PMID- 12695064
OWN - NLM
STAT- MEDLINE
DCOM- 20040112
LR  - 20210520
IS  - 0168-3659 (Print)
IS  - 0168-3659 (Linking)
VI  - 89
IP  - 1
DP  - 2003 Apr 14
TI  - Stabilization of alpha-chymotrypsin at the CH2Cl2/water interface and upon 
      water-in-oil-in-water encapsulation in PLGA microspheres.
PG  - 71-85
AB  - Protein inactivation and aggregation are serious drawbacks in the encapsulation 
      of proteins in bioerodible polymers by water-in-oil-in-water (w/o/w) 
      encapsulation. The model protein alpha-chymotrypsin was employed to investigate 
      whether its stabilization towards the major stress factors in the w/o/w 
      encapsulation procedure would allow for the encapsulation and release of 
      non-aggregated and active protein. Due to the formation of amorphous aggregates 
      alpha-chymotrypsin is an excellent sensor to probe unfolding events. Furthermore, 
      its enzymatic activity is highly sensitive towards the presence of organic 
      solvents. alpha-Chymotrypsin in aqueous solution showed substantial aggregation 
      and activity loss when it was homogenized with CH(2)Cl(2) due to adsorption to 
      the interface. Its w/o/w encapsulation in poly(lactic-co-glycolic)acid (PLGA) 
      microspheres caused formation of 35% non-covalent aggregates and reduced the 
      specific activity by 14%. Screening for efficient excipients revealed that 
      co-dissolving the protein with maltose and polyethylene glycol (PEG, M(w) 5000) 
      in the first aqueous phase reduced interface-induced protein aggregation and 
      inactivation. Employing these excipients during encapsulation led to a reduction 
      in alpha-chymotrypsin inactivation (10%) and aggregation (12%). Optimizing the 
      effect of PEG by also dissolving the excipient in the organic phase prior to 
      encapsulation further decreased the amount of non-covalent aggregates to 7% and 
      loss in activity to 5%. The data obtained demonstrate that the w/o emulsification 
      step is the main stress-factor in the w/o/w encapsulation procedure but 
      subsequent encapsulation steps also cause some protein aggregation.
FAU - Perez-Rodriguez, Caroline
AU  - Perez-Rodriguez C
AD  - University of Puerto Rico, Rio Piedras Campus, Department of Chemistry, P.O. Box 
      23346, San Juan, PR 00931-3346, USA.
FAU - Montano, Nashbly
AU  - Montano N
FAU - Gonzalez, Karilys
AU  - Gonzalez K
FAU - Griebenow, Kai
AU  - Griebenow K
LA  - eng
GR  - P20 RR16439-01/RR/NCRR NIH HHS/United States
GR  - S06 GM08102/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - Netherlands
TA  - J Control Release
JT  - Journal of controlled release : official journal of the Controlled Release 
      Society
JID - 8607908
RN  - 0 (Capsules)
RN  - 0 (Emulsifying Agents)
RN  - 0 (Emulsions)
RN  - 0 (Glycolates)
RN  - 0 (Suspensions)
RN  - 059QF0KO0R (Water)
RN  - 1SIA8062RS (Polylactic Acid-Polyglycolic Acid Copolymer)
RN  - 26009-03-0 (Polyglycolic Acid)
RN  - 27432CM55Q (Serum Albumin, Bovine)
RN  - 33X04XA5AT (Lactic Acid)
RN  - 34346-01-5 (Polyglactin 910)
RN  - 3WJQ0SDW1A (Polyethylene Glycols)
RN  - 69-79-4 (Maltose)
RN  - A6R43525YO (Methyl Chloride)
RN  - EC 3.4.21.1 (Chymotrypsin)
RN  - EC 3.4.21.1 (alpha-chymotrypsin)
SB  - IM
MH  - Animals
MH  - Capsules
MH  - Chymotrypsin/*antagonists & inhibitors/*metabolism
MH  - Emulsifying Agents/chemistry
MH  - Emulsions/chemistry
MH  - *Enzyme Stability
MH  - Freeze Drying
MH  - Glycolates/chemistry
MH  - In Vitro Techniques
MH  - Lactic Acid
MH  - Maltose/pharmacokinetics
MH  - Methyl Chloride/*chemistry/pharmacokinetics
MH  - Microspheres
MH  - Polyethylene Glycols/pharmacokinetics
MH  - Polyglactin 910/chemistry
MH  - Polyglycolic Acid
MH  - Polylactic Acid-Polyglycolic Acid Copolymer
MH  - Protein Denaturation/drug effects
MH  - Serum Albumin, Bovine
MH  - Suspensions
MH  - Water
EDAT- 2003/04/16 05:00
MHDA- 2004/01/13 05:00
CRDT- 2003/04/16 05:00
PHST- 2003/04/16 05:00 [pubmed]
PHST- 2004/01/13 05:00 [medline]
PHST- 2003/04/16 05:00 [entrez]
AID - S0168365903000749 [pii]
AID - 10.1016/s0168-3659(03)00074-9 [doi]
PST - ppublish
SO  - J Control Release. 2003 Apr 14;89(1):71-85. doi: 10.1016/s0168-3659(03)00074-9.