[onionfart]

Hi I ran an alanine standard through an HPLC column using MeCN/H2O as a mobile phase. Started out 0 MeCN and ramped up from 40 to 100 at that end portion. Should I be starting out with high MeCN to H2O? Is the Alanine one of the first two peaks or located within the last set of peaks. This is an old column and have not been guaranteed that it is still functioning correctly.
https://imgur.com/a/TsPWpoS

[Babcock_Hall]

In reverse phase, which solvent is more strongly eluting, water or acetonitrile?  Did you run a blank prior to your sample?

Determining which peak is alanine is not a straightforward matter, but you might try changing the composition of the mobile phase slightly as an experiment.