Phenylketonuria (PKU) is an inborn error of metabolism characterized by a loss of phenylalanine hydroxylase activity; an enzyme that metabolizes phenylalanine to tyrosine. Phenylalanine ammonia lyase (PAL) is currently being evaluated as a possible therapy for the management of PKU. PAL catalyzes the conversion of phenylalanine to transcinnamic acid (TCA). Our proposed therapy involves encapsulation of PAL enzyme in ethyl cellulose microcapsules using the spray drying method. PAL activity is markedly reduced due to product inhibition. We hypothesized that the addition of albumin to the PAL reaction mixture would bind the TCA and prevent it from inhibiting PAL. In the first phase of our research, we developed an HPLC assay to quantitate phenylalanine and TCA in the presence of albumin. In the second phase, we determined that albumin completely alleviated product inhibition and enhanced PAL activity. Subsequent ultrafiltration studies showed that albumin acted by extensively binding to and sequestering TCA. PAL microcapsules will be taken orally. In the final phase, we studied the activity of encapsulated PAL in simulated GIT conditions to evaluate the ability of microcapsules to protect PAL enzyme against pH and protease mediated degradation. The activity of encapsulated PAL was lower than an equivalent amount of free PAL possibly due to diffusional limitations to the entry of phenylalanine into the microcapsules. Encapsulation of PAL in ethyl cellulose microcapsules did not protect against acidic pH mediated reduction of PAL activity or pepsin mediated proteolytic degradation.
|School:||University of the Sciences in Philadelphia|
|School Location:||United States -- Pennsylvania|
|Source:||MAI 58/01M(E), Masters Abstracts International|
|Subjects:||Biochemistry, Pharmaceutical sciences|
|Keywords:||Albumin, Encapsulation, Oral enzyme substitution therapy, Phenylalanine ammonialyase, Phenylketonuria|
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