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M. Anitha Sri,Y11MPH448, I/II M. Pharmacy, II Semester, Industrial Pharmacy, Chalapathi Institute Of Pharmaceutical Sciences.

In-vitro Characterisation

3. 4. 5. 6. 7. 8. 9. 10.

Drug Content In-vitro drug and Hb release Osmotic Fragility Osmotic Shock Turbulence Shock Morphology % Cell Recovery Entrapped Magnetite ESR ZSR

5mL) Deproteinised with Acetonitrile (2mL) Centrifuged at 2500-3000 rpm for 10 min. Clear supernatant is analyzed for drug content .Drug Content: Packed loaded cells (0.

.Retaining the entrapped magnetite: Magnet placed adjacent to the base of centrifuge tube.

In-vitro Drug Content Incubation Periodic withdrawl of samples Deproteinisation with Acetonitrile Centrifugation Supernatant was assayed for drug .

4) at 50% haematocrit in a metabolic rotating wheel incubator.Incubation of loaded normocytes at 37 0C in PBS (pH 7. Metabolic Shaker Incubator .

0.8 µ spectropore membrane filter is fitted to a hypodermic syringe to withdraw the samples. Hypodermic syringe with a membrane filter. .

45 µ filter Deproteinisation with methanol Centrifuge Supernatant assayed for Hb at 540 nm .In-vitro Hb release: 5 % Hematocrit in PBS Withdrawl of samples using 0.

Osmotic Fragility Exposing to solutions of varying tonicities Isotonic to hypotonic Centrifuge Supernatant assayed for drug and Hb Based on resistance of cells to haemolysis .

Change of shape due to osmotic imbalance Osmotic Fragility .

Osmotic Shock Sudden exposure to non-isotonic environment Incubation of resealed erythrocytes Centrifugation Supernatant assayed for Hb release .

Turbulence shock Measure of simulating destruction of loaded erythrocytes during injection Loaded cells passed through hypodermic needle Flow rate – 10 mL/min Withdrawl of suspensions Centrifuge Supernatant assayed for Hb .

Morphology & % cell recovery Phase contrast optical microscopy Transmission electron microscopy Scanning electron microscopy Haemocytometer .

Magnetite concentration Magnetite loaded cells treated with HCl Heated at 60 0 C for 2 hrs + 20% w/v trichloroacetic acid Centrifuge Supernatant assayed for magnetite .

ESR & ZSR ESR is the estimation of suspension stability in plasma Depends on the no. and size of RBC and other plasma proteins Normal Blood ESR: 0 to 15 mm/hr ZSR is a measure of the closeness with which RBC will approach each other .

NOVEL APPROACHES Erythrosomes Nanoerythrosomes Biotinylated erythrocytes .

.Erythrosomes:  Erythrosomes are specially engineered vesicular systems in which chemically cross-linked human erythrocyte cytoskeletons are used as a support upon which a lipid bilayer (phosphatidyl choline)is coated.

Nanoerythrosomes Normal erythrocytes Depletion of Hb Preparation of erythrocyte ghost Extrusion / Sonication / Electrical Breakdown Nanoerythrosomes .

2.  Methods: 1. Using spacer arm.  .Biotinylated erythrocytes Biotin molecules are attached to the amino groups present on the RBC. Direct interaction.

Jain. R. K. Jain. 256 to 291. 2. Controlled and Novel Drug Delivery. Pg No. N. Pg No. Jain. Vyas. K. Advances in controlled and Novel Drug Delivery. Novel carrier systems. Progress in controlled and Novel Drug Delivery Systems. Pg No. K. Pg No. 4. 387 to 416. K. 332 to 360. 3. N. . N.P. Targeted and controlled drug delivery. Khar.References: 1. 387 to 416. S.