Oct 10, 2021 at 17:14

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Yes. Adding $5$ mL pure acetic acid to $50$ mL phosphate base saline (PBS) produces a pH value situated between $2$ and $3$, depending upon the exact concentrations of the phosphates of the PBS. Different mixtures are known. The buffer capacity of PBS is exceeded by the huge amount of added acetic acid.

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  • $\begingroup$ The acetic acid/acetate concentration in the final solution will be about 290 mM. 10 x PBS could be about 100 mM phosphate, and it is diluted sixfold, so it will be less than 20 mM phosphate. At a pH of 4.3, the final solution is an acetate buffer rather than a hydrogen phosphate/dihydrogen phosphate buffer (PBS stands for phosphate buffered saline). $\endgroup$
    – Karsten ♦
    Commented Oct 10, 2021 at 16:16
  • $\begingroup$ Thanks for the answers. The reason I am puzzled is actually if this passes the buffer capacity, why is the buffer used ? Does it have a benefit as a buffer other than providing saline at the end ? $\endgroup$
    – mete
    Commented Oct 10, 2021 at 16:38
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    $\begingroup$ @mete This solution is easy to make (because the stock solutions are typically already available in the lab) and it works. It is very possible that nobody checked whether there is any effect of leaving out one or the other ingredient. $\endgroup$
    – Karsten ♦
    Commented Oct 10, 2021 at 23:56

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