In vitro study of alcohol dehydrogenase and acetaldehyde dehydrogenase encapsulated into human erythrocytes by an electroporation procedure.
The optimal conditions for electroporated/resealed loading of alcohol dehydrogenase (ADH) and/or acetaldehyde dehydrogenase (ALDH) into human erythrocytes were established prior to the study, with the following characteristics: 300 V, 1 ms pulse time, eight pulses every 15 min and 1 h resealing at 37 degreesC. High encapsulation yield and carrier cell recoveries were achieved. Cell volumes increase while hemoglobin contents decrease; in consequence a decrease in cell hemoglobin concentration was observed. A lower hypotonic resistance of loading erythrocytes (throughout osmotic fragility curves) and unaltered oxygen transport capability (as given by oxygen equilibrium curves) were observed. The stability against time (up to 168 h-7 days) of encapsulated individual enzymes, either ADH- or ALDH-red blood cells (RBCs), was studied at 4 degreesC and 37 degreesC, in comparison with that of free enzyme solutions. Both enzymes were released from carrier RBCs to the incubation medium. The stability of carrier RBCs was studied under similar conditions. Non-significant variations in hematological parameters were observed. However, the hemoglobin derivative forms showed modifications. The continuous degradation of ethanol by ADH-RBCs and coencapsulated ADH- and ALDH-RBCs, as a function of time (up to 70 h) suggests the use of these carrier RBCs as agents for complete metabolization of ethanol. The mentioned properties bare the possibility of using ADH and ALDH as carrier systems in in vivo situations.
Lizano
C
C
Dpt. Bioquímica y Biología Molecular, Campus Universitario, Universidad de Alcalá, 28871 Alcalá de Henares, Madrid, Spain. carmen.lizano@alubioqui.alcala.es
Sanz
S
S
Luque
J
J
Pinilla
M
M
D000426:Alcohol Dehydrogenase / Q000737:chemistry* / Q000627:therapeutic use; D000445:Aldehyde Oxidoreductases / Q000737:chemistry* / Q000627:therapeutic use; D019149:Bioreactors; D018274:Electroporation / Q000379:methods*; D004795:Enzyme Stability; D004909:Erythrocyte Indices; D004912:Erythrocytes / Q000737:chemistry / Q000201:enzymology*; D000431:Ethanol / Q000737:chemistry* / Q000378:metabolism; D006454:Hemoglobins / Q000032:analysis; D006801:Humans; D008107:Liver Diseases / Q000517:prevention & control; D009996:Osmotic Fragility
D006454:Hemoglobins; D000431:Ethanol; D000426:Alcohol Dehydrogenase; D000445:Aldehyde Oxidoreductases; C018699:aldehyde dehydrogenase (NAD(P)+)