USP Malic acid standard by HPLC has two separate peaks

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I run a pure Malic acid standard from USP by using C18 column (Phenomenex Kinetex 2.6µ, P/N 00F-4496-E0 ). My mobile phase is 0.012N HCL and 1 mL/minute. There are two peaks in chromatogram: one is 1.92 minute and one is at 3.78 minute. the sample prep. is 15 mg/ 25 mL of DI water, and inject at 1 uL. Anyone know why I get a second peak. Thanks in advance.
I'm only experienced with organic acids like this on a specialty organic acid column, not on RP-18. We used a conductivity detector.

1. Detail what happens when you just just inject mobile phase.

2. Detail what happens when you make standard up at 3 times the concentration and inject that.

3. Have you used this assay before and this is just a new container of malic acid, or is this the first time you've tried to do this?
The column may be overloaded or there may be a solvent mismatch problem. Try to decrease the concentration of the standard solution or to decrease the injection volume. Also try to dissolve the standard in the mobile phase instead of water. Check the chromatograms of the injected solvent (mobile phase or water) as well. Is the detector a UV detector or a RID? What is the wavelength? What is the height of the main peak (at 3.78 min?)?
By the way, HCl is not a good mobile phase (since it is corrosive) for steel columns and capillaries. H3PO4 solution with appropriate pH (about 2) would be better.
Another possible cause of the poor chromatogram is the possible dewetting of C18 stationary phase in the aqueous eluent.
We use UV detector at 210 nm. We only use the peak at 1.9 minute for Malic acid, and the area is 30 mAu at 1 ul injeciton (15 mg/25 mL). The peak at 3.7 is 22 mAu.
This is my first time to work with this method, we have been used this for years now. I ask the question around, nobody has an answer. I agree that HCL isn't a good choice. But the method was set up that way for years, and to change we have to go through a validation process.
I don't why I did think of a blank injection and increase the concentration. I will try those suggestions. I will try phosphate solution too. But I don't see how 0.012 N HCL could cause the extra peak. Thank you for the suggestions.
HCl itself does not cause extra peaks. The only possible problem with HCl in comparison with H3PO4 is that HCl is more corrosive to steel than H3PO4; hence, HCl is not recommended for acidifying mobile phases.

Another possible cause of the two peaks is the fact that malic acid usually contains an impurity of fumaric acid. At 210 nm fumaric acid absorbs UV light much stronger than malic acid. Thus, even minor impurity of fumaric acid may result in a peak with the size comparable to the size of the peak of malic acid. Check whether the peak at 3.78 min is the peak of fumaric acid.
I run a water blank and 0.012 N HCL blank. There isn't any interference. I used Potassium Phosphate Dibasic as solvent too which has a pH at 2.2. The second peak still there and no RT change. I will try to run Fumaric acid later. Thank you very much.
HCL solution is not a good mobile phase choice. It can be easily replaced with 0.1% H3PO4. Why is the PH of potassium dibadic solution 2.2? It should be basic. Did you adjust it with h3po4? Why not just use h3po4?

I have used 0.1% H3PO4 for the analysis of organic acids multiple times. The method is very robust and works very well. A step gradient to 100% ACN after elution of the target is important to have as this can 1) remove more hydrophobic components in the sample from the column and 2) prevent dewetting as long as the pump is not stopped.

Try cleaning the column with ACN and reequilibrate it with the mobile phase before injection of the malic acid standard.
HCL solution is not a good mobile phase choice. It can be easily replaced with 0.1% H3PO4. Why is the PH of potassium dibadic solution 2.2? It should be basic. Did you adjust it with h3po4? Why not just use h3po4?

I have used 0.1% H3PO4 for the analysis of organic acids multiple times. The method is very robust and works very well. A step gradient to 100% ACN after elution of the target is important to have as this can 1) remove more hydrophobic components in the sample from the column and 2) prevent dewetting as long as the pump is not stopped.

Try cleaning the column with ACN and reequilibrate it with the mobile phase before injection of the malic acid standard.
Yes, you are right that I use phosphoric acid to adjust the pH for the buffer. I was hoping by using the phosphate buffer to eliminate the second peak in my standard injection, but it didn't work. I agree that phosphate is a better choice. My major problem is that a second peak in my chromatogram which I couldn't explain it.
I'm almost sure that the second peak is due to fumaric acid.
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