Experiments for the determination of blood concentration and half-life of drugs
Measurement of blood concentration and half-life of drugs is used (1) in order to guide the clinical selection of the optimal therapeutic regimen and the most appropriate therapeutic dosage for different individuals; (2) to improve the clinical drug efficacy; and (3) to reduce the clinical adverse drug reactions.
Operation method
colorimetric method
Principle
Phenolphthalein (phenolsufon phthaleinl, psp), known by the trade name of phenol red (molecular weight 354.38), is a commonly used indicator, which is purplish-red in alkaline environments. After intravenous injection of psp, because it is not metabolized in the body, it is not easy to enter the tissue through the capillaries, so we can use colorimetric method to determine the absorbance of plasma psp at different times after the administration of the drug, and calculate the concentration of plasma psp through the external labeling method; and because of psp to meet the elimination kinetics of the first level of the body, so by the time of the administration of the drug and the corresponding concentration of the blood drug, the calculation of the drug half-life of the apparent volume of distribution.
Materials and Instruments
Rabbit Move I. Plotting the standard curve of phenol red Caveat ⒈ before taking blood, the rabbit ear part of the hair should be removed, so as not to affect blood coagulation.Peck before taking blood, the rabbit ear ear margin vein should be fully expanded and filled to facilitate blood collection.3) When centrifuging, pay attention to the balance of the test tube, so as not to damage the centrifuge.Singed prohibit touching the smooth surface of the cuvette with hands, if the solution flows out, can only be wiped with mirror paper to avoid glossy surface. Common Problems For more product details, please visit Aladdin Scientific website.
PSP solution NaCl solution NaOH solution Ethanol
Rabbit Box Scalpel Anticoagulation Tube Test Tube Rack Pipettes Ear Wash Balls Microsampler Syringe No. 6 Needle Surgical Lights Artery Clamps Mirror Wipes Cotton Balls Benchtop Centrifuge Model 721 Spectrophotometer Infant Pound Scale
Take 1, 2, 4, 8, 16 umol-Lˉ1PSP standard solution and distilled water 1.55 ml each, and then add 1 mol-Lˉ1NaOH solution 0.05 ml uniform. Determine the absorbance of the standard solution at 560 nm colorimetrically and plot the standard curve of phenol red, or use a calculator or different standard concentrations of phenol red and its corresponding absorbance for linear regression, the linear regression equation of the standard curve can be obtained:Y=a+bX X: standard concentration of phenol red; Y: absorbance (A)
II. Determination of phenol red concentration in blood at different times after drug administration
A healthy rabbit was taken, weighed and placed in a rabbit box. 1 ml of blood was taken from one ear vein and placed in a dry test tube containing heparin and shaken for blank control, then 6 mg/kg of 0.6% phenol red solution was injected into the ear vein of the same side, and 1 ml of blood was taken from the other ear vein and placed in a test tube containing heparin and shaken at 5, 10, 20 and 30 min after the administration of the drug respectively. The blood was centrifuged for 10 min at 1500 r/min. 0.1 ml of the supernatant was placed in 5 test tubes, and 1.5 ml of diluent was added to each tube, shaken well and left for 5 min. The absorbance was measured at 560 nm by colorimetric measurement with a 721 spectrophotometer.
Results
The plasma concentration of phenol red at different times was calculated by using a calculator or computer to call the linear regression program (attached):
1. Phenol red standard curve (concentration-absorbance relationship curve).
2. Using the above standard curve, find the concentration value corresponding to different absorbance, multiply this concentration value by 16 to find the plasma concentration of phenol red at different times after the administration of the drug.
3. According to the kinetic equation for primary elimination ㏒ C1= ㏒ C0-Kt/2.303
The slope (-K/2.303) and intercept (㏒ C0) of this regression equation can be obtained by linearly regressing the time of administration, t, on the logarithmic phytochemicals of the different psp plasma concentrations that have been obtained, ㏒ C1. By bringing K and C0 into the equations t1/2 = 0.693/K and V4 = D/ C0 (D is the administered dose), t1/2 and Vd can be obtained.