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Identification of Subvisible Particles in Biopharmaceutical Formulations Using Raman Spectroscopy Provides Insight into Polysorbate 20 Degradation Pathway.

Saggu M, Liu J, Patel A - Pharm. Res. (2015)

Bottom Line: To study composition and heterogeneity of insoluble subvisible particles in Mab formulations resulting from degradation of polysorbate 20 and to develop a better understanding of the mechanisms of polysorbate degradation leading to particle formation.Most of the subvisible particles identified were comprised of mixtures of fatty acids with no observable signal from fatty acid esters consistent with hydrolysis being the predominant degradation mechanism leading to particulate formation under these storage conditions.Our methodology is generally applicable for identification of particles in antibody formulations and, in particular, has the potential to give detailed information about particle heterogeneity and insight into mechanistic aspects of particle formation.

View Article: PubMed Central - PubMed

Affiliation: Late Stage Pharmaceutical Development, Genentech Inc., South San Francisco, California, 94080, USA, saggu.miguel@gene.com.

ABSTRACT

Purpose: To study composition and heterogeneity of insoluble subvisible particles in Mab formulations resulting from degradation of polysorbate 20 and to develop a better understanding of the mechanisms of polysorbate degradation leading to particle formation.

Methods: In this study, we exploit the potential of Raman microscopy for chemical identification of particles in monoclonal antibody formulations. Through a combination of experiments and density functional theory (DFT) calculations, we identified unique spectral marker bands for insoluble degradation products of polysorbate 20. We first applied our methodology to identify particles in model systems containing complex mixtures of fatty acids and then to subvisible particles in antibody formulations stored at 5°C for several years.

Results: Most of the subvisible particles identified were comprised of mixtures of fatty acids with no observable signal from fatty acid esters consistent with hydrolysis being the predominant degradation mechanism leading to particulate formation under these storage conditions.

Conclusions: Our methodology is generally applicable for identification of particles in antibody formulations and, in particular, has the potential to give detailed information about particle heterogeneity and insight into mechanistic aspects of particle formation.

No MeSH data available.


Experimental Raman spectra of dry fatty acids (a) capric acid (b) lauric acid (c) myristic acid and (d) palmitic acid. Experimental conditions: 4 mW laser power, T = 298 K, 60 s accumulation time.
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Fig1: Experimental Raman spectra of dry fatty acids (a) capric acid (b) lauric acid (c) myristic acid and (d) palmitic acid. Experimental conditions: 4 mW laser power, T = 298 K, 60 s accumulation time.

Mentions: Raman spectra of the dry fatty acids capric acid, lauric acid, myristic acid and palmitic acid were obtained using low laser power and are shown in Fig. 1. Characteristic for all fatty acids is the presence of two intense bands between 1050 and 1150 cm−1 arising from ν(C-C) stretch vibrations, one band at 1299 cm−1 arising from γ(CH2) twisting and a group of bands between 1400 and 1500 cm−1 caused by γ(CH2) wagging, γ(CH2) scissoring as well as δ(CH2) and δ(CH3) deformation vibrations (21). Typically carbonyl stretch frequencies of aliphatic compounds are found above 1700 cm−1. The carbonyl stretch frequency ν(C = O) of the fatty acids is red-shifted to 1640 cm−1 and shows only moderate Raman intensity indicating a local environment in which the carbonyl is hydrogen bonded (22). This peak is split into two peaks for lauric, myristic and palmitic acid suggesting heterogeneity of hydrogen bonding in the particle structure.Fig. 1


Identification of Subvisible Particles in Biopharmaceutical Formulations Using Raman Spectroscopy Provides Insight into Polysorbate 20 Degradation Pathway.

Saggu M, Liu J, Patel A - Pharm. Res. (2015)

Experimental Raman spectra of dry fatty acids (a) capric acid (b) lauric acid (c) myristic acid and (d) palmitic acid. Experimental conditions: 4 mW laser power, T = 298 K, 60 s accumulation time.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4526581&req=5

Fig1: Experimental Raman spectra of dry fatty acids (a) capric acid (b) lauric acid (c) myristic acid and (d) palmitic acid. Experimental conditions: 4 mW laser power, T = 298 K, 60 s accumulation time.
Mentions: Raman spectra of the dry fatty acids capric acid, lauric acid, myristic acid and palmitic acid were obtained using low laser power and are shown in Fig. 1. Characteristic for all fatty acids is the presence of two intense bands between 1050 and 1150 cm−1 arising from ν(C-C) stretch vibrations, one band at 1299 cm−1 arising from γ(CH2) twisting and a group of bands between 1400 and 1500 cm−1 caused by γ(CH2) wagging, γ(CH2) scissoring as well as δ(CH2) and δ(CH3) deformation vibrations (21). Typically carbonyl stretch frequencies of aliphatic compounds are found above 1700 cm−1. The carbonyl stretch frequency ν(C = O) of the fatty acids is red-shifted to 1640 cm−1 and shows only moderate Raman intensity indicating a local environment in which the carbonyl is hydrogen bonded (22). This peak is split into two peaks for lauric, myristic and palmitic acid suggesting heterogeneity of hydrogen bonding in the particle structure.Fig. 1

Bottom Line: To study composition and heterogeneity of insoluble subvisible particles in Mab formulations resulting from degradation of polysorbate 20 and to develop a better understanding of the mechanisms of polysorbate degradation leading to particle formation.Most of the subvisible particles identified were comprised of mixtures of fatty acids with no observable signal from fatty acid esters consistent with hydrolysis being the predominant degradation mechanism leading to particulate formation under these storage conditions.Our methodology is generally applicable for identification of particles in antibody formulations and, in particular, has the potential to give detailed information about particle heterogeneity and insight into mechanistic aspects of particle formation.

View Article: PubMed Central - PubMed

Affiliation: Late Stage Pharmaceutical Development, Genentech Inc., South San Francisco, California, 94080, USA, saggu.miguel@gene.com.

ABSTRACT

Purpose: To study composition and heterogeneity of insoluble subvisible particles in Mab formulations resulting from degradation of polysorbate 20 and to develop a better understanding of the mechanisms of polysorbate degradation leading to particle formation.

Methods: In this study, we exploit the potential of Raman microscopy for chemical identification of particles in monoclonal antibody formulations. Through a combination of experiments and density functional theory (DFT) calculations, we identified unique spectral marker bands for insoluble degradation products of polysorbate 20. We first applied our methodology to identify particles in model systems containing complex mixtures of fatty acids and then to subvisible particles in antibody formulations stored at 5°C for several years.

Results: Most of the subvisible particles identified were comprised of mixtures of fatty acids with no observable signal from fatty acid esters consistent with hydrolysis being the predominant degradation mechanism leading to particulate formation under these storage conditions.

Conclusions: Our methodology is generally applicable for identification of particles in antibody formulations and, in particular, has the potential to give detailed information about particle heterogeneity and insight into mechanistic aspects of particle formation.

No MeSH data available.