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Identification and Assessment of Octreotide Acylation in Polyester Microspheres by LC-MS/MS.

Shirangi M, Hennink WE, Somsen GW, van Nostrum CF - Pharm. Res. (2015)

Bottom Line: Release profiles of octreotide from hydrophilic microspheres were compared with that of PLGA microspheres.Nucleophilic attack of the peptide can also occur to the carbamate bond presented in (PC-PEG-PC)-(PL) since 1,4-butanediisocyanate was used as the chain extender.LC-ITMS provided detailed structural information of octreotide modifications via mass analysis of ion fragments.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands.

ABSTRACT

Purpose: Polyesters with hydrophilic domains, i.e., poly(D,L-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA) and a multiblock copolymer of poly(ε-caprolactone)-PEG-poly(ε-caprolactone) and poly(L-lactide) ((PC-PEG-PC)-(PL)) are expected to cause less acylation of encapsulated peptides than fully hydrophobic matrices. Our purpose is to assess the extent and sites of acylation of octreotide loaded in microspheres using tandem mass spectrometry analysis.

Methods: Octreotide loaded microspheres were prepared by a double emulsion solvent evaporation technique. Release profiles of octreotide from hydrophilic microspheres were compared with that of PLGA microspheres. To scrutinize the structural information and localize the actual modification site(s) of octreotide, liquid chromatography ion-trap mass spectrometry (LC-ITMS) was performed on the acylated adducts.

Results: Hydrophilic microspheres showed less acylated adducts in comparison with PLGA microspheres. LC-MS/MS showed that besides the N-terminus and primary amine of lysine, the primary hydroxyl of the end group of octreotide was also subjected to acylation. Nucleophilic attack of the peptide can also occur to the carbamate bond presented in (PC-PEG-PC)-(PL) since 1,4-butanediisocyanate was used as the chain extender.

Conclusions: Hydrophilic polyesters are promising systems for controlled release of peptide because substantially less acylation occurs in microspheres based on these polymers. LC-ITMS provided detailed structural information of octreotide modifications via mass analysis of ion fragments.

No MeSH data available.


Structural formulas of the polymers used in this study: (a) Poly(d,l-lactic-co-glycolic acid) (PLGA 50/50), (b) Poly(d,l-lactic-co-hydroxymethyl glycolic acid) (PLHMGA 82/18), (c) poly(d,l-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA 64/18/18), (d) poly((ε-caprolactone-b-PEG-b-ε-caprolactone)-b-l-lactide)) (PC-PEG-PC)-(PL).
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Fig2: Structural formulas of the polymers used in this study: (a) Poly(d,l-lactic-co-glycolic acid) (PLGA 50/50), (b) Poly(d,l-lactic-co-hydroxymethyl glycolic acid) (PLHMGA 82/18), (c) poly(d,l-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA 64/18/18), (d) poly((ε-caprolactone-b-PEG-b-ε-caprolactone)-b-l-lactide)) (PC-PEG-PC)-(PL).

Mentions: PLGA (acid terminated 5004A with d,l-lactide/glycolide molar ratio 50:50, IV = 0.4 dl/g) was obtained from Purac, The Netherlands. Poly(lactic-co-hydroxymethyl glycolic acid) (PLHMGA) (82/18 lactic/hydroxymethyl glycolic acid ratio (L/HMG) and Mw of 22.5 kg/mol) and poly(lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA) (64/18/18 lactic/glycolic/hydroxymethyl glycolic acid (L/G/HMG) ratio, Mw of 44 kg/mol) were synthesized as described by Leemhuis et al. (23,24). The Polymers are further referred to as PLGA, PLHMGA and PLGHMGA, respectively. The company Innocore (The Netherlands) kindly provided 20CP10C20-LL40 (IV = 0,67 dl/g), which is a multiblock copolymer that was prepared by chain extension of a ABA-triblock copolymer of poly(ε-caprolactone) and 1000 Da PEG with a poly(l-lactide) block with a molecular weight of 4000, using 1,4-butanediisocyanate as a chain extender (26,27). This copolymer is further referred to as (PC-PEG-PC)-(PL). The polymer structures are shown in Fig. 2.Fig. 2


Identification and Assessment of Octreotide Acylation in Polyester Microspheres by LC-MS/MS.

Shirangi M, Hennink WE, Somsen GW, van Nostrum CF - Pharm. Res. (2015)

Structural formulas of the polymers used in this study: (a) Poly(d,l-lactic-co-glycolic acid) (PLGA 50/50), (b) Poly(d,l-lactic-co-hydroxymethyl glycolic acid) (PLHMGA 82/18), (c) poly(d,l-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA 64/18/18), (d) poly((ε-caprolactone-b-PEG-b-ε-caprolactone)-b-l-lactide)) (PC-PEG-PC)-(PL).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4526596&req=5

Fig2: Structural formulas of the polymers used in this study: (a) Poly(d,l-lactic-co-glycolic acid) (PLGA 50/50), (b) Poly(d,l-lactic-co-hydroxymethyl glycolic acid) (PLHMGA 82/18), (c) poly(d,l-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA 64/18/18), (d) poly((ε-caprolactone-b-PEG-b-ε-caprolactone)-b-l-lactide)) (PC-PEG-PC)-(PL).
Mentions: PLGA (acid terminated 5004A with d,l-lactide/glycolide molar ratio 50:50, IV = 0.4 dl/g) was obtained from Purac, The Netherlands. Poly(lactic-co-hydroxymethyl glycolic acid) (PLHMGA) (82/18 lactic/hydroxymethyl glycolic acid ratio (L/HMG) and Mw of 22.5 kg/mol) and poly(lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA) (64/18/18 lactic/glycolic/hydroxymethyl glycolic acid (L/G/HMG) ratio, Mw of 44 kg/mol) were synthesized as described by Leemhuis et al. (23,24). The Polymers are further referred to as PLGA, PLHMGA and PLGHMGA, respectively. The company Innocore (The Netherlands) kindly provided 20CP10C20-LL40 (IV = 0,67 dl/g), which is a multiblock copolymer that was prepared by chain extension of a ABA-triblock copolymer of poly(ε-caprolactone) and 1000 Da PEG with a poly(l-lactide) block with a molecular weight of 4000, using 1,4-butanediisocyanate as a chain extender (26,27). This copolymer is further referred to as (PC-PEG-PC)-(PL). The polymer structures are shown in Fig. 2.Fig. 2

Bottom Line: Release profiles of octreotide from hydrophilic microspheres were compared with that of PLGA microspheres.Nucleophilic attack of the peptide can also occur to the carbamate bond presented in (PC-PEG-PC)-(PL) since 1,4-butanediisocyanate was used as the chain extender.LC-ITMS provided detailed structural information of octreotide modifications via mass analysis of ion fragments.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands.

ABSTRACT

Purpose: Polyesters with hydrophilic domains, i.e., poly(D,L-lactic-co-glycolic-co-hydroxymethyl glycolic acid) (PLGHMGA) and a multiblock copolymer of poly(ε-caprolactone)-PEG-poly(ε-caprolactone) and poly(L-lactide) ((PC-PEG-PC)-(PL)) are expected to cause less acylation of encapsulated peptides than fully hydrophobic matrices. Our purpose is to assess the extent and sites of acylation of octreotide loaded in microspheres using tandem mass spectrometry analysis.

Methods: Octreotide loaded microspheres were prepared by a double emulsion solvent evaporation technique. Release profiles of octreotide from hydrophilic microspheres were compared with that of PLGA microspheres. To scrutinize the structural information and localize the actual modification site(s) of octreotide, liquid chromatography ion-trap mass spectrometry (LC-ITMS) was performed on the acylated adducts.

Results: Hydrophilic microspheres showed less acylated adducts in comparison with PLGA microspheres. LC-MS/MS showed that besides the N-terminus and primary amine of lysine, the primary hydroxyl of the end group of octreotide was also subjected to acylation. Nucleophilic attack of the peptide can also occur to the carbamate bond presented in (PC-PEG-PC)-(PL) since 1,4-butanediisocyanate was used as the chain extender.

Conclusions: Hydrophilic polyesters are promising systems for controlled release of peptide because substantially less acylation occurs in microspheres based on these polymers. LC-ITMS provided detailed structural information of octreotide modifications via mass analysis of ion fragments.

No MeSH data available.