caa a22 4500 477126502 CHVBK 20180405111651.0 cr unu---uuuuu 170330e19971201xx s 000 0 eng 10.1023/A:1012115426306 doi (NATIONALLICENCE)springer-10.1023/A:1012115426306 The Stability and Degradation Pathway of Recombinant Human Parathyroid Hormone: Deamidation of Asparaginyl Residue and Peptide Bond Cleavage at Aspartyl and Asparaginyl Residues [Elektronische Daten] [Yoshiaki Nabuchi, Eri Fujiwara, Hitoshi Kuboniwa, Yoshinori Asoh, Hidetoshi Ushio] Purpose. The stability of recombinant human parathyroid hormone (rhPTH) was examined under acidic to alkaline conditions; its degradation pathways were elucidated from resultant products. Methods. Degradation assay was performed in the pH range 2 to 10 at 40, 50 and 60°C. The approximate molecular mass and pI values of the degradation products were estimated by electrophoresis. FAB-MS peptide mapping and amino acid composition analysis were used to determine these structures. The amount of each respective product was determined by HPLC. Results. At pH2, eight degradation products were found: l-30rhPTH, l-74rhPTH, l-71rhPTH, l-56rhPTH, l-45rhPTH, 46-84rhPTH, 31-84rhPTH and Asp76-rhPTH; these were mainly as a consequence of peptide bond cleavage of the amide bond of Asp. At pH9, five products were found: isoAsp16-rhPTH, Aspl6-rhPTH, Asp57-rhPTH, Asp76-rhPTH, 17-84rhPTH; the main degradation pathway was deamidation of Asn via a cyclic imide intermediate. Degradation products resulting from cleavage at Asp were increased in proportion to the extent that pH was lowered below 5. As pH was increased above 5, so were products resulting from deamidation of Asn. Correspondingly, levels of intact rhPTH were at a peak at pH5. Conclusions. Degradation of rhPTH under acidic conditions predominantly occurs by cleavage at Asp, whereas, above pH5, deamidation of Asn is the more prominent. rhPTH is most stable at pH5. Plenum Publishing Corporation, 1997 human parathyroid hormone nationallicence deamidation nationallicence cleavage nationallicence asparaginyl nationallicence aspartyl nationallicence Nabuchi Yoshiaki Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut Fujiwara Eri Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut Kuboniwa Hitoshi Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut Asoh Yoshinori Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut Ushio Hidetoshi Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut Pharmaceutical Research Kluwer Academic Publishers-Plenum Publishers 14/12(1997-12-01), 1685-1690 0724-8741 14:12<1685 1997 14 11095 https://doi.org/10.1023/A:1012115426306 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1023/A:1012115426306 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Nabuchi Yoshiaki Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut NATIONALLICENCE 700 1- Fujiwara Eri Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut NATIONALLICENCE 700 1- Kuboniwa Hitoshi Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut NATIONALLICENCE 700 1- Asoh Yoshinori Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut NATIONALLICENCE 700 1- Ushio Hidetoshi Fuji Gotemba Research laboratories, Chugai Pharmaceutical Co., Ltd., Gotemba, 412, Shizuoka, Japan aut NATIONALLICENCE 773 0- Pharmaceutical Research Kluwer Academic Publishers-Plenum Publishers 14/12(1997-12-01), 1685-1690 0724-8741 14:12<1685 1997 14 11095 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer