The role of citric acid in oral peptide and protein formulations: relationship between calcium chelation and proteolysis inhibition

Søren H Welling; František Hubálek; Jette Jacobsen; David J Brayden; Ulrik L Rahbek; Stephen T Buckley

doi:10.1016/j.ejpb.2013.12.017

The role of citric acid in oral peptide and protein formulations: relationship between calcium chelation and proteolysis inhibition

Søren H Welling, František Hubálek, Jette Jacobsen, David J Brayden, Ulrik L Rahbek, Stephen T Buckley

Department of Drug Design and Pharmacology

34 Citations (Scopus)

Abstract

The excipient citric acid (CA) has been reported to improve oral absorption of peptides by different mechanisms. The balance between its related properties of calcium chelation and permeation enhancement compared to a proteolysis inhibition was examined. A predictive model of CA's calcium chelation activity was developed and verified experimentally using an ion-selective electrode. The effects of CA, its salt (citrate, Cit) and the established permeation enhancer, lauroyl carnitine chloride (LCC) were compared by measuring transepithelial electrical resistance (TEER) and permeability of insulin and FD4 across Caco-2 monolayers and rat small intestinal mucosae mounted in Ussing chambers. Proteolytic degradation of insulin was determined in rat luminal extracts across a range of pH values in the presence of CA. CA's capacity to chelate calcium decreased ~10-fold for each pH unit moving from pH 6 to pH 3. CA was an inferior weak permeation enhancer compared to LCC in both in vitro models using physiological buffers. At pH 4.5 however, degradation of insulin in rat luminal extracts was significantly inhibited in the presence of 10mM CA. The capacity of CA to chelate luminal calcium does not occur significantly at the acidic pH values where it effectively inhibits proteolysis, which is its dominant action in oral peptide formulations. On account of insulin's low basal permeability, inclusion of alternative permeation enhancers is likely to be necessary to achieve sufficient oral bioavailability since this is a weak property of CA.

Original language	English
Journal	European Journal of Pharmaceutics and Biopharmaceutics
Volume	86
Issue number	3
Pages (from-to)	544-51
Number of pages	8
ISSN	0939-6411
DOIs	https://doi.org/10.1016/j.ejpb.2013.12.017
Publication status	Published - Apr 2014

Keywords

Administration, Oral
Animals
Caco-2 Cells
Calcium
Calcium Chelating Agents
Chemistry, Pharmaceutical
Citric Acid
Humans
Insulin
Intestinal Mucosa
Male
Peptides
Proteolysis
Rats
Rats, Wistar
Serum Albumin, Bovine

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The role of citric acid in oral peptide and protein formulations: relationship between calcium chelation and proteolysis inhibition. / Welling, Søren H; Hubálek, František; Jacobsen, Jette et al.
In: European Journal of Pharmaceutics and Biopharmaceutics, Vol. 86, No. 3, 04.2014, p. 544-51.

Research output: Contribution to journal › Journal article › Research › peer-review

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abstract = "The excipient citric acid (CA) has been reported to improve oral absorption of peptides by different mechanisms. The balance between its related properties of calcium chelation and permeation enhancement compared to a proteolysis inhibition was examined. A predictive model of CA's calcium chelation activity was developed and verified experimentally using an ion-selective electrode. The effects of CA, its salt (citrate, Cit) and the established permeation enhancer, lauroyl carnitine chloride (LCC) were compared by measuring transepithelial electrical resistance (TEER) and permeability of insulin and FD4 across Caco-2 monolayers and rat small intestinal mucosae mounted in Ussing chambers. Proteolytic degradation of insulin was determined in rat luminal extracts across a range of pH values in the presence of CA. CA's capacity to chelate calcium decreased ~10-fold for each pH unit moving from pH 6 to pH 3. CA was an inferior weak permeation enhancer compared to LCC in both in vitro models using physiological buffers. At pH 4.5 however, degradation of insulin in rat luminal extracts was significantly inhibited in the presence of 10mM CA. The capacity of CA to chelate luminal calcium does not occur significantly at the acidic pH values where it effectively inhibits proteolysis, which is its dominant action in oral peptide formulations. On account of insulin's low basal permeability, inclusion of alternative permeation enhancers is likely to be necessary to achieve sufficient oral bioavailability since this is a weak property of CA.",

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AU - Welling, Søren H

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AU - Rahbek, Ulrik L

AU - Buckley, Stephen T

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AB - The excipient citric acid (CA) has been reported to improve oral absorption of peptides by different mechanisms. The balance between its related properties of calcium chelation and permeation enhancement compared to a proteolysis inhibition was examined. A predictive model of CA's calcium chelation activity was developed and verified experimentally using an ion-selective electrode. The effects of CA, its salt (citrate, Cit) and the established permeation enhancer, lauroyl carnitine chloride (LCC) were compared by measuring transepithelial electrical resistance (TEER) and permeability of insulin and FD4 across Caco-2 monolayers and rat small intestinal mucosae mounted in Ussing chambers. Proteolytic degradation of insulin was determined in rat luminal extracts across a range of pH values in the presence of CA. CA's capacity to chelate calcium decreased ~10-fold for each pH unit moving from pH 6 to pH 3. CA was an inferior weak permeation enhancer compared to LCC in both in vitro models using physiological buffers. At pH 4.5 however, degradation of insulin in rat luminal extracts was significantly inhibited in the presence of 10mM CA. The capacity of CA to chelate luminal calcium does not occur significantly at the acidic pH values where it effectively inhibits proteolysis, which is its dominant action in oral peptide formulations. On account of insulin's low basal permeability, inclusion of alternative permeation enhancers is likely to be necessary to achieve sufficient oral bioavailability since this is a weak property of CA.

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KW - Caco-2 Cells

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KW - Calcium Chelating Agents

KW - Chemistry, Pharmaceutical

KW - Citric Acid

KW - Humans

KW - Insulin

KW - Intestinal Mucosa

KW - Male

KW - Peptides

KW - Proteolysis

KW - Rats

KW - Rats, Wistar

KW - Serum Albumin, Bovine

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VL - 86

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EP - 551

JO - European Journal of Pharmaceutics and Biopharmaceutics

JF - European Journal of Pharmaceutics and Biopharmaceutics

IS - 3

ER -

The role of citric acid in oral peptide and protein formulations: relationship between calcium chelation and proteolysis inhibition

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