New HPLC Amino Acids Separation Compatible With Carbon Dating Technique



Hydroxyproline seems to be the most promising amino acid used in carbon dating when isolated from bone collagen. Separation of amino acids is challenging, especially without the use of ions or inorganic buffers that can interfere with Mass spectrometer (MS) or contaminate the sample with modern carbon. Amino acids are also not retained in reverse-phase chromatography. The ideal solution would be using water only to separate the amino acids. This would allow a direct coupling to MS. We were able to separate hydroxyproline from proline and other simple amino acids like glycine and alanine in HPLC on Newcrom AH column using water only as a mobile phase. Using water also allowed UV detection at 205 nm which can’t be done if using a buffer based on acetic or formic acid.
See more information on radiocarbon dating here.
The same method can be modified to get symmetrical peaks and higher efficiency if a mobile phase with ionic modifier such as formic acid is used.


Column Newcrom AH, 4.6×150 mm, 5 µm, 100A
Mobile Phase MeCN/H2O
Flow Rate 1.0 ml/min
Detection UV, 205 nm, CAD



Class of Compounds
Drug, Acid, Hydrophilic, Ionizable, Vitamin, Supplements, Amino acid
Analyzing Compounds Alanine, Glycine, Proline, Hydroxyproline


Application Column

Newcrom AH

The Newcrom columns are a family of reverse-phase-based columns. Newcrom A, AH, B, and BH are all mixed-mode columns with either positive or negative ion-pairing groups attached to either short (25 Å) or long (100 Å) ligand chains. Newcrom R1 is a special reverse-phase column with low silanol activity.

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Application Analytes:
SIELC Technologies usually develops more than one method for each compound. Therefore, this particular method may not be the best available method from our portfolio for your specific application. Before you decide to implement this method in your research, please send us an email to so we can ensure you get optimal results for your compound/s of interest.