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Drug Content
Packed loaded erythrocytes (0.5 ml) are first deproteinized with acetonitrile
(2.0 ml) and subjected to centrifugation at 2500 rpm for 10 min. The clear
supernatant is analyzed for the drug content by spectroscopy method.
In vitro Drug and Haemoglobin Release
Normal and loaded erythrocytes are incubated at 37± 2°C in phosphate buffer
saline (pH 7.4) at 50% haematocrit in a metabolic rotating wheel incubator
bath. Periodically, the samples are withdrawn with the help of a hypodermic
syringe fitted with a 0.8µ Spectropore membrane filter.
Percent haemoglobin can similarly be calculated at various time intervals at
540 run spectrophotometrically.
Osmotic Fragility
When red blood cells are exposed to solutions of varying tonicities their
shape changes (swell in hypotonic and shrink in hypertonic
environments) due to osmotic imbalance. Assayed for Hb and/or drug
release.
Osmotic Shock
Osmotic shock describes a sudden exposure of drug loaded
erythrocytes to an environment, which is far from isotonic to evaluate
the ability of resealed erythrocytes to withstand the stress and
maintain their integrity as well as appearance.
Turbulence Shock
1. The parameter indicates the effects of shear force and pressure by
which resealed erythrocytes formulations are injected, on the
integrity of the loaded cells.
2. Loaded erythrocytes (10% haematocrit, 5 ml) are passed through a
23-gauge hypodermic needle at a flow rate of 10 ml/min . After every
pass, aliquote of the suspension is withdrawn and centrifuged for 15
min, and haemoglobin content, leached out are estimated
spectrophotometrically.
Morphology and Percent Cellular Recovery
Phase-contrast optical microscopy, transmission electron
microscopy and scanning electron microscopy are the microscopic
methods used to evaluate the shape, size and the surface features
of the loaded erythrocytes.
The most common storage media include Hank’s balanced salt solution and
acid–citrate–dextrose at 4° C.
Cells remain viable in terms of their physiologic and carrier characteristics for
at least 2 weeks at this temperature.
The addition of calcium-chelating agents or the purine nucleosides improve
circulation survival time of cells upon reinjection.
Exposure of resealed erythrocytes to membrane stabilizing agents such as
dimethyl sulfoxide, dimethyl,3,3-di-thio-bispropionamide, gluteraldehyde,
toluene-2-4-diisocyanate followed by lyophilization or sintered glass filtration
has been reported to enhance their stability upon storage.
The various mechanisms proposed for drug release include:
Passive diffusion.
Specialized membrane associated carrier transport.
Phagocytosis of resealed cells by macrophages of RES, subsequent
accumulation of drug into the macrophage interior, followed by slow
release.
Accumulation of erythrocytes in lymph nodes upon subcutaneous
administration followed by hemolysis to release the drug.
Erythrocytes as carrier for enzymes.
Erythrocytes as carrier for drugs.
Erythrocytes for drug targeting.
1. Drug targeting to reticuloendothelial system.
2. Drug targeting to liver.
-Treatment of liver tumors.
-Treatment of parasitic diseases.
-Removal of RES iron overload.
-Removal of toxic agents.
Enzyme Deficiency/Replacement Therapy: Gaucher’s disease
(glucocerebrosidase), replacement of enzyme in lysosomes
(glucuronidase, galactosidase, glucosidase)
Treatment of Liver Tumours
Treatment of Parasitic diseases
Removal of Toxic Agents : enzyme to hydrolyze organophosphorous
compounds.
The damaged erythrocytes are quickly removed from circulation by
phagocytic Kupffer cells located in liver and spleen.
Chemically modified RBC can be targeted to organs of the MPS.
1. Surface Modification with Antibodies.
2. Surface Modification with Glutaraldehyde.
3. Surface Modification-involving Carbohydrates .
4. Surface Modification with Sulphydryls.
5. Surface chemical cross-linking.
Delivery of Antiviral Agents
Delivery of Azidothymidine Derivative
Delivery of Deoxycytidine Derivatives
Macrophage Activation
Thrombolytic Therapy
Oxygen Deficiency Therapy
Delivery of Interleukins
Nanoerythrosomes
Extrusion of RBC ghosts to produce small vesicles having
an average diameter of 100nm.
Erythrosomes
Specially engineered vesicular systems in which
chemically cross linked human erythrocyte cytoskeletons
are used as a support upon which a lipid bilayer is
coated.
Jain.S., Jain.N.K., resealed erythrocytes as drug carriers, Edited Jain
N.K., Controlled And Novel Drug Delivery, New Delhi, CBS
publishers, New Delhi, 2004, 256-281.
Vyas S.P., Khar R.K., Targeted And Controlled Drug Delivery: Novel
Carrier Systems, New Delhi, CBS publisher, 2004, 387-413.
Indian Journal of Pharmaceutical Education & Research Vol. 43(4),
Oct-Dec, 2009 , 375-386.
Journal of Controlled Release 95 (2004) 27– 49.
Evaluation n application of resealed erythrocytes

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Evaluation n application of resealed erythrocytes

  • 1.
  • 2.
  • 3. Drug Content Packed loaded erythrocytes (0.5 ml) are first deproteinized with acetonitrile (2.0 ml) and subjected to centrifugation at 2500 rpm for 10 min. The clear supernatant is analyzed for the drug content by spectroscopy method. In vitro Drug and Haemoglobin Release Normal and loaded erythrocytes are incubated at 37± 2°C in phosphate buffer saline (pH 7.4) at 50% haematocrit in a metabolic rotating wheel incubator bath. Periodically, the samples are withdrawn with the help of a hypodermic syringe fitted with a 0.8µ Spectropore membrane filter. Percent haemoglobin can similarly be calculated at various time intervals at 540 run spectrophotometrically.
  • 4. Osmotic Fragility When red blood cells are exposed to solutions of varying tonicities their shape changes (swell in hypotonic and shrink in hypertonic environments) due to osmotic imbalance. Assayed for Hb and/or drug release. Osmotic Shock Osmotic shock describes a sudden exposure of drug loaded erythrocytes to an environment, which is far from isotonic to evaluate the ability of resealed erythrocytes to withstand the stress and maintain their integrity as well as appearance.
  • 5. Turbulence Shock 1. The parameter indicates the effects of shear force and pressure by which resealed erythrocytes formulations are injected, on the integrity of the loaded cells. 2. Loaded erythrocytes (10% haematocrit, 5 ml) are passed through a 23-gauge hypodermic needle at a flow rate of 10 ml/min . After every pass, aliquote of the suspension is withdrawn and centrifuged for 15 min, and haemoglobin content, leached out are estimated spectrophotometrically.
  • 6. Morphology and Percent Cellular Recovery Phase-contrast optical microscopy, transmission electron microscopy and scanning electron microscopy are the microscopic methods used to evaluate the shape, size and the surface features of the loaded erythrocytes.
  • 7.
  • 8.
  • 9. The most common storage media include Hank’s balanced salt solution and acid–citrate–dextrose at 4° C. Cells remain viable in terms of their physiologic and carrier characteristics for at least 2 weeks at this temperature. The addition of calcium-chelating agents or the purine nucleosides improve circulation survival time of cells upon reinjection. Exposure of resealed erythrocytes to membrane stabilizing agents such as dimethyl sulfoxide, dimethyl,3,3-di-thio-bispropionamide, gluteraldehyde, toluene-2-4-diisocyanate followed by lyophilization or sintered glass filtration has been reported to enhance their stability upon storage.
  • 10. The various mechanisms proposed for drug release include: Passive diffusion. Specialized membrane associated carrier transport. Phagocytosis of resealed cells by macrophages of RES, subsequent accumulation of drug into the macrophage interior, followed by slow release. Accumulation of erythrocytes in lymph nodes upon subcutaneous administration followed by hemolysis to release the drug.
  • 11. Erythrocytes as carrier for enzymes. Erythrocytes as carrier for drugs. Erythrocytes for drug targeting. 1. Drug targeting to reticuloendothelial system. 2. Drug targeting to liver. -Treatment of liver tumors. -Treatment of parasitic diseases. -Removal of RES iron overload. -Removal of toxic agents.
  • 12. Enzyme Deficiency/Replacement Therapy: Gaucher’s disease (glucocerebrosidase), replacement of enzyme in lysosomes (glucuronidase, galactosidase, glucosidase) Treatment of Liver Tumours Treatment of Parasitic diseases Removal of Toxic Agents : enzyme to hydrolyze organophosphorous compounds.
  • 13. The damaged erythrocytes are quickly removed from circulation by phagocytic Kupffer cells located in liver and spleen. Chemically modified RBC can be targeted to organs of the MPS. 1. Surface Modification with Antibodies. 2. Surface Modification with Glutaraldehyde. 3. Surface Modification-involving Carbohydrates . 4. Surface Modification with Sulphydryls. 5. Surface chemical cross-linking.
  • 14. Delivery of Antiviral Agents Delivery of Azidothymidine Derivative Delivery of Deoxycytidine Derivatives Macrophage Activation Thrombolytic Therapy Oxygen Deficiency Therapy Delivery of Interleukins
  • 15. Nanoerythrosomes Extrusion of RBC ghosts to produce small vesicles having an average diameter of 100nm. Erythrosomes Specially engineered vesicular systems in which chemically cross linked human erythrocyte cytoskeletons are used as a support upon which a lipid bilayer is coated.
  • 16. Jain.S., Jain.N.K., resealed erythrocytes as drug carriers, Edited Jain N.K., Controlled And Novel Drug Delivery, New Delhi, CBS publishers, New Delhi, 2004, 256-281. Vyas S.P., Khar R.K., Targeted And Controlled Drug Delivery: Novel Carrier Systems, New Delhi, CBS publisher, 2004, 387-413. Indian Journal of Pharmaceutical Education & Research Vol. 43(4), Oct-Dec, 2009 , 375-386. Journal of Controlled Release 95 (2004) 27– 49.