ABSTRACT
Objective To establish and validate the Caco-2 cell in vitro absorption model,so as to play a foundation for next study of drug absorption and transport. Methods Caco-2 cells were cultured on the polycarbonate membrane inserts fixed in the 24-well transwell transport chamber with three types cell density of low,medium and high concentration (5×104,1×105,2× 105·mL-1) respectively.After being cultured for 3,6,9,12,15,18,21 d,the integrity of monolayer were assessed and compared by the cell morphology,growth characters and transepithelial electrical resistance (TEER),aiming to determine the inoculum density and culture time.Then the permeability and polarity were validated by the apical-to-basolateral amount of Lucifer Yellow across cell monolayer,the alkaline phosphatase activity in the apical side (AP),the basolateral side (BL) and intracellular activity. Results The cells of low,medium and high concentration group had fusion into a integrate cell monolayer and the maximum absorbance after being cultured for 15,12,9 d respectively.However,conglobated and dead cells were observed at the later growth stage in the medium and high concentration group and the TEER of cell monolayer were smaller than the low concentration group,which could reach 300 Ω·cm2after cultured 15 d and keep a relatively stable value, then cells were cultured with 5×104·mL-1cell density for 21 d.The Lucifer Yellow apparent permeability coefficient (Papp) was 3.57×10-7 cm·s-1which was lower than 5. 0×10-7cm·s-1according to provision. And the intracellular alkaline phosphates’ activity increased,AP/BL increased by 5 times in day 21. Conclusion The integrity,permeability and polarity of the established Caco-2 cell model in our laboratory was validated,and it can be used as an in vitro model similar with small intestinal epithelium for absorption and transport studies.
ABSTRACT
Objective To develop a method for determination of serumal copper ion in rabbit with implantation of copper intrauterine device.Methods At different time points after implantation,the serumal copper ion concentrations were determined by microwave digestion-inductively coupled plasma mass spectrometry(ICP-MS) using scandium(Sc) and indium(In) as internal standards to compensate sample matrix effects,and the pharmacokinetics parameters were calculated in order to reflect copper ion release and metabolic rule.Results The serumal copper ion concentrations were kept at a low and stable level.The recoveries were in the range of 98.7%-113.3%,with the relative standard deviations of less than 5.0%.Conclusion The analytical method is simple,fast,and can be used for the determination of serumal copper ion in rabbit.
ABSTRACT
Objective To study the antipyretic,anti-inflammatory and analgesic effects of Chaigeshubiao granules. Methods Animals were randomly divided into 6 groups:blank control group,indomethacin group,Fengreganmao granules group and the high, the medium and the low dosage groups of Chaigeshubiao granules ( 26, 13, 6. 5 g . kg-1 ) . Each group was administered via intragastric administration once a day for 5 days.Rat model suffering from fever by dried yeast,relieving fever of Chaigeshubiao granules was investigated;Rats with toe swelling by 1% carrageenin and mice with ear swelling by dimethylbenzene were applied to observe anti-inflammatory effects of Chaigeshubiao granules were observed;The pain models induced by 0. 6%acetic acid and the hot-plate tests in mice were used to observe the analgesic effects of Chaigeshubiao granules. Results Compared with negative control group,Chaigeshubiao granules in high dosage could obviously decrease the temperature of rat with fever induced by dried yeast 0.5 h later,and the medium dosage group decreased 4-6 h later. Moreover,Chaigeshubiao granules in high dosage could inhibit inflammatory reaction of rat with toe swelling caused by albumen at 1 h,2 h,3 h,when the medium dosage group inhibited at 2 h,3 h,4 h. The inhibition ratio of the mice with ear swelling induced by dimethylbenzene was 58.2%, 52.0% and 53.9%,respectively at the high,medium and low dose groups. And the inhibition ratio of retortion of mice by 0.6%acetic acid was 50.5%,68.8%,58.1%,respectively, at the high,medium and low dose groups. In addition,the high,medium and low dose groups reduced the pain reaction latency of mice in the hot-plate tests. Conclusion Chaigeshubiao granules have antipyretic,anti-inflammatory and analgesic actions.