CN112545991A

CN112545991A - Andrographolide solution capable of being self-emulsified and preparation method thereof

Info

Publication number: CN112545991A
Application number: CN202110032599.7A
Authority: CN
Inventors: 陈建新; 李江北; 曾振灵; 黄显会; 武力; 刘小凡; 杨霞
Original assignee: South China Agricultural University
Current assignee: South China Agricultural University
Priority date: 2021-01-08
Filing date: 2021-01-08
Publication date: 2021-03-26

Abstract

The invention discloses a stable andrographolide solution capable of self-emulsifying and a preparation method thereof. The andrographolide solution is composed of andrographolide, surfactant, cosurfactant and vegetable oil. The surfactant is a nonionic liquid surfactant with the HLB value of 10-15, and the cosurfactant is one or a mixture of two of polyethylene glycol 400, ethanol, propylene glycol, diethylene glycol monoethyl ether and dimethyl sulfoxide (DMSO). The content of andrographolide solution medicine is in the range of 0.1-5.0%, the andrographolide solution medicine can be self-emulsified into O/W type microemulsion after water is added, and the grain diameter of the medicine-containing liquid drop after emulsification is less than 1 micron. The andrographolide solution can obviously improve the solubility and bioavailability of andrographolide. The preparation method is simple in preparation process, the used auxiliary materials are safe and non-toxic, industrialization is easy to realize, and the preparation method has a good application prospect.

Description

Andrographolide solution capable of being self-emulsified and preparation method thereof

Technical Field

The invention belongs to the field of medicines and veterinary medicines, relates to a novel anti-inflammatory and anti-infective medicament, and particularly relates to an andrographolide oil solution capable of being self-emulsified into an oil-in-water microemulsion and a preparation method thereof.

Background

The common andrographis herb is a heat-clearing and detoxifying traditional Chinese medicine commonly used in traditional Chinese medicine and veterinary medicine, and is commonly used for treating various infectious diseases clinically, including traumatic infection, furuncle, carbuncle, erysipelas, upper respiratory tract infection, acute and chronic tonsillitis, acute and chronic pharyngolaryngitis, acute and chronic bronchitis, acute bacillary dysentery, acute gastroenteritis and the like. Andrographolide is the main effective component of herba Andrographitis, and belongs to diterpene lactone compounds. Modern pharmacological research shows that andrographolide has pharmacological effects of resisting inflammation, regulating immunity, resisting virus, resisting bacteria, treating cardiovascular diseases, protecting liver, etc. In clinical medicine, andrographolide is widely used in treating upper respiratory tract infection, bacillary dysentery and other diseases, has definite curative effect and is known as natural antibiotic. The clinical andrographolide preparation forms comprise tablets, dripping pills, capsules and the like. However, andrographolide has very poor water solubility, and its solubility in water at room temperature is only 0.07mg/mL (Chenling, King Shaohua). Fundamental physicochemical property of andrographolide is examined, pharmaceutical today, 2010, vol.20 (1): 41-43 ]; the Bioavailability of andrographolide in rats orally administered by Chen et al is only 1.19% [ Haw-Wen Chen, Chin-Shiu Huang, Chien-Chun Li et al, Bioavailability of andrographolide and protection against carbon tetrachloride-induced oxidative metabolism in rats, oxygenation and Applied Pharmacology, 2014, 280: 1-9], the extremely low dissolution rate and bioavailability severely restrict the clinical efficacy of andrographolide.

The bioavailability of the insoluble drug can be improved by adopting various pharmaceutical technologies, such as preparation technologies of forming soluble salts, micronizing or nanocrystallizing to increase the surface area, emulsifying and microemulsifying, solid dispersion taking a water-soluble surfactant as a carrier, cyclodextrin molecule inclusion and the like. The soluble salt prepared from andrographolide as raw material is in various forms, including Xiyanping, Yanhuning and Chuanhuning for clinical use. Although the andrographolide salt injection has the characteristics of fast distribution, wide distribution and the like, along with the increasingly wide clinical application of the andrographolide salt injection, the clinical compliance is poor, the adverse reaction reports are increased day by day, and the safety of the injection is concerned (Hexianmin, Cheng nationality seal. adverse reaction analysis of different salts of andrographolide injection. Chinese medicine industry, 2011, 20 (19): 60]. In addition, veterinary clinical administration by drinking water and mixing is a more convenient way, and injection administration has high application cost and low efficiency. In order to improve the dissolution rate and bioavailability of andrographolide, several patent applications relate to the preparation of andrographolide emulsions, suspensions and nano-emulsions, and although the bioavailability of andrographolide is improved to a certain extent, the process is complicated, the requirements on equipment are high, the drug-loading rate of the preparation is low, the stability is poor, and the clinical application of the preparation is limited.

The microemulsion is a novel medicinal preparation formed by water, oil, surfactant, cosurfactant and the like. The nano-scale microemulsion (nanoemulsion) can directly permeate intestinal mucosa to enter a blood circulation system, so that the microemulsion can not only obviously improve the solubility of a water-insoluble medicament, but also obviously improve the bioavailability of the water-insoluble medicament. However, complete microemulsion solutions tend to have low drug content, may break emulsions during storage and transportation, and may degrade unstable drugs in water during storage. In order to overcome the problems, the invention provides an andrographolide solution capable of self-emulsifying, and water is added during clinical use to form microemulsion spontaneously. The solution provided by the invention has high drug content and stable performance.

Disclosure of Invention

The invention aims to overcome the defects and shortcomings of the prior art and provide an andrographolide solution which has high effective concentration, good stability, high bioavailability and simple preparation process and can be self-emulsified into microemulsion in water.

The inventor systematically evaluates the solubility of various cosolvents, vegetable oils and surfactants in andrographolide, screens out the cosolvents, surfactants and oils with higher solubility, and optimizes the formula of the self-emulsifiable andrographolide solution by applying a ternary phase diagram method to obtain an optimized formula and an optimal preparation process. The preparation is subjected to comprehensive performance and pharmacodynamic evaluation, including drug loading, stability, particle size distribution and concentration after self-emulsification, anti-inflammatory efficacy, pharmacokinetic parameters and the like. Aims to provide an andrographolide solution preparation which has stable performance, high bioavailability, good drug effect and simple and convenient process and is suitable for industrial production.

The technical scheme for realizing the aim of the invention is to prepare an andrographolide solution capable of self-emulsifying to form microemulsion, wherein the solution consists of andrographolide, surfactant, cosurfactant and vegetable oil. The mass percentages of the components are as follows: 0.1-5.0% of andrographolide, 25-75% of surfactant, 10-50% of cosurfactant and 10-35% of vegetable oil.

The surfactant is a nonionic liquid surfactant with the HLB value of 10-15, and the selected surfactant is one or a mixture of two of polyethylene glycol fatty acid ester (including polyethylene glycol 400 monooleate, polyethylene glycol 600 monooleate, polyethylene glycol 400 monolaurate and caprylic/capric polyethylene glycol glyceride), polyoxyethylene 40 hydrogenated castor oil, polyethylene glycol vitamin E succinate, lecithin, soybean lecithin and Tween 80;

the cosurfactant is one or a mixture of two of polyethylene glycol 400, ethanol, 1, 2-propylene glycol, 1, 3-propylene glycol, diethylene glycol monoethyl ether and dimethyl sulfoxide (DMSO);

the vegetable oil is one or mixture of two of olive oil, camellia oil and isopropyl myristate.

The prescription of the andrographolide solution provided by the invention comprises the types and the proportions of a surfactant, a cosurfactant and vegetable oil, which are determined by finding out a micro-emulsion or nano-emulsion area through a ternary phase diagram method. Selecting one or more oils according to the HLB value of the surfactant phase, changing the ratio of the surfactant to the oils according to the ratio of 9: 1-1: 9, adding a certain amount of andrographolide solution dissolved in cosurfactant, slowly adding distilled water at room temperature, and fully stirring until an O/W type pale yellow or colorless andrographolide microemulsion solution with good fluidity is formed.

Another object of the present invention is to provide a method for preparing a self-emulsifiable andrographolide solution, comprising the following steps in sequence: (1) weighing a certain amount of andrographolide, adding a cosurfactant, and heating to dissolve at 60-90 ℃; (2) adding a surfactant into the solution, and fully stirring to make the solution uniform; (3) then adding vegetable oil into the above solution, stirring thoroughly to make it uniform, to obtain self-emulsifiable andrographolide solution; (4) the obtained solution is subpackaged into penicillin bottles.

The andrographolide solution provided by the invention is a light yellow transparent or semitransparent solution, water is added according to the volume ratio of 1: 50-1: 1000, and the diameter of the oily liquid drop containing the medicine is measured by a nanometer particle size analyzer to be within the range of 10-600 nm. Compared with the prior art, the andrographolide solution has the following advantages:

(1) effectively overcomes the defects of poor water solubility, poor in vitro dissolution and low in vivo bioavailability of the andrographolide, and obviously improves the anti-inflammatory and anti-infection drug effects of the andrographolide.

(2) The medicine content is high, different use concentrations can be prepared according to different diseases and treatment purposes clinically, and the medicine packaging and transportation cost is greatly saved.

(3) The selected drug carrier is transparent or semitransparent solution prepared from vegetable oil, cosurfactant and nonionic surfactant. The solution has high thermodynamic stability, and the medicinal components are not easy to deteriorate. The present invention selects non-ionic surfactant with no toxicity and good biocompatibility, is not easy to be affected by strong electrolyte and inorganic salt, and has small hemolytic effect.

(4) The andrographolide solution provided by the invention has the advantages of simple preparation process and simple requirements on equipment, high-speed stirring and shearing or high-pressure homogenization are not needed, and the heating temperature is not more than 100 ℃.

Drawings

FIG. 1 is a graph showing the particle size measurement of an emulsion formed by adding water to a solution prepared in example 1;

FIG. 2 is a graph showing the particle size measurement of an emulsion formed by adding water to the solution prepared in example 2;

FIG. 3 is a graph showing the particle size measurement of an emulsion formed by adding water to the solution prepared in example 3;

FIG. 4 is the survival curve of LPS-inoculated mice inoculated with either the perigastric andrographolide suspension or solution in example 7.

Detailed Description

The following detailed description of the embodiments and effects of the present invention is given by way of specific examples, which are intended to illustrate the present invention and should not be construed as limiting the scope of the claims of the present invention.

Example 1 preparation of a solution of Andrographolide I

Weighing 2.0g of andrographolide, adding 50ml of 1.2-propylene glycol, heating and dissolving in a water bath at 80-90 ℃, adding 20ml of polyethylene glycol 600 monooleate, 20ml of polyoxyethylene 40 hydrogenated castor oil and 10ml of isopropyl myristate, and fully stirring to obtain an andrographolide solution of formula I, wherein the content of andrographolide in the solution is 2.0%.

Example 2 preparation of formula II Andrographolide solution

Weighing 2.0g of andrographolide, adding 30ml of diethylene glycol monoethyl ether, heating and dissolving in a water bath at 80-90 ℃, adding 30ml of polyethylene glycol 600 monooleate, 10ml of soybean phospholipid and 10ml of olive oil, and fully stirring to obtain an andrographolide solution of formula II, wherein the content of andrographolide in the solution is 2.5%.

EXAMPLE 3 preparation of prescription III Andrographolide solution

Weighing 2.0g of andrographolide, adding 8ml of DMSO and 40012ml of polyethylene glycol, heating and dissolving in 80-90 ℃ water bath, adding 30ml of polyethylene glycol 600 monooleate and 5ml of camellia oil, and stirring completely to obtain andrographolide solution with andrographolide content of 3.6% in the formula III.

EXAMPLE 4 determination of the particle size of emulsion after emulsification of Andrographolide solution with Water

1ml of the andrographolide solution prepared in example 1 was added to 100ml of distilled water, 0.8ml of the andrographolide solution prepared in example 2 was added to 100ml of distilled water, 0.56ml of the andrographolide solution prepared in example 3 was added to 100ml of distilled water, shaking was performed, and 200ppm of emulsion was prepared, wherein the emulsion prepared from the andrographolide solutions of formula I and formula II was translucent, and the emulsion prepared from the andrographolide solution of formula III was colorless and transparent. The particle sizes of the three emulsion droplets are measured by adopting a Zetasizer Nano-ZS90 nanometer particle size analyzer of British Markov company, and the particle sizes of the emulsion droplets formed by adding water into formulas I, II and III are respectively in the ranges of 120-450 nm, 150-400 nm and 10-30 nm, as shown in the graph from 1 to 3.

EXAMPLE 5 determination of the stability of Andrographolide solution

The stability of the andrographolide solution is inspected by measuring the change of the drug content of the andrographolide solution after being placed for different time and the change of the droplet size of the nanoemulsion formed by adding water into the solution. Taking 1ml, 0.8ml and 0.56ml of andrographolide solutions of formulas I, II and III, respectively, standing for different time (0 day, 90 days and 180 days), adding methanol to constant volume to 100ml, and measuring the content of andrographolide in the diluent at each time point by using a High Performance Liquid Chromatography (HPLC), wherein the detection wavelength of an HPLC ultraviolet detector is 224 nm. The external standard method is adopted for quantification, and the drug contents of the three prescription andrographolide solutions at each time point are shown in table 1.1 ml, 0.8ml and 0.56ml of andrographolide solutions of formulas I, II and III, which were placed for different periods of time (0 day, 90 days and 180 days), were taken, 100ml of distilled water was added, shaken well, and the particle size distribution of emulsion droplets was measured with a nanometer particle size analyzer, the results are shown in Table 1.

TABLE 1 determination of the drop size distribution of andrographolide solutions after standing for different days and emulsion formation with water

The results in the table show that the andrographolide solutions of formula I, II and III have a 20%, 16% and 8.3% decrease after 180 days at room temperature, and the stability of formula III is better than that of formula I and formula II. After the andrographolide solutions in the formulas I, II and III are placed at room temperature for 90 days and 180 days, the andrographolide solutions can still be self-emulsified into microemulsion, and the particle size of the microemulsion is slightly increased compared with that of the microemulsion on the preparation day.

Example 6 evaluation of bioavailability of Andrographolide solution by oral administration

The bioavailability of the andrographolide solution of formula III prepared in example 3 in rats was examined using andrographolide drug as a control group. The method is to divide 12 male SD rats into two groups at random, each group comprises 6 male SD rats, andrographolide bulk drug (dispersion liquid) dispersed in 0.5% sodium carboxymethylcellulose solution and andrographolide solution of formula III diluted by 3 times with distilled water at one time, wherein the dose of intragastric administration is 100 mg/kg. Blood was periodically collected from the orbit, and after treatment, the plasma concentration of andrographolide was measured by LC-MS method, and pharmacokinetic parameters were calculated, with the results shown in table 2. The results in Table 2 show that the bioavailability of the andrographolide solution of formula III is 5 times higher than that of the andrographolide material.

TABLE 2 oral bioavailability test results of andrographolide dispersion and solution rats

Example 7 evaluation of anti-inflammatory Activity of Andrographolide solution by oral administration

The number of Kunming mice at 6 weeks is 60, half male and half female, and the Kunming mice are randomly divided into 6 groups, 10 mice in each group, and half male and female. In the other 5 groups, except for the normal control group (Mock), each group was intraperitoneally injected with 0.2mL of a physiological saline solution containing 10mg of bacterial Lipopolysaccharide (LPS). Except for the LPS control group (LPS control), which was intragastrically administered with 0.2mL of physiological saline 30min before LPS inoculation, the other 4 groups were intragastrically administered with high and low dose of andrographolide dispersion (dispersed in 0.5% carboxymethylcellulose water solution) or formula III andrographolide solution diluted 3 times with distilled water 30min before LPS inoculation, respectively. Gavage with saline or drug was continued on days 2 and 3 for 3 total times, observed for 6 days, and the number of deaths recorded. The experimental grouping and results are shown in Table 3, where LPS control mice survived only 20% (2/10), Andrographolide Dispersion (AD) survived 40% (4/10) and 50% (5/10) in the high and low dose groups, respectively, while prescription III andrographolide solution (AD-NE) survived 70% (7/10) and 60% (6/10) in the high and low dose groups, respectively.

TABLE 3 protective Effect of the dispersions or solutions of andrographolide gavage on LPS-inoculated mice

Claims

1. An andrographolide solution capable of self-emulsifying comprises the following components in percentage by mass: 0.1-5.0% of andrographolide, 25-75% of surfactant, 10-50% of cosurfactant, 10-35% of vegetable oil and 100% of the total composition.

2. The andrographolide solution of claim 1, wherein the surfactant is one or a mixture of polyethylene glycol 400 monooleate, polyethylene glycol 600 monooleate, polyethylene glycol 400 monolaurate, caprylic/capric macrogol glyceride, polyoxyethylene 40 hydrogenated castor oil, polyethylene glycol vitamin E succinate, lecithin, soya lecithin, Tween 80.

3. The andrographolide solution of claim 1, wherein the co-surfactant is one or a mixture of polyethylene glycol 400, ethanol, 1, 2-propanediol, 1, 3-propanediol, diethylene glycol monoethyl ether, and dimethyl sulfoxide (DMSO).

4. The andrographolide solution of claim 1, wherein the vegetable oil is one or a mixture of olive oil, camellia oil and isopropyl myristate.

5. The andrographolide solution according to any one of claims 1-5, which is self-emulsifiable to an O/W microemulsion after being added with water, wherein the diameter of drug-containing droplets of the microemulsion is less than 1 micron.

6. The method for preparing the self-emulsifiable andrographolide solution of claim 1, comprising the following steps:

(1) weighing a certain amount of andrographolide, adding a cosurfactant, and heating to dissolve at 60-90 ℃;

(2) according to the HLB value of the surfactant phase, the proportion of the surfactant to the oil is changed according to the proportion of 9: 1-1: 9, and a proper proportion is determined by utilizing a ternary phase diagram method;

(3) adding surfactant and vegetable oil into andrographolide solution dissolved in cosurfactant, and stirring to obtain andrographolide solution capable of self-emulsifying.