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Buffer acetate solution pH=4.40 for UPLC

Discussions about HPLC, CE, TLC, SFC, and other "liquid phase" separation techniques.

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Dear colleagues,
I need to know how to prepare a buffer acetate solution at pH 4.40 for using in BEH C18 column in UPLC.

Thank you in advance.
Hello mcervera,

there are some ways you can go from here but everything starts with the definition of the concentration of the solution you desire. Always remember: when thinking of a buffer you have to think of pH value and concentration... and then check out for molecules that have a pKa that makes it feasible to create the buffer. remember, always consider buffering range as pKa +-1 and the closer you are to this value higher the concentration you may need to maintain buffer capacity.

Now, you have some lots of ways to think this through. some ways are more correct than others but as long as you don't use a crazy amount of salt in your mobile phase and pH is well controlled, you should not have a problem. please be advised however that once you have one method going you should stick to it forever (considering the scenario as the method you have developed) since there might be changes in actual values from one buffer preparation procedure to other. also, always use acids and bases that correspond to what you have (example: for amonium acetate buffers we correct pH values only with ammonium hydroxide and acetic acid, nothing more).

Many people will basically measure a quantity of salt that is equivalent to the quantity in mols of the desired buffer concentration, measure the pH and titrate it with either a basic or acidic solution until it reaches pH.

if you want to be more precise you can make a two system equation. first equation should be the Henderson-Hasselbalch equation, second should be the sum of mols from the acid and the salt (conjugate base) addition should be equal to your chosen buffer concentration. from this you have 2 equations and 2 values (mols of acid and mols of conjugated base). you may use an acid or base solution to correct the value if necessary. remember to have high purity acids and/or consider the purity when calculating a volume (simple conversion using molecular weight and density from the mol quantity) and to consider the hydration of the acetate (if there is anything on the label) when calculating the salt weight.

i hope this helps!

best regards,
Luccas Name

YMC applications specialist (LATAM)
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