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Method for evaluating safety of in-vitro compound and applications thereof

A technology for safety evaluation and compounds, applied in biochemical equipment and methods, animal cells, vertebrate cells, etc., to achieve the effects of simple evaluation methods, avoiding environmental pollution, and low cost

Inactive Publication Date: 2010-05-26
GUANGDONG PHARMA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no LD50 acute toxicity prediction model established by using primary neuron and cardiomyocyte toxicological and pharmacological experimental data, and the application of primary cells has great advantages in drug safety evaluation in vitro

Method used

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  • Method for evaluating safety of in-vitro compound and applications thereof
  • Method for evaluating safety of in-vitro compound and applications thereof
  • Method for evaluating safety of in-vitro compound and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1 primary cell isolation and culture

[0040] 1. Rat primary cortical neuron cell culture

[0041] Aseptically isolate the cerebral cortex of newborn SPF Wistar rats, wash twice with dissection solution, and cut the brain tissue to 1mm with ophthalmic scissors in the dissection solution 3 Size, transfer to a 15ml centrifuge tube, let stand, remove the supernatant, add 0.25% trypsin, digest at 37°C for 5 minutes, suck out the supernatant and transfer it to a 15ml centrifuge tube containing a small amount of calf serum, stop the digestion, repeat several times until Complete digestion. Collect all the liquid and pass through a 180-mesh sieve, then transfer to a centrifuge tube, 100rpm, 7min. Aspirate the supernatant, resuspend the cells in DMEM medium containing 10% fetal bovine serum and 5% horse serum, and adjust the cell density to 5×10 after cell counting. 5 , inoculated in a 96-well plate covered with polylysine (one well of which was placed with a cell sl...

Embodiment 2

[0044] Rat acute oral LD50 and IC50 (primary cortical neuron cells) of embodiment 2 medicine or compound

[0045] Table 1 shows the LD50 (mg / kg) of 17 test compounds once orally administered to rats and the IC50 (mg / ml) values ​​obtained by using drugs or compounds to act on primary cortical neuron cells in this example.

[0046] Rat acute oral LD50 and IC50 of table 117 kinds of drugs or compounds

[0047]

[0048]

[0049] * Rat acute oral LD50 value (mg; kg, derived from chemical safety database).

[0050] It can be seen from the above table that the LD50 of propafenone hydrochloride, chlorpromazine hydrochloride and haloperidol are respectively: propafenone hydrochloride > chlorpromazine hydrochloride > haloperidol, and the above three drugs act on primary cortical nerves respectively. The IC50 after metabolites is: propafenone hydrochloride > chlorpromazine hydrochloride > haloperidol, which is consistent with the order of LD50.

[0051] The LD50 of paroxetine an...

Embodiment 3

[0056] Rat acute oral LD50 and IC50 (primary cardiomyocytes) of embodiment 3 medicine or compound

[0057] Table 2 shows the LD50 (mg / kg) of 20 test compounds once orally administered to rats and the IC50 (mg / ml) values ​​obtained by using drugs or compounds to act on primary cardiomyocytes in this example.

[0058] Rat acute oral LD50 and IC50 of table 220 kinds of drugs or compounds

[0059]

[0060]

[0061] * Rat acute oral LD50 value (mg; kg, derived from chemical safety database).

[0062] It can be seen from the above table that the LD50 of verapamil hydrochloride, chlorpromazine hydrochloride, propafenone hydrochloride, haloperidol, paroxetine hydrochloride, phenobarbital sodium and lidocaine hydrochloride are respectively: propafenone hydrochloride Ketone > phenobarbital sodium > paroxetine hydrochloride > lidocaine hydrochloride > chlorpromazine hydrochloride > haloperidol > verapamil hydrochloride, the IC50 of the above drugs acting on primary cardiomyocytes...

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Abstract

The invention discloses a method for evaluating safety of in-vitro compound. The method is implemented by culturing primary culture cells in separation, co-incubating the compound to the tested and the separately cultured primary culture cells, and testing the relative vitality of the primary culture cells with MTT process. The invention also discloses applications of the above method for evaluating safety of in vitro compound in in-vitro safety evaluation of cosmetics, foods, new medicines or unknown property compounds. The inventive method for evaluating safety of in-vitro compound is easy to implement, short in period, high in test efficiency, low in expenditure and stable and reliable in results, avoids environment pollution and has considerable application prospect.

Description

technical field [0001] The invention relates to the field of pharmacological and toxicological detection, in particular to an in vitro compound safety evaluation method and application thereof. Background technique [0002] For a long time, the preclinical safety evaluation of drugs has relied on animal experiments. However, from an economic point of view, animal experiments are time-consuming and expensive. Animal protectionism has also raised questions about the extensive use of laboratory animals. In June 2007, the European Commission’s legislation on the approval and registration of compounds came into effect. About 30,000 compounds are to be tested. If all animal experiments are used, it is estimated that about 3,900,000 animals will be needed. Therefore, the EU advocates the use of in vitro methods to replace animal experiments. In cosmetics and The skin irritation test has taken the lead in adopting 3R experiments (Refinement, Reduction, Replacement), that is, optim...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/02C12N5/0793C12N5/077
Inventor 臧林泉巫玮潘琴潘雪刁石磊郭姣
Owner GUANGDONG PHARMA UNIV