著者

Sae SOTOMATSU Tomohiro YAMADA Hajime MIZUNO Hideki HAYASHI Toshimasa TOYO’OKA Kenichiro TODOROKI

出版者

The Society for Chromatographic Sciences

雑誌

CHROMATOGRAPHY (ISSN:13428284)

巻号頁・発行日

pp.2019.013, (Released:2019-06-21)

参考文献数

16

被引用文献数

6

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To construct liquid chromatography (LC)-based bioanalytical method for therapeutic monoclonal antibodies (mAbs) and antibody-drug conjugates (ADCs), twelve commercially available therapeutic mAbs and one ADC were chemically reduced, and the generated fragments were analyzed by high-temperature reversed-phase LC. For most therapeutic mAbs, single peaks of light and heavy chains were detected, indicating a possibility of homogeneous LC analysis using light chains. However, characteristic fragmentations were observed in infliximab, pembrolizumab, ramucirumab, and trastuzumab emtansine. We also performed a simple validation using the fragmented light chains for the bioanalysis of bevacizumab. The limit of detection (LOD) and limit of quantification (LOQ) of bevacizumab were 0.63 and 2.10 µg/mL, respectively, with dithiothreitol reduction, and 0.74 and 2.48 µg/mL, respectively, with tris (2-carboxyethyl) phosphine reduction. These results indicate that both the reductants confer sufficient linearity, LOQ, and LOD for the light chain analysis of bevacizumab. Thus, this method, combined with affinity purification, can be used for the bioanalysis of bevacizumab.

著者

Sachise KARAKAWA Masashi HARADA Rumi NISHIMOTO

出版者

The Society for Chromatographic Sciences

雑誌

CHROMATOGRAPHY (ISSN:13428284)

巻号頁・発行日

vol.44, no.1, pp.21-26, 2023-02-20 (Released:2023-03-11)

参考文献数

47

被引用文献数

1

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Three different types of analytical methods for D,L-amino acid were developed. First, a method for simultaneous analysis of D,L-amino acids in food was constructed by combining pre-column derivatization with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), UHPLC separation, and UV and circular dichroism (CD) detection. Using this method, D,L-amino acids in food samples was determined at a very short time of 5.5 minutes. Second, a pre-column derivatization LC/MS method (direct method) for D-alanine, D-serine, and D-aspartic acid and L-isomers in biological samples was developed. In this method, D,L-amino acids were derivatized with an achiral reagent, p-N,N,N-trimethylammonioanilyl N′-hydroxysuccinimidyl carbamate iodide (TAHS), separated with a chiral column, and detected using mass spectrometry (MS). Third, a pre-column derivatization LC/MS method (indirect method) for highly sensitive and selective analysis of 20 D,L-amino acids using a chiral derivatization reagent, (R)-4-nitrophenyl-N-[2-(diethylamino)-6,6-dimethyl-[1,1-biphenyl]-2-yl] carbamate hydrochloride ((R)-BiAC), was developed. The unique structure of the (R)-BiAC reagent, which has a biaryl axially chiral structure, allowed for higher resolution and a high sensitive analysis of the 20 D,L-amino acids within 12 minutes. This method has been applied to various D-amino acid research studies, and it can be applied to a wide range of analyses from food to biological samples. Lastly, an LC/MS method was established that simultaneously analyzes tryptophan and 15 of its metabolites, which is involved in neurotransmission and immunity in vivo.

著者

Naoki NISHIMURA Toyohiro NAITO Takuya KUBO Koji OTSUKA

出版者

The Society for Chromatographic Sciences

雑誌

CHROMATOGRAPHY (ISSN:13428284)

巻号頁・発行日

vol.39, no.3, pp.113-118, 2018-10-20 (Released:2018-11-08)

参考文献数

36

被引用文献数

6 4

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In this study, we reveal the suppression of non-specific hydrophobic interaction in poly(ethylene-co-glycidylmethacrylate) (PEGM) based spongy monolith (SPM), PEGM-SPM, which has recently be reported as a new platform of separation medium for affinity chromatography in our previous study, by an simple acidic treatment. Additionally, the immobilization procedures of protein-A toward the PEGM-SPM and the separation conditions for immunoglobulin G (IgG) were optimized for further effective affinity separations. As a result of treatment by a mixture of trifluoroacetic acid and acetonitrile, the hydrophobicity was dramatically suppressed in the PEGM-SPM. The optimizations for the density of PEGM in the PEGM-SPM, the protein A immobilization, and the binding/releasing conditions showed that variety of proteins and peptides were not retained on the protein A immobilized spongy column at all while IgG was absolutely separated by a simple stepwise pH gradient condition.

著者

Yasushi ISHIHAMA

出版者

The Society for Chromatographic Sciences

雑誌

CHROMATOGRAPHY (ISSN:13428284)

巻号頁・発行日

pp.2019.023, (Released:2019-10-10)

参考文献数

64

被引用文献数

10

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Mass spectrometry-based proteomics platforms have been widely used as ‘proteome sequencers’ to characterize the proteomes of a wide range of organisms. The work-flow generally involves multiple steps of sample preparation, peptide purification/concentration/pre-fractionation, and nanoLC/MS/MS measurement. This review focuses on our contributions to the technical development of current proteomics platforms, and includes a consideration of the limitations of these systems, together with the prospects for developing superior new-generation proteome sequencers.

著者

Takako HAYASHI Teruhisa FUJIMAKI Maki MIYAZAWA

出版者

The Society for Chromatographic Sciences

雑誌

CHROMATOGRAPHY (ISSN:13428284)

巻号頁・発行日

pp.2018.011, (Released:2018-09-29)

参考文献数

16

被引用文献数

4

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We examined the utility of LC-MS/MS/MS in the rapid quantitative analysis of clenbuterol in pig liver. Compared with LC-MS/MS in the SRM mode, which gave a chromatogram of pig liver containing interference peaks near the target component peak, LC-MS/MS/MS generated a clear chromatogram with no interference peaks. Validation of the method yielded favorable results for both LC-MS/MS in the SRM mode and LC-MS/MS/MS, with trueness values of 95.5% and 102%, repeatability of 1.9% and 5.5%, and within-laboratory reproducibility of 2.0% and 5.1%, respectively. Moreover, the recovery of clenbuterol-d9 used as surrogate was ≥70% in both measurement methods and thus, validation was achieved in terms of both selectivity and limit of quantification. The results suggest that LC-MS/MS/MS has highly selectivity for quantitative analysis as the influence of interference peaks is suppressed, and can be used when quantification is difficult with LC-MS/MS in the SRM mode due to matrix effects.