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To preface this, I am a biologist.

I need to make a $\mathrm{pH}=6.8$, $\pu{1 M}$ Tris buffer. We have access to Tris base and Tris Hydrochloride. (Unfortunately, none of the fancy Tris compounds will work in my situation.)

I tried making a straight Tris base buffer, but before I could lower the $\mathrm{pH}$ to 6.8 (using concentrated $\ce{HCl}$), it crashed drastically. I assume that I got past the buffering capabilities.

Are there any recommendations for this? Thank you.

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  • $\begingroup$ You need $\mathrm{pH}$ of your Tris buffer @ 37 degrees, I believe. $\endgroup$ – Mathew Mahindaratne Mar 31 '18 at 3:10
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From Wikipedia...

The conjugate acid of tris has a pKa of 8.07 at 25 °C, which implies that the buffer has an effective pH range between 7.5 and 9.0.

So you're outside the range where TRIS is an effective buffer.

I assume by "crashing" you mean that a precipitate formed. A buffer is also rated based on its "buffer capacity." Effectively this depends on the the concentration of the buffer. Thus solubility of the buffers comes into consideration.

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Tris is short for tris(hydroxymethyl)aminomethane. Tris has a $\mathrm {pK_a}$ of $\mathrm {8.1}$ and a $\mathrm {pH}$ level between $\mathrm {7}$ and $\mathrm {9}$. Although that range may change at physological temperature, I doubt it go as low as $\mathrm {pH} = 6.8$ without sacrificing $\mathrm {[B]/[A]}$, which may go as low as $\mathrm {\approx0.05}$.

Thus, I think you should use $\mathrm {BIS-TRIS-propane}$ instead. $\mathrm {BIS-TRIS-propane}$ (1,3-Bis[tris(hydroxymethyl)methylamino]propane) is two Tris molecules connected by 1,3-diaminopropane. It has $\mathrm {pK_a}$s of $\mathrm {6.8}$ and $\mathrm {9.0}$, and effective $\mathrm {pH}$ range of $\mathrm {6.3-9.5}$.

I'm not sure how soluble it in deionized water, but worth trying ($\mathrm {$1/g}$). Probably a good idea to keep your stock solution in $\mathrm {0.5 M}$ range.

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Another choice: Ypu may use N,N-Bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (BES) (CAS Number: 10191-18-1), instead. It has p$K_\mathrm{a}$ of $7.193$ at $\pu{25 ^{\circ}C}$ (Ref.1) and useful pH range of $6.4-7.8$. BES is therefore better suited for pH control in the buffer region of biochemical interest (Ref.1,2). Sigma-Aldrich sells it in 99% pure form (~$1.0/g).

BES is soluble to the extent of $\pu{3.2 M}$ at $\pu{0 ^{\circ}C}$ (Ref.2) and can be easily purified from aqueous alcohol. Two equimolal buffer solutions consist of BES and sodium besate (sodium salt of BES), each at a molality of $\pu{0.02 mol kg^{-1}}$ and $\pu{0.10 mol kg^{-1}}$, respectively have shown p$K_\mathrm{a}$ value of each solution as $6.97$ at $\pu{37 ^{\circ}C}$ (Ref.1). On the other hand, a solution prepared with $\pu{0.03 mol kg^{-1}}$ of BES and $\pu{0.02 mol kg^{-1}}$ of sodium besate has shown p$K_\mathrm{a}$ value of $6.76$ at $\pu{37 ^{\circ}C}$ (Ref.1).

References:

  1. R. N. Roy, E. E. Swensson, G. LaCross, Jr., C. W. Krueger, “Standard buffer of N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (Bes) for use in the physiological pH range 6.6 to 7.4,” Anal. Chem. 1975, 47(8), 1407–1410 (DOI: 10.1021/ac60358a076).
  2. N. E. Good, G. D. Winget, W. Winter, T. N. Connolly, S. Izawa, R. M. M. Singh, “Hydrogen Ion Buffers for Biological Research,” Biochemistry 1966, 5(2), 467–477 (DOI: 10.1021/bi00866a011).
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