Determination of amino acids in diverse polymeric matrices using HPLC, with emphasis on agars and agaroses.
Determination of amino acids in polymers with varying structure and charge was performed using vapor phase acid hydrolysis and subsequent precolumn derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC). Percent load of neutral, cationic and anionic peptide-modified synthetic polymers was accurately determined using this technique. Assay utility was shown for glycosaminoglycans and other sulfated polymers, neutral carbohydrate polymers such as agar, agarose, and cellulose, and polymers such as lipopolysaccharide and deoxyribonucleic acid. The carboxylated and sulfated molecules included chondroitin sulfate, hyaluronic acid, dermatan sulfate, and heparin, and the sulfated polymers included fucoidan, carrageenan, and dextran sulfate, as examples. Assayed cumulative amino acid concentrations (i.e. protein levels) are reported, although amino acid distribution data was also available from the analysis. Recovery was acceptable for the various compounds tested and did not correlate with structure. However, different sample sizes were necessary to achieve acceptable recovery, depending on the level of protein present in the matrix. While some matrices contained peaks in addition to the amino acids and amino sugars, they were not found to interfere using the standard gradient separation. Assayed amino acid profiles were compared for agaroses with differing electroendosmosis values and for agar samples from different parts of the globe. While the amounts of protein varied depending on source, the relative distribution of amino acids was very similar across the agar samples surveyed.
Palace
G P
GP
Analytical Chemistry and Chemistry Departments, GelTex Pharmaceuticals, Incorporated, Waltham, MA 02451, USA. gpalace@geltex.com
Fitzpatrick
R
R
Tran
K V
KV
Phoebe
C H
CH
Norton
K
K
D000362:Agar / Q000737:chemistry; D000596:Amino Acids / Q000032:analysis*; D000634:Aminoquinolines; D002219:Carbamates; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002942:Circular Dichroism; D005782:Gels / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D011108:Polymers / Q000737:chemistry*; D011506:Proteins / Q000032:analysis / Q000737:chemistry*; D012685:Sepharose / Q000737:chemistry
C078178:6-aminoquinolyl-N-hydroxysuccinimidyl carbamate; D000596:Amino Acids; D000634:Aminoquinolines; D002219:Carbamates; D005782:Gels; D010455:Peptides; D011108:Polymers; D011506:Proteins; D000362:Agar; D012685:Sepharose