Effects of cryoprotectants on the viability and activity of freeze dried recombinant yeasts as novel oral drug delivery systems assessed by an artificial digestive system.
The aim of this study was to investigate, in a gastric-small intestinal system TIM-1, the effect of cryoprotectants on the survival of freeze-dried Saccharomyces cerevisiae expressing the heterologous P450 73A1 and their ability to convert trans-cinnamic acid into p-coumaric acid. Yeasts were lyophilized in suspensions of trehalose, maltose, lactose, or a milk proteins/trehalose mix. Freeze-dried or native yeasts and trans-cinnamic acid were introduced simultaneously into TIM-1 at the beginning of digestion. Yeast survival rate was evaluated by cell counting in the ileal effluents. P450 73A1 activity was followed by HPLC assay of p-coumaric acid. Freeze-dried yeasts showed high tolerance to digestive conditions. Nevertheless, their survival rate was lower than that of non-dried cells (around 80% whatever the protective agent vs. 96%). The ability of recombinant freeze-dried S. cerevisiae to perform a bioconversion reaction in the digestive tract was shown with all the protectants. The highest trans-cinnamic acid conversion rate (24 vs. 41% for native yeasts) was obtained with the milk proteins/trehalose mix. These results show that freeze-drying might be considered for the pharmaceutical formulation of new drug delivery systems based on orally administered recombinant yeasts and that TIM-1 could be a helpful tool for the pre-screening of oral dosage forms.
Blanquet
Stéphanie
S
Equipe de Recherche Technologique Conception, Ingénierie et Développement de l'Aliment et du Médicament (ERT CIDAM), Centre de Recherche en Nutrition Humaine (CRNH) d'Auvergne, Faculté de Pharmacie, Université d'Auvergne, Clermont-Ferrand, France.
Garrait
Ghislain
G
Beyssac
Erick
E
Perrier
Céline
C
Denis
Sylvain
S
Hébrard
Géraldine
G
Alric
Monique
M
D003451:Cryoprotective Agents / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis*; D016503:Drug Delivery Systems / Q000379:methods*; D005612:Freeze Drying; D041981:Gastrointestinal Tract / Q000378:metabolism*; D005818:Genetic Engineering; D006368:Helianthus / Q000201:enzymology / Q000235:genetics; D006801:Humans; D006899:Mixed Function Oxygenases / Q000096:biosynthesis*; D008954:Models, Biological; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics; D050564:Trans-Cinnamate 4-Monooxygenase
D003451:Cryoprotective Agents; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D050564:Trans-Cinnamate 4-Monooxygenase