Incorporation and release of chemically intact mitomycin C from albumin microspheres: a high performance liquid chromatography evaluation.
Preparation of mitomycin C-loaded human serum albumin (HSA) microspheres using a new technique that avoids the use of heat denaturation, which is known chemically to degrade incorporated drug, is described. This method is based on cross-linking of protein by glutaraldehyde (2.2%) during emulsification (W/O) at room temperature. The resultant particles have a mean (s.d.) diameter of 16.9 (0.34) microns (50% weight average), contain mean (s.d.) 1.15 (0.05%) mitomycin C (MMC) (w/w, n = 17) and maintain sustained release of drug over 20 h. High performance liquid chromatography (HPLC) with diode array detection was used to study the chemical integrity of the drug. Two classes of decomposition products were evaluated: chemical degradation products and drug/nucleophile covalent adducts. The HPLC separation was validated by a number of standards of proposed degradation products. To examine incorporated drug, a complete microsphere system was solubilized with 0.4% trypsin for 24 h, while to examine released drug, microspheres were immobilized on a flow-through glass wool column and fractions were collected. No evidence of significant chemical degradation or covalent coupling to protein was detected in microsphere digests. Two candidate decomposition products, representing approximately 10% of drug released from microspheres (assuming similar molar extinction coefficients to MMC), were identified in column fractions. One of these products appeared to be a covalent adduct, the other possibly an isomeric form of intact MMC. Thus, MMC is predominantly incorporated into and released (90%) chemically intact from HSA microspheres prepared by the technique described.
Allan
L
L
Imperial Cancer Research Fund, Western General Hospital, Edinburgh, UK.
Cummings
J
J
Willmott
N
N
Whateley
T L
TL
Smyth
J F
JF
D002851:Chromatography, High Pressure Liquid; D006801:Humans; D008863:Microspheres; D016685:Mitomycin / Q000008:administration & dosage / Q000737:chemistry*; D012709:Serum Albumin; D013056:Spectrophotometry, Ultraviolet
D012709:Serum Albumin; D016685:Mitomycin