Lactate dehydrogenase (LDH) was entrapped in phosphatidylcholine liposomes to evaluate the protective effect of liposomes against protease digestion. Three different preparations of LDH either encapsulated in liposomes, unencapsulated in liposomes or in the absence of liposomes were incubated with the protease trypsin. The loss of LDH activity was measured at intervals over a 12-hour period. The degradation rate of LDH was found to be the same when LDH was unencapsulated in the presence or absence of liposomes. However, when LDH was entrapped in liposomes the degradation rate was 4 to 24 folds slower. This finding indicates that encapsulation of a protein in a liposome protects the protein from the degrading effects of a protease enzyme.

Drug Carriers↗

PubMedDate not supplied

Loss of LDH activity during membrane filtration.

Membrane filters rendered hydrophilic and composed of biphenyl polycarbonate, polyvinylidene fluoride (PVDF), acrylic copolymer, polysulfone, and mixed esters of cellulose were evaluated to determine which type of filter best can be used for the filtration of lactate dehydrogenase (LDH) solution. Also, the effect of the membrane pore size was evaluated. LDH solution was passed through the filters at a controlled flow rate, after which the filtrate was assayed for LDH activity and protein content using the Bradford method. Polycarbonate and PVDF filters generally showed low loss of protein, except 5 microns PVDF filters. Mixed esters of cellulose, acrylic copolymer, and polysulfone caused considerable loss of protein during passage of the LDH solution through the filter. Interestingly, it was also found that, generally, as the pore size increased the amount of protein loss decreased. However, 5 microns PVDF and mixed esters of cellulose filters showed more loss than their corresponding 0.65 microns and 3 microns pore size filters, respectively. In all cases, more protein was recovered in the filtrate fractions as the volume of LDH solution filtered was increased, suggesting that the mechanism of loss is adsorption and that the magnitude of loss is related to saturation of the matrix polymer surfaces.

Adsorption↗

PubMedDate not supplied

Evaluation of closure integrity after multiple penetrations.

The purpose of this investigation was to simulate in-use testing of closures to determine how many doses could be reasonably withdrawn from a multiple dose vial of a product without compromising the integrity of the closure. Four types of studies were done: 1) a simulated in-use test, 2) a coring test development study, 3) effect of sterilization on coring, and 4) exploratory studies of closure leakage after multiple penetrations. A modified drill press was used for the studies. West 1888 and West 1535 closures were evaluated by the simulated in-use test using a 26G Needle. The results indicated that few particles were generated after 20 penetrations of both closures. There was a marked increase in particles after 30 penetrations. When sterilized closures were tested, it was found that West 1888 generated no particles after 10 insertions with 21G and 18G needles, but there was an increase in the number of particles after 20 insertions. With West 1535, particles were generated after only 10 insertions. Interestingly, autoclaved closures released fewer particles than closures that were not autoclaved. Leakage was observed only from West 850 closures and only under relatively high pressure differential conditions.

Drug Packaging↗

PubMedDate not supplied

Freeze-thaw studies of a model protein, lactate dehydrogenase, in the presence of cryoprotectants.

The objective of this study was to investigate the behavior of lactate dehydrogenase (LDH) upon freezing and thawing, alone or in the presence of several selected cryoprotectants. Also, the influence of the freezing rate on retainment of LDH activity was investigated. It was observed that fast freezing caused less loss of LDH activity than slow freezing. The probable mechanisms of loss of activity after freeze-thaw cycles were discussed. Selected cryoprotectants were evaluated for their ability to protect LDH during freeze-thaw cycles. Surface tension and pH change measurements upon freezing of the cryoprotectant solutions were carried out. Based on the results of these experiments, a potential mechanism of cryoprotection has been developed.

Cryopreservation↗

PubMedDate not supplied

Protection of a model protein, lactate dehydrogenase by encapsulation in liposome.

The objective of this study was to determine if encapsulating a model protein, lactate dehydrogenase (LDH), inside liposomes can protect it during storage, upon freeze-thaw cycles and during freeze-drying. LDH, which undergoes cold denaturation, was entrapped insides liposomes prepared using modification of Bangham's method. It was found that when LDH was entrapped in liposomes and subjected to a single freeze-thaw treatment, there was about 7 times more recovery of activity than when LDH was free in solution. Similarly, LDH encapsulated in liposomes showed 60% recovery after freeze-drying compared to 1-2% recovery for the unentrapped LDH. The role of phospholipid content, residual moisture content and protein content were evaluated. Possible mechanism(s) of cold temperature inactivation of proteins and role of liposomes in preventing such cryodenaturation will be discussed in light of these results.

Drug Compounding↗

PubMedDate not supplied

Extraction rates of marker compounds from rubber closures for parenteral use. II. mechanism of extraction and evaluation of select extraction parameters.

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Diffusion↗