CN109453166B

CN109453166B - Solid dispersion of rifamycin-quinolizinone coupled molecules and application thereof

Info

Publication number: CN109453166B
Application number: CN201811200949.0A
Authority: CN
Inventors: 刘宇; 徐向毅; 马振坤
Original assignee: Danuo Pharmaceutical Suzhou Co ltd
Current assignee: Danuo Pharmaceutical Suzhou Co ltd
Priority date: 2018-10-16
Filing date: 2018-10-16
Publication date: 2021-03-12
Anticipated expiration: 2038-10-16
Also published as: CN109453166A

Abstract

The invention provides a solid dispersion of rifamycin-quinolizidone coupling molecules, which comprises rifamycin-quinolizidone coupling molecules with a structure shown in a formula I, a high molecular carrier, functional auxiliary materials and a solvent; the high molecular carrier comprises one or a combination of more of povidone K30, povidone VA64, hydroxypropyl cellulose L, polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer and polymethacrylate; the functional auxiliary materials comprise one or a combination of more of vitamin E polyethylene glycol succinate, sodium dodecyl sulfate, meglumine and Tween 80;

the solid dispersion of the rifamycin-quinolizinone conjugate molecule of the invention can be used as a formulation of a medicament for treating bacterial infections.

Description

Solid dispersion of rifamycin-quinolizinone coupled molecules and application thereof

Technical Field

The invention relates to a solid dispersion of rifamycin-quinolizidone coupling molecules and application thereof, belonging to the technical field of medicines.

Background

The rifamycin-quinolizidone coupling molecule has a structure shown in a formula I in the specification, is a semi-synthetic medium-broad-spectrum antibacterial drug, is formed by connecting rifamycin and quinolizidone pharmacophores through stable covalent bonds, and is a novel antibacterial drug with a multi-target action mechanism. The rifamycin-quinolizinone conjugate molecule can selectively react with DNA-dependent RNA polymerase to inhibit the transcription process of bacterial DNA. Meanwhile, mutual transformation between DNA topoisomers is prevented by interaction with DNA topoisomerase (including DNA gyrase and DNA topoisomerase IV), and DNA replication, recombination and transcription processes are inhibited. Through the synergistic effect of the coupling pharmacophore, the rifamycin-quinolizinone coupling molecule can effectively inhibit the formed single-drug-resistant or multi-drug-resistant clinical strains and greatly reduce the frequency of drug resistance generated by bacteria. However, in the prior art, a preparation of the rifamycin-quinolizinone coupling molecule with high dissolution rate and reliable curative effect is still lacked.

Disclosure of Invention

In view of the above-mentioned drawbacks of the prior art, it is an object of the present invention to provide a solid dispersion of rifamycin-quinolizinone coupling molecules, which can be used as a good formulation of drugs for treating bacterial infections, and its use.

The purpose of the invention is realized by the following technical scheme:

a solid dispersion of rifamycin-quinolizidone coupling molecules comprises rifamycin-quinolizidone coupling molecules with a structure shown in formula I, a high molecular carrier, functional auxiliary materials and a solvent;

the polymer carrier comprises one or a combination of more of povidone K30(PVP K30), povidone VA64(PVP-VA64), hydroxypropyl cellulose L (HPC-L), polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer (Soluplus) and polymethacrylate (Eudragit EPO);

the functional auxiliary materials comprise one or more of vitamin E polyethylene glycol succinate (VE-TPGS), sodium dodecyl sulfate (SLS), meglumine and Tween 80 (Tween-80);

in the above solid dispersion, preferably, the polymethacrylate includes yuteqi EPO, but is not limited thereto.

In the solid dispersion described above, preferably, the solid dispersion is characterized by one or more XRPDs substantially similar to figure 8.

In the solid dispersion described above, preferably, the solid dispersion is characterized by one or more thermograms substantially similar to those of fig. 9, 10, 11.

In the solid dispersion, preferably, the polymer carrier comprises one or more of hydroxypropyl cellulose L, polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer and ewing EPO in combination.

In the solid dispersion, preferably, the functional adjuvant is vitamin E polyethylene glycol succinate (VE-TPGS).

In the solid dispersion, preferably, the dosage of the rifamycin-quinolizidone coupling molecule is 23.8% -71.2%, the dosage of the polymeric carrier is 23.8% -71.2%, and the dosage of the functional adjuvant is 3% -7% (preferably 5%) based on 100% of the total mass of the rifamycin-quinolizidone coupling molecule, the polymeric carrier and the functional adjuvant.

In the above solid dispersion, the solvent may be a commonly used agent in the art; preferably, the solvent comprises one or a combination of several of butanone, tetrahydrofuran, dichloromethane and methyl isobutyl ketone; tetrahydrofuran, dichloromethane or acetone are preferred.

In the above solid dispersion, preferably, the solid dispersion remains amorphous according to XRPD when it is left to stand for 4 weeks at 40 ℃/75% RH or 60 ℃/open.

The invention also provides a microgranule comprising a solid dispersion of the rifamycin-quinolizinone coupling molecule described above.

The invention also provides a medicinal composition which comprises the microgranule.

The present invention also provides a pharmaceutical composition comprising a solid dispersion of the rifamycin-quinolizinone coupling molecule described above.

The invention also provides application of the solid dispersion of the rifamycin-quinolizinone coupling molecule in preparing a medicament for treating diseases caused by human bacterial infection.

The invention also provides application of the particle preparation in preparing a medicament for treating diseases caused by bacterial infection of a human body.

The invention also provides application of the medicinal composition in preparing a medicament for treating diseases caused by bacterial infection of a human body.

The main indications of the oral preparation formed by the rifamycin-quinolizinone coupled molecule solid dispersant include common and serious gastrointestinal tract infection caused by gram-positive bacteria and microaerophilic bacteria, including but not limited to helicobacter pylori infection and liver cirrhosis related hyperammonemia, small intestine bacterial overgrowth and irritable bowel syndrome, etc. A number of in vitro tests have shown that rifamycin-quinolizinone conjugate molecules have good antibacterial activity against a variety of aerobic and anaerobic gram-positive or gram-negative bacteria. The activity of the rifamycin-quinolizinone coupled molecule on main intestinal ammonia-producing bacteria (bacteria possibly related to intestinal ammonia production) shows that the rifamycin-quinolizinone coupled molecule has the same or stronger bacteriostatic activity on main intestinal ammonia-producing bacteria such as bifidobacterium infantis subspecies, bacteroides fragilis, clostridium difficile, clostridium perfringens, eggpt bacterium, escherichia coli, helicobacter pylori, lactobacillus salivarius, clostridium necrophorum, streptococcus praecox, morganella morganii, proteus vulgaris, salmonella, yersinia colitis and the like. Compared with ciprofloxacin, the rifamycin-quinolizinone coupled molecule has longer effect after antibiotic, sub-inhibitory concentration effect and sub-inhibitory concentration effect after antibiotic, lower spontaneous drug resistance frequency, faster sterilization speed and better capacity of preventing the generation of drug resistance gene mutation. The results of pharmacodynamic studies in vitro and animal models show that the rifamycin-quinolizinone coupled molecule has significant bactericidal effect and good drug effect on clinical isolated strains related to indications, including drug-resistant strains.

The invention has the outstanding effects that:

Drawings

FIG. 1 is a standard solution high performance liquid chromatogram for dissolution test assay analysis of rifamycin-quinolizinone coupling molecules of the examples performed according to method 1;

FIG. 2 is a standard solution high performance liquid chromatogram for dissolution test assay analysis of rifamycin-quinolizinone coupling molecules of the examples performed according to method 2;

FIG. 3 is a standard solution high performance liquid chromatogram for dissolution test assay analysis of rifamycin-quinolizinone coupling molecules of the examples performed according to method 3;

FIG. 4 is a graph of dissolution behavior (25 ℃) of a solid dispersion film in simulated gastric fluid at pH 1.2;

FIG. 5 is a graph of dissolution behavior (25 ℃) of a solid dispersion film in acetate buffered salt at pH 4.5;

FIG. 6 is a comparison of three kinds of solid dispersion powders (A) prepared by spray drying and dissolution residues (B) obtained by the spray drying method;

FIG. 7 is a scanning electron microscope comparison of three formulations of solid dispersion powder prepared by spray drying and original rifamycin-quinolizidone coupling molecule;

FIG. 8 is a powder X-ray diffraction pattern of a solid dispersion prepared by three-component spray drying;

FIG. 9 is a MDSC curve for the solid dispersion powder of formulation 3;

FIG. 10 is a MDSC curve for the solid dispersion powder of formulation 2;

FIG. 11 is a MDSC curve for the solid dispersion powder of formula 1;

FIG. 12 is a graph showing the dissolution behavior of a solid dispersion powder prepared by a spray drying method in acetate buffer at pH 4.5;

FIG. 13 is a comparative graph of the polarized light microscope showing the solid dispersion powder (A) prepared by spray drying of recipes 4 and 5 and the dissolution residue (B) thereof;

FIG. 14 is a scanning electron micrograph of the powders of the solid dispersions of formulas 4 and 5;

FIG. 15 is a graph of powder X-ray diffraction contrast for solid dispersions of different drug loadings;

FIG. 16 is a MDSC curve for a solid dispersion powder with an API drug load of 47.5%;

FIG. 17 is a MDSC curve for an API drug load of 72.1% solid dispersion powder;

FIG. 18 is a graph of the dissolution behavior of solid dispersion powders at different drug loadings in acetate buffer pH 4.5;

FIG. 19 is an XRPD pattern for a 5 day stability experiment for the resulting solid dispersions of

formulas

3, 4 and 5;

FIG. 20 is a polarized light microscope image of 5-day stability experiment of solid dispersions obtained from

recipes

3, 4, and 5 under different stability conditions

FIG. 21 is a graph of the MDSC profile of the solid dispersion from formulation 3 after experimental conditions of stability at 40 deg.C/75% RH;

FIG. 22 is a graph of the MDSC profile of the solid dispersion from formulation 4 after experimental conditions of stability at 40 deg.C/75% RH;

FIG. 23 is a graph of the MDSC profile of the solid dispersion from formulation 5 after experimental conditions of 40 deg.C/75% RH stability;

FIG. 24 is a graph of the MDSC profile of the solid dispersion from formulation 3 after 50 deg.C/open stability test conditions;

FIG. 25 is a graph of the MDSC profile of the solid dispersion from formulation 4 after 50 deg.C/open stability test conditions;

FIG. 26 is a graph of the MDSC profile of the solid dispersion from formulation 5 after 50 deg.C/open stability test conditions;

FIG. 27 is a plot of PLM at 2 weeks and 4 weeks for the solid dispersions obtained from formulation 3 under different stability conditions;

FIG. 28 is an XRPD pattern for 2 weeks of the solid dispersion from formula 3 under different stability conditions;

FIG. 29 is an XRPD pattern for 4 weeks of the solid dispersion from formula 3 under different stability conditions;

FIG. 30 is a MDSC curve for the solid dispersion of formula 3 at 25 deg.C/60% RH for 2 weeks;

FIG. 31 is a MDSC curve for the solid dispersion of formula 3 at 40 ℃/75% RH for 2 weeks;

FIG. 32 is a MDSC curve of the solid dispersion of formula 3 at 60 ℃ for 2 weeks;

FIG. 33 is a MDSC curve for the solid dispersion of formula 3 at 25 deg.C/60% RH for 4 weeks;

FIG. 34 is a MDSC curve for the solid dispersion of formula 3 at 40 deg.C/75% for 4 weeks;

FIG. 35 is a MDSC curve of the solid dispersion of formula 3 at 60 ℃ for 4 weeks.

Detailed Description

The technical solutions of the present invention will be described in detail below in order to clearly understand the technical features, objects, and advantages of the present invention, but the present invention is not limited to the practical scope of the present invention. The experimental methods described in the following examples are all conventional methods unless otherwise specified; the reagents and materials are commercially available, unless otherwise specified.

As used herein, "similar, analogous" with respect to forms exhibiting similar characteristics as, for example, XRPD, IR, raman spectroscopy, DSC, TGA, NMR, SSNMR, etc., means that the polymorph or co-crystal can be identified by the method and can range from similar to substantially similar, as long as the material is identified by the method with the variability expected by one skilled in the art (based on experimental variations including, for example, the instrument used, time of day, humidity, season, pressure, room temperature, etc.).

Polymorphism, as used herein, refers to the appearance of different crystalline forms of a single compound in different hydration states (e.g., the nature of some compounds and complexes). Thus, polymorphs are distinct solids sharing the same molecular formula, yet each polymorph may have unique physical properties. Thus, a single compound may result in multiple polymorphs, each of which has different and unique physical properties, such as solubility profile, melting point temperature, hygroscopicity, particle shape, density, flowability, compactibility, and/or x-ray diffraction peaks. The solubility of each polymorph can vary, and thus identification of the presence of a pharmaceutically acceptable polymorph is essential to provide a drug with a predictable solubility profile. It is desirable to investigate all solid state forms of a drug including all polymorphs and to determine the stability, dissolution and flowability of each polymorph. Polymorphs of a compound can be distinguished in the laboratory by X-ray diffraction spectroscopy and by other methods such as infrared spectroscopy.

The materials and equipment conditions used in the examples of this specification, and the methods involved, are as follows:

TABLE 1 reagents

TABLE 2 auxiliary materials

Name (R)	Manufacturer of the product	Batch number
			Polyvinylpyrrolidone K30	ASHLAND	0001910711
Polyvinylpyrrolidone VA64	BASF	13968068E0
			Hydroxypropyl cellulose-L	Nippon Soda	NCA-2021
Hydroxypropyl methylcellulose E3 premix LV	Colorcon	PD280165
			Soluplus	BASF	84414368E0
Ewing EPO	EVONIK	G120231025
			Sodium dodecyl sulfate	BASF	0008985310
Meglumine	Alfa Aesar	10151696
			Vitamin E-polyethylene glycol succinate	Healthcare	C071710801
Tween-80	Sigma-Aldrich	172402112

TABLE 3 Instrument Equipment

Instrument and equipment parameters:

line powder diffractometer (XRPD)

Light pipe Cu: K-Alpha

A generator: voltage is 40 kV; current is 40 mA;

scanning angle: 3to 40 deg;

sample rotation speed: 0 rpm;

scanning speed: 10 deg./min.

Modulation type differential calorimeter (MDSC)

Equilibrating at 0 ℃; modulation was. + -. 1 ℃ per 60 seconds; keeping the temperature constant for five minutes; the temperature is raised to 200 ℃ at the speed of 2 ℃/min.

Thermogravimetric analyzer (TGA)

The sample was warmed from room temperature at 10 ℃ to 300 ℃.

Polarizing microscope (PLM)

Nikon LV100POL with 5 Mpixel CCD;

magnification: 20X.

Scanning Electron Microscope (SEM)

The model is Phenom Prox; the accelerating voltage is 10 KV-Point.

Dynamic moisture adsorption apparatus (DVS)

And (3) balancing conditions: dm/dt is 0.01%/min; humidity setting: 0,10,20,30,40,50,60,70,80,90,80,70,60,50,40,30,20,10,0.

Tap density instrument

The instrument comprises the following steps: 10mL measuring cylinder, ERWEKA tap density instrument; lifting speed: 300 times per minute; fixing the height: 14 mm.

Bulk density: the powder was passed slowly through a sieve (1.0mm, No.18) to a 10mL graduated cylinder. Each sample was tested in triplicate.

Tap density: the tap density measurement according to USP was performed as follows: the cylinder was vibrated 500 times and the volume, Va, was measured. The cylinder was vibrated 750 times again and the volume, Vb, was measured. If the difference in volume between the two times is less than 2%, Vb is the final tapped volume. If the volume change is more than 2%, repeating the vibration mode 1250 times in each time until the volume change of two adjacent times is less than 2%, and the volume of the last time is the final tap volume.

The compression coefficient and the Hawson index are calculated according to the following formula:

carl index (compression factor) 100 × (1- ρ)_bulk/ρ_tapped)

Hawson index rho_tapped/ρ_bulk

ρ_bulkAs bulk density value, p_tappedIs a tap density value

(reference: USP29-NF24<1174>)

pH 1.2 simulated gastric fluid preparation:

400 μ L of 12N concentrated HCl was pipetted to 100mL of pH 1.8 simulated gastric fluid using a pipette gun. The pH meter measures the pH of the solution to be 1.21.

Simulated gastric fluid (pH 1.8) 950mL of ultrapure water was weighed into a 1L volumetric flask, 1.4mL of 12N concentrated hydrochloric acid and 2g of sodium chloride were added, and the mixture was stirred uniformly. And the volume is determined by ultrapure water.

pH 4.5 acetate buffer preparation:

869.05mg of sodium hydroxide solid particles were weighed, 1L of ultrapure water was weighed into a 1L glass bottle, 3.15mL of glacial acetic acid was added to the bottle using a pipette, and the pH of the solution was measured by a pH meter to be 4.53.

The high performance liquid chromatography determination method of the rifamycin-quinolizinone coupled molecule comprises the following steps:

the hplc content and impurity detection methods are shown in tables 4, 6 and 8, where method 2 in table 6 effectively improves the resolution of the chromatographic peaks. The HPLC spectra corresponding to the 3 liquid phase methods are respectively shown in FIG. 1, FIG. 2 and FIG. 3. The retention times for the rifamycin-quinolizinone conjugate molecule were 2.74 minutes (assay, method 1), 4.32 minutes (assay, method 2), and 22.27 minutes (impurity assay, method 3), respectively. The system adaptability test results can meet the experimental requirements (tables 5, 7 and 9).

TABLE 4 dissolution test assay high performance liquid chromatography method (method 1)

TABLE 5 dissolution test content analysis liquid phase method system adaptability results

Limit of quantitation being 0.5. mu.g/mL

TABLE 6 liquid phase method after correction of dissolution test content analysis (method 2)

TABLE 7 liquid phase method system adaptability results after correction of dissolution test content analysis

Parameter(s)	Standard of merit	Results
			Blank (Diluent)	No significant interference peaks (<LOQ)	By passing
Sensitivity of the probe	S/N≥10	By passing
			Recovery rate	98.0％-102.0％	100.2
Resolution ratio	≥1.2	3.22
			Tailing factor	0.8≤T≤2.0	1.2
Precision degree	≤2.0	0.11
			Linear (mg/mL)	R²≥0.9990	1

TABLE 8 stability experiments impurity analysis high performance liquid chromatography method (method 3)

TABLE 9 stability experiment impurity analysis liquid phase method System adaptability results

Examples

This example provides a solid dispersion of a rifamycin-quinolizinone coupled molecule, which comprises a rifamycin-quinolizinone coupled molecule having a structure shown in formula I, a polymeric carrier, and a solvent.

The rifamycin-quinolizinone conjugate molecule of this example (abbreviated as API or drug substance for convenience as shown in formula I) is produced by dano medicine (suzhou) ltd.

The polymer carrier comprises one or a combination of more of povidone K30 (namely polyvinylpyrrolidone K30), povidone VA64 (namely polyvinylpyrrolidone VA64), hydroxypropyl cellulose L (HPC-L), polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer (Soluplus), polymethacrylate (Eudragit EPO) and vitamin E-polyethylene glycol succinate (VE-TPGS).

In this example, a solvent was first subjected to a screening assay: the solvent comprises one or a combination of more of butanone, tetrahydrofuran, dichloromethane and methyl isobutyl ketone. Weighing about 10mg of raw material medicine into 2mL of liquid phase vials, adding different solvents with proper volumes respectively, observing and judging whether the raw material medicine is completely dissolved or not, and judging the approximate solubility of the raw material medicine.

The results are shown in Table 10 below. According to the approximate solubility test results, dichloromethane, tetrahydrofuran or acetone was used to prepare the appropriate concentration of drug substance/adjuvant mixture solution.

TABLE 10 approximate solubilities of drug substances in different solvents

Solvent(s)	Security level	Approximate solubility (mg/mL)
			Methanol	2	<1
Acetone (II)	3	2.3～3.0
			Butanone	3	9.7～19.4
Tetrahydrofuran (THF)	3	>200
			Methylene dichloride	2	>200
Methyl isobutyl ketone	3	20～50

Reference ICH Q3c

In this example, the polymer carrier was screened:

different polymers (povidone K30, povidone VA64, hydroxypropyl cellulose L, hydroxypropyl methyl cellulose E3, Soluplus, ewt EPO) were used as dispersants and binary solid dispersions were prepared by solvent evaporation and their solubilization of the drug substance was evaluated by vial dissolution experiments.

Preparation of binary solid dispersions (abbreviation ASD): approximately 12.5mg of the rifamycin-quinolizinone conjugate molecule drug and 37.5mg of the polymer are weighed into a 40mL glass vial and dissolved completely by the addition of an appropriate amount of solvent to give a clear solution. To remove the solvent as soon as possible to obtain a dry drug/polymer film, the open 40mL vials were wrapped in perforated aluminum foil paper and placed in a vacuum oven at 35 ℃ for 40 hours to obtain binary solid dispersion films, the preparation of which is shown in table 11 below:

TABLE 11 preparation of binary solid Dispersion films

The prepared binary solid dispersion film was subjected to dissolution test:

5mL of simulated gastric fluid at pH 1.2 or acetate buffer at pH 4.5 was added to a 40mL vial prepared with a thin film of solid dispersion (target concentration 2.5mg/mL), vortexed by a vortexer for 5 seconds (as a first point, 0 min), and then the 40mL vial was placed on a shaker. Pipette 500. mu.L (pH 1.2 system) or 1mL (pH 4.5 system) of the suspension into a centrifuge tube at 0 min, 30 min, 60 min, respectively, and centrifuge for five minutes at 140000 rpm. The supernatant was taken to a liquid phase vial and diluted with methanol by an appropriate fold for liquid phase analysis (method 1).

In either the pH 1.2SGF solution or pH 4.5 buffer system, all solid dispersion systems (23.8% drug loading) were in suspension with or without incompletely dissolved solids, except that the API/EPO system appeared as a clear solution containing two phases. Based on the dissolution test results shown in tables 12-13 and fig. 4-5, the pure raw material drug has lower solubility, and the solubilizing effect of PVP K30, PVP VA64 or HPMC E3 on the rifamycin-quinolizinone coupling molecule is not obvious. However, for the HPC-L, Soluplus and Ettky EPO systems, dissolution experiments show that the corresponding solid dispersion film has very obvious solubilizing effect on the rifamycin-quinolizinone coupling molecules.

TABLE 12 dissolution results of solid dispersion film in simulated gastric fluid at pH 1.2 (25 ℃ C.)

TABLE 13 dissolution results of solid dispersion film in acetate buffer salt having pH 4.5 (25 ℃ C.)

In this example, functional excipients were screened.

To evaluate the solubilization of the functional excipients on the rifamycin-quinolizinone conjugate molecule, different functional excipients (sodium dodecyl sulfate, meglumine, vitamin E-polyethylene glycol succinate, or tween-80) were pre-dissolved in a pH 1.2SGF solution and a pH 4.5 buffer at a target concentration of 3 mg/mL. Taking about 20mg of API, putting the API into a 1.5mL liquid phase vial, respectively adding 1mL of pH 1.2SGF solution or 1mL of pH 4.5 buffer solution containing different functional auxiliary materials, and putting the solution on a constant temperature mixer at 37 ℃ at the rotating speed of 700 revolutions per minute. After two hours, the solution is taken, centrifuged for five minutes at the rotating speed of 14000 r/min, the supernatant is taken for dilution and injected into a high performance liquid analyzer for analysis, a chromatogram is recorded, and the concentration is calculated according to the peak area of an external standard method (method 1).

The results of the 2 hour solubility experiments for rifamycin-quinolizinone conjugate molecules in solutions containing different functional excipients pH 1.2SGF and pH 4.5 buffer are shown in table 14. The result shows that compared with other functional auxiliary materials, the TPGS can respectively improve the solubility of the API to 761.0 mu g/mL and 80.08 mu g/mL in the pH 1.2SGF solution and the pH 4.5 buffer solution, and has obvious solubilization effect.

TABLE 14 2 hour solubility of rifamycin-quinolizinone coupling molecules in functional adjuvant solution

In this example, a solid dispersion was prepared using a spray drying method:

in view of good solubility and suspension status, Ettky EPO, Soluplus and HPC-L were each chosen as a solid dispersion vehicle adjuvant. For a greater increase in the dissolution rate, VE-TPGS was also added as a functional adjuvant to the solid dispersion formulation. Acetone or dichloromethane was used as the spray-dried solution. The formulation composition and spray drying process parameters are shown in table 15 below. The resulting product was placed in a vacuum oven at 35 ℃ for 38 hours and then stored in a refrigerator at 5 ℃.

TABLE 15 spray drying Process parameters

Physical characterization of the amorphous solid Dispersion powder obtained

All spray dried solids were characterized by HPLC, PLM, XRD, SEM, MDSC to examine dispersion drug loading, microstructure (amorphous or crystalline), surface morphology, particle size and glass transition temperature. The test results are shown in Table 16 and FIGS. 6-11.

The drug loading of the solid dispersion system containing HPC-L, Soluplus or Ettky EPO was 23.5%, 24.7% or 22.4%, respectively, close to the theoretical value of 23.8%. PLM (fig. 6(a)) and XRPD results (fig. 8) show that all spray-dried systems are amorphous. The MDSC maps (FIGS. 9-11) show that the solid dispersion systems containing HPC-L, Soluplus or Ewing EPO all have only one glass transition temperature, which is 86.0 ℃, 119.9 ℃ and 64.6 ℃, respectively, indicating that the rifamycin-quinolizinone coupling molecule and the three excipients can form a uniform amorphous state, and good miscibility is shown. The SEM topography (figure 7) shows that the rifamycin-quinolizinone conjugate molecule/HPC/TPGS spray-dried powder is a severely depressed particle, whereas the spray-dried powder containing Soluplus or ewing EPO is a sphere with a smooth or partially depressed surface.

TABLE 16 characterization of rifamycin-quinolizinone coupled molecules/adjuvants/surfactants spray dried dispersions

Dissolution test of the amorphous solid dispersion powder obtained

About 421mg of the solid dispersion was taken, placed in a 40mL glass vial, taken as dissolution medium (5 mg/mL of API target concentration) with 20mL of pH 4.5 acetate buffer (preheated to 37 ℃), sealed, vortexed for 5 seconds, added with a stirrer, placed on a heating magnetic stirrer at 37 ℃ and rotated at 300 rpm. After 10 minutes, 20 minutes, 40 minutes, 60 minutes, 90 minutes, 120 minutes and 180 minutes, 500 mu L of liquid is taken out and put into a centrifuge tube, the centrifuge tube is centrifuged for 3 minutes at the rotating speed of 14000 r/min, supernatant liquid is taken out and put into a liquid phase bottle, the liquid phase bottle is diluted by methanol and injected into a high performance liquid chromatograph, chromatogram is recorded, and the concentration is calculated by peak area according to an external standard method. In addition, the suspensions after the dissolution test were subjected to pH measurement, and the remaining residue was subjected to PLM characterization, and the data results are shown in table 17.

Bulk drug/ewt's EPO/VE-TPGS solid dispersions significantly increased API dissolution rate (10 min dissolution rate of 2.44mg/mL) in pH 4.5 acetate buffer compared to solid dispersion systems containing HPC-L and Soluplus (10 min dissolution rates of 0.007 and 0.084mg/mL, respectively), and the supersaturation was maintained throughout the dissolution process. After 180 minutes dissolution experiments, the residue of all solid dispersion systems was all amorphous characterized by PLM (as shown in fig. 6). Therefore, drug substance/eudragit EPO/TPGS was chosen as the dominant prescription for subsequent drug loading screening.

TABLE 17 dissolution results of solid dispersion powder in acetate buffer solution of pH 4.5 (37 ℃ C.)

In this example, the drug loading of the solid dispersion is screened

Based on the results of the 23.8% drug loading spray-dried dispersion dissolution test, ewing EPO was selected as the carrier for the solid dispersion formulation. API/yuteqi EPO 1:1 and 1:3, and formula 4 and formula 5 with 5% TPGS were used to prepare amorphous solid dispersions. Acetone was used as the solvent for the preparation of the spray-dried solution. The formulation composition and process parameters are shown in table 18 below. The resulting product was dried at 35 ℃ for 20 hours and then stored in a refrigerator at 5 ℃.

Watch 18

Characterization of the solid dispersions obtained according to recipes 4, 5

All spray dried solid powders were characterized by HPLC, PLM, XRD, SEM, MDSC to examine dispersion drug loading, microstructure (amorphous or crystalline), surface morphology, particle size and glass transition temperature. The test results are shown in Table 19 below.

According to the result of high performance liquid chromatography, the drug loading of the prescription 4 and the drug loading of the prescription 5 are respectively 39.7 percent and 52.3 percent, and are slightly smaller than the theoretical values thereof, namely 47.5 percent and 71.2 percent. PLM and XRPD results (as shown in fig. 13(a) and fig. 15) show that the spray dried dispersion is amorphous. As shown in fig. 16 and 17, recipe 4 and recipe 5 have only one glass transition temperature occurring at 97.1 c or 126.9 c, respectively, indicating good mixing between the components in the system. According to the SEM spectrum (as shown in FIG. 14), the solid dispersion powder was a particle having severe dishing.

TABLE 19 physical characterization results of solid dispersion powders with different drug loadings

Powder dissolution test (Artificial gastroduodenal model) was performed on the solid dispersions obtained according to formulas 4 and 5

Taking appropriate amount of solid dispersion formula 4 and formula 5, respectively, taking pH 4.5 acetate buffer (preheated to 37 ℃) as dissolution medium (API target concentration is 5mg/mL), sealing, performing vortex treatment for 5 seconds, adding a stirrer, and placing on a heating magnetic stirrer at 37 ℃ with the rotating speed of 300 revolutions per minute. At 10 min, 20 min, 40 min, 60 min, 90 min, 120 min and 180 min, 500 μ L of the liquid was taken into a centrifuge tube, centrifuged at 14000 rpm for 3 min, the supernatant was taken into a liquid phase vial, diluted five times with methanol and injected into a high performance liquid chromatograph (method 1). The dissolution results are shown in table 20 below and fig. 18. To compare the dissolution behavior of the different drug loaded solid dispersions, the dissolution results of formula 1 at 23.8% drug load tested in section 3.2.3 are also presented here.

In this dissolution experiment, the dissolution rates of the 47.5% and 71.2% drug-loaded solid dispersion systems were much less than that of the 23.8% drug-loaded solid dispersion formulation 1. The drug loading is increased from 23.8 percent to 71.2 percent, and simultaneously the dissolution concentration is reduced from 2.4mg/mL to 0.03 mg/mL. The pH values of the dissolution solution are all less than 6, which eliminates the influence of EPO dissolution process during dissolution. The large differences in dissolution behavior of solid dispersions of different drug loading may be due to differences in wettability of the particles, distribution of the components, and the ability of the API to recrystallize under different microenvironments. The 3 hour dissolution residue PLM results (as shown in fig. 13 (B)) show that there are significant crystals for both formula 4 and formula 5, which also explains its lower dissolution rate from another perspective.

TABLE 20 dissolution results of solid dispersion powder in acetate buffer solution of pH 4.5 (37 ℃ C.)

Short-term physical stability studies of the solid dispersions obtained according to

recipes

3, 4 and 5

Taking a certain amount of solid dispersion, respectively putting into 2mL liquid-phase small bottles, covering the bottle openings with a pricked aluminum foil paper, and respectively placing the small bottles in an open stabilization box with 40 ℃/75% RH and 50 ℃/for five days. XRPD, PLM and MDSC characterization was performed on all 5 day stable samples. XRPD results are shown in fig. 19, showing that all solid dispersions are still amorphous. The PLM results show that the remaining dispersion system is also amorphous except that the solid dispersion 4 samples were treated at 50 deg.C/open and the solid dispersion 5 was treated at 40 deg.C/75% RH with a small amount of crystals appearing (FIG. 20). The MDSC results showed (as shown in table 21 and fig. 21-26) that almost all samples had only 1 glass transition temperature indicating no phase separation, while formula 3 exhibited two tgs at 40 ℃/75% RH indicating that the samples were unstable and phase separated to some extent under high humidity conditions.

TABLE 21 change in glass transition temperature of solid dispersion powder in physical stability experiment

Long-term physical stability studies of the solid dispersions obtained according to formula 3

A small amount of the solid dispersion powder (bulk drug/Ewing EPO/VE-TPGS) obtained in the formula 3 is taken into a 40mL bottle, the opening of the bottle is covered by a perforated aluminum foil, and the bottle is respectively placed in a 25 ℃/60% RH stabilizing box or an electrothermal blowing drying box at 40 ℃/75% RH and 60 ℃. After 2 and 4 weeks, XRPD, PLM and MDSC characterization was performed on the stability samples, respectively.

According to XRPD and PLM (as shown in fig. 27-29), all solid dispersion stability samples remained amorphous after 4 weeks. The MDSC results (as shown in fig. 30-35) show that the samples subjected to three different temperature and humidity controls have only one glass transition temperature, but the values are distributed between 61.8 ℃ and 82.4 ℃ and deviate from the glass transition temperature (69.4 ℃) of the initial solid dispersion, which indicates that the solid dispersion does not phase separate but the single-phase microstructure of the system is changed under different temperature and humidity conditions.

In embodiments of the present disclosure, there is also provided a microparticle comprising a solid dispersion of a rifamycin-quinolizinone coupling molecule as described above. The granulation machine can be a conventional granulation machine, can also be an intra-particle controlled release agent, and is prepared into granules by using a solid dispersion of rifamycin-quinolizinone coupling molecules; all disintegrate through the microgranules to release the active ingredient rifamycin-quinolizinone conjugate molecule.

In another embodiment of the present invention, there is provided a pharmaceutical composition comprising the above fine granules.

Also provided in embodiments of the present specification is a pharmaceutical composition comprising a solid dispersion of a rifamycin-quinolizinone coupling molecule as described above. Namely, the composition is a composition containing a solid dispersion of the rifamycin-quinolizinone coupling molecule and other auxiliary materials, and can also comprise a pharmaceutically acceptable carrier. Further, the excipients comprise excipients such as one or more of diluents, binders, lubricants, intragranular controlled release agents, disintegrants, colorants, flavorants or sweeteners. The above compositions can be formulated for the preparation of coated and uncoated tablets, hard and soft gelatin capsules, dragees, lozenges, wafers, pellets, powders in sealed packets and the like. The above compositions may be formulated for topical use in preparations such as ointments, pomades, creams, gels and lotions. A "pharmaceutically acceptable carrier" includes a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, which is involved in carrying or transporting a chemical of the invention from one organ or portion of the body to another organ or portion of the body. Each carrier is preferably "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the subject. Some examples of materials that can be used as pharmaceutically acceptable carriers include: (1) sugars such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered gum tragacanth; (5) malt; (6) gelatin; (7) talc powder; (8) excipients, such as cocoa butter and suppository waxes; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols such as glycerol, sorbitol, mannitol and polyethylene glycol; (12) esters such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) ringer's solution; (19) ethanol; (20) a phosphate buffer solution; and (21) other non-toxic, compatible substances for use in pharmaceutical formulations.

The specific embodiment of the specification also provides application of the solid dispersion of the rifamycin-quinolizinone coupling molecule or the composition thereof in preparing a medicament for treating diseases caused by bacterial infection of a human body.

The specific embodiment of the specification also provides application of the particle preparation in preparing a medicament for treating diseases caused by human bacterial infection.

The specific embodiment of the specification also provides application of the medicinal composition in preparing a medicament for treating diseases caused by bacterial infection of a human body.

In the specific embodiment of the present specification, the main indications of the oral preparation formed by the rifamycin-quinolizinone coupled molecule solid dispersant include common and serious gastrointestinal tract infections caused by gram-positive bacteria and microaerophilic bacteria, including but not limited to helicobacter pylori infection, liver cirrhosis associated with hyperammonemia, small intestine bacterial overgrowth, irritable bowel syndrome and the like. A number of in vitro tests have shown that rifamycin-quinolizinone conjugate molecules have good antibacterial activity against a variety of aerobic and anaerobic gram-positive or gram-negative bacteria. The activity of the rifamycin-quinolizinone coupled molecule on main intestinal ammonia-producing bacteria (bacteria possibly related to intestinal ammonia production) shows that the rifamycin-quinolizinone coupled molecule has the same or stronger bacteriostatic activity on main intestinal ammonia-producing bacteria such as bifidobacterium infantis subspecies, bacteroides fragilis, clostridium difficile, clostridium perfringens, eggpt bacterium, escherichia coli, helicobacter pylori, lactobacillus salivarius, clostridium necrophorum, streptococcus praecox, morganella morganii, proteus vulgaris, salmonella, yersinia colitis and the like. Compared with ciprofloxacin, the rifamycin-quinolizinone coupled molecule has longer effect after antibiotic, sub-inhibitory concentration effect and sub-inhibitory concentration effect after antibiotic, lower spontaneous drug resistance frequency, faster sterilization speed and better capacity of preventing the generation of drug resistance gene mutation. The results of pharmacodynamic studies of in vitro and animal models show that the solid dispersion preparation of the rifamycin-quinolizinone coupled molecule has significant bactericidal effect and good drug effect on clinical isolated strains related to indications, including drug-resistant strains.

Claims

1. A solid dispersion of rifamycin-quinolizidone coupling molecules comprises rifamycin-quinolizidone coupling molecules with a structure shown in a formula I, a high molecular carrier, functional auxiliary materials and a solvent; based on the total mass of the rifamycin-quinolizidone coupling molecules, the polymer carriers and the functional auxiliary materials being 100%, the dosage of the rifamycin-quinolizidone coupling molecules is 23.8% -71.2%, the dosage of the polymer carriers is 23.8% -71.2%, and the dosage of the functional auxiliary materials is 3% -7%;

the polymer carrier comprises one or a combination of more of hydroxypropyl cellulose L, polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer and Ettky EPO;

the functional auxiliary materials comprise one or a combination of more of vitamin E polyethylene glycol succinate, meglumine and Tween 80;

the solvent comprises one or a combination of more of butanone, tetrahydrofuran, dichloromethane and methyl isobutyl ketone;

2. the solid dispersion of claim 1, wherein: the solid dispersion is characterized by an XRPD substantially similar to figure 8.

3. The solid dispersion of claim 1, wherein: the solid dispersion is characterized by a thermogram substantially similar to fig. 9, 10 or 11.

4. The solid dispersion of claim 1, wherein:

the functional auxiliary material is vitamin E polyethylene glycol succinate.

5. The solid dispersion of claim 1, wherein: the solvent is tetrahydrofuran, dichloromethane or acetone.

6. The solid dispersion of claim 1, wherein: the solid dispersion was still amorphous according to XRPD when left to stand for 4 weeks at 40 ℃/75% RH or 60 ℃/open.

7. A microparticle comprising a solid dispersion of a rifamycin-quinolizinone coupling molecule according to any one of claims 1 to 6.

8. A pharmaceutical composition comprising the microgranule of claim 7.

9. A pharmaceutical composition comprising a solid dispersion of a rifamycin-quinolizinone coupling molecule according to any one of claims 1 to 6.

10. Use of a solid dispersion of a rifamycin-quinolizinone coupling molecule according to any one of claims 1 to 6 in the manufacture of a medicament for the treatment of a disease caused by a bacterial infection in a human.

11. Use of the microgranule of claim 7 in the manufacture of a medicament for the treatment of a disease caused by a bacterial infection in a human.

12. Use of a pharmaceutical composition according to claim 9 for the manufacture of a medicament for the treatment of a disease caused by a bacterial infection in a human.