MONTELUKAST BERBERINE QUATERNARY AMMONIUM SALT COMPOUND AND DOUBLE SALT COMPOSITION, AND SYNTHESIS METHOD THEREFOR AND USE THEREOF

Abstract

Disclosed is a montelukast berberine double salt or composition with anti-inflammatory, antibacterial and immunomodulatory effects. Particularly, disclosed are a montelukast berberine quaternary ammonium salt with a structure as represented by formula (I), a double salt or a composition, a solvate, a hydrate, an isotope substitution or an isomer thereof.

##STR00001##

Claims

1-10. (canceled)

11. A quaternary ammonium salt conjugated compound as represented by formula (I) or a pharmaceutically acceptable salt, a solvate, a composition, an enantiomer and an isotope substitution or a double salt thereof, ##STR00036## wherein R.sub.1 and R.sub.2 are independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.1 or any two adjacent R.sub.2 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; R.sub.3 and R.sub.4 are independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.3 or any two adjacent R.sub.4 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, m is an integer arbitrarily selected from 0, 1, 2, 3 and 4; n is an integer arbitrarily selected from 0, 1, 2, 3 and 4; P is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5; and t is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5.

12. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 11, having a structure as represented by formula (IA), ##STR00037## wherein R.sub.1 and R.sub.2 are independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.1 or any two adjacent R.sub.2 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, m is an integer arbitrarily selected from 0, 1, 2, 3 and 4; and n is an integer arbitrarily selected from 0, 1, 2, 3 and 4.

13. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 11, having a structure as represented by formula (IB), ##STR00038## wherein R.sub.1 is independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.1 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and m is an integer arbitrarily selected from 0, 1, 2, 3 and 4.

14. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 12, having a structure as represented by formula (IB), ##STR00039## wherein R.sub.1 is independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.1 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and m is an integer arbitrarily selected from 0, 1, 2, 3 and 4.

15. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 11, having a structure as represented by formula (IC), ##STR00040## wherein R.sub.2 is independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally optimally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.2 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and n is an integer arbitrarily selected from 0, 1, 2, 3 and 4; preferably, the quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof, having a structure as represented by formula (ID), ##STR00041## wherein R.sub.3 and R.sub.4 are independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally optimally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.3 or any two adjacent R.sub.4 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and P is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5; and t is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5.

16. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 12, having a structure as represented by formula (IC), ##STR00042## wherein R.sub.2 is independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally optimally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.2 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and n is an integer arbitrarily selected from 0, 1, 2, 3 and 4; preferably, the quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof, having a structure as represented by formula (ID), ##STR00043## wherein R.sub.3 and R.sub.4 are independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally optimally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.3 or any two adjacent R.sub.4 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and P is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5; and t is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5.

17. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 13, having a structure as represented by formula (IC), ##STR00044## wherein R.sub.2 is independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally optimally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.2 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, and 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and n is an integer arbitrarily selected from 0, 1, 2, 3 and 4; preferably, the quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof, having a structure as represented by formula (ID), ##STR00045## wherein R.sub.3 and R.sub.4 are independently selected from hydrogen, deuterium, halogen, CN, C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl, wherein the C.sub.1-10 alkyl, C.sub.2-10 alkenyl, C.sub.2-10 alkynyl, C.sub.1-10 alkoxyl, 6-10 membered aryl, 5-8 membered heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl and 3-8 membered saturated or partially saturated heterocyclyl are optionally optimally substituted by one to more substituent groups; and the substituent groups are arbitrarily selected from hydrogen, deuterium, halogen, alkyl, haloalkyl, alkoxyl, alkylamino, O?, CN, OH, C.sub.3-10 saturated or partially saturated cycloalkyl, C.sub.3-10 saturated or partially saturated heterocyclyl, 6-10 membered aryl and 5-8 membered heteroaryl; or any two adjacent R.sub.3 or any two adjacent R.sub.4 form 5-6 membered aryl or heteroaryl, 3-8 membered saturated or partially saturated cycloalkyl, 3-8 membered saturated or partially saturated heterocyclyl together with carbon attached thereto, wherein the cycloalkyl and the heterocyclyl are optionally substituted by one to more groups selected from hydrogen, deuterium, halogen, oxo, CN, CF.sub.3, OH, OCH.sub.3, and OCH.sub.2CH.sub.3; heteroatoms in the heteroaryl and heterocyclyl represent any heteroatom independently selected from O, N, S, and P, and isotopes thereof, the halogen is arbitrarily independently selected from F, Cl, Br, and I, and isotopes thereof, and P is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5; and t is an integer arbitrarily selected from 0, 1, 2, 3, 4 and 5.

18. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 11, wherein each R.sub.1 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.1 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.1 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.1 form ##STR00046## together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.2 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.2 form ##STR00047## or together with carbons respectively connected thereto; each R.sub.3 is the same or different, and is each independently selected from hydrogen, hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; each R.sub.3 is the same or different, and is each independently selected from fluorine, chlorine, bromine or iodine, preferably chlorine; each R.sub.4 is the same or different, and is each independently selected from hydrogen or hydroxy C.sub.1-6 alkyl; R.sub.4 is selected from hydrogen or ##STR00048## m is selected from 0, 1, 2 or 3; n is selected from 0, 1, 2, or 3; t is selected from 0 or 1; and p is selected from 0 or 1.

19. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 12, wherein each R.sub.1 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.1 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.1 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.1 form ##STR00049## or together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.2 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.2 form ##STR00050## together with carbons respectively connected thereto; each R.sub.3 is the same or different, and is each independently selected from hydrogen, hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; each R.sub.3 is the same or different, and is each independently selected from fluorine, chlorine, bromine or iodine, preferably chlorine; each R.sub.4 is the same or different, and is each independently selected from hydrogen or hydroxy C.sub.1-6 alkyl; R.sub.4 is selected from hydrogen or ##STR00051## m is selected from 0, 1, 2 or 3; n is selected from 0, 1, 2, or 3; t is selected from 0 or 1; and p is selected from 0 or 1.

20. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 13, wherein each R.sub.1 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.1 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.1 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.1 form ##STR00052## together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.2 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.2 form ##STR00053## together with carbons respectively connected thereto; each R.sub.3 is the same or different, and is each independently selected from hydrogen, hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; each R.sub.3 is the same or different, and is each independently selected from fluorine, chlorine, bromine or iodine, preferably chlorine; each R.sub.4 is the same or different, and is each independently selected from hydrogen or hydroxy C.sub.1-6 alkyl; R.sub.4 is selected from hydrogen or ##STR00054## m is selected from 0, 1, 2 or 3; n is selected from 0, 1, 2, or 3; t is selected from 0 or 1; and p is selected from 0 or 1.

21. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 14, wherein each R.sub.1 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.1 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.1 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.1 form ##STR00055## together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; or two R.sub.2 form a 3-8 membered heterocyclic ring together with carbons respectively connected thereto; each R.sub.2 is the same or different, and is each independently selected from hydrogen, methyl or methoxyl; or two R.sub.2 form ##STR00056## together with carbons respectively connected thereto; each R.sub.3 is the same or different, and is each independently selected from hydrogen, hydrogen, C.sub.1-6 alkyl, and C.sub.1-6 alkoxyl; each R.sub.3 is the same or different, and is each independently selected from fluorine, chlorine, bromine or iodine, preferably chlorine; each R.sub.4 is the same or different, and is each independently selected from hydrogen or hydroxy C.sub.1-6 alkyl; R.sub.4 is selected from hydrogen or ##STR00057## m is selected from 0, 1, 2 or 3; n is selected from 0, 1, 2, or 3; t is selected from 0 or 1; and p is selected from 0 or 1.

22. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to any one of claims claim 11, which is selected from the following ##STR00058## ##STR00059## ##STR00060## ##STR00061## ##STR00062## ##STR00063## ##STR00064## ##STR00065##

23. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to any one of claims claim 12, which is selected from the following ##STR00066## ##STR00067## ##STR00068## ##STR00069## ##STR00070## ##STR00071## ##STR00072## ##STR00073##

24. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to any one of claims claim 13, which is selected from the following structural compound or a corresponding enantiomer or composition thereof: ##STR00074## ##STR00075## ##STR00076## ##STR00077## ##STR00078## ##STR00079## ##STR00080## ##STR00081##

25. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to any one of claims claim 14, which is selected from the following structural compound or a corresponding enantiomer or composition thereof: ##STR00082## ##STR00083## ##STR00084## ##STR00085## ##STR00086## ##STR00087## ##STR00088## ##STR00089##

26. The quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to any one of claims claim 15, which is selected from the following ##STR00090## ##STR00091## ##STR00092## ##STR00093## ##STR00094## ##STR00095## ##STR00096## ##STR00097##

27. A synthesis method for the quaternary ammonium salt conjugated compound, the solvate, the enantiomer and the isotope substitution according to any one of claims claim 11, comprising the following steps: 1) preparation of various free berberine alkaloid acetone addition products: weighing berberine alkaloid quaternary ammonium salt compounds of various acid radicals on demand and placing in a reaction flask, adding an inorganic base, then adding acetone dropwise, and stirring for reaction until the raw materials are completely reacted; performing suction filtration on the reaction mixture, washing a filter cake with water until neutral, and drying to obtain various free berberine alkaloid acetone addition products; and 2) preparation of the quaternary ammonium salt conjugated compound, solvate or composition: weighing montelukast on demand and placing in a reaction flask, adding ethyl acetate to dissolve fully, and then adding an 8-acetonyl dihydroberberine alkaloid compound for reaction under stirring until the raw materials are completely reacted, followed by concentration under reduced pressure or crystallization; or adding an appropriate anti-solvent to the reaction mixture to obtain a berberine montelukast double salt composition.

28. A pharmaceutical composition, wherein the pharmaceutical composition comprises a therapeutically effective amount of at least one of the quaternary ammonium salt conjugated compound or the pharmaceutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to any one of claims claim 11 or a pharmaceutically acceptable carrier or excipient.

29. A method for prevention or treatment of diseases related to inflammation, immunity, infection, allergy, metabolism and others, comprising: administering a patient with a preventively or therapeutically effective amount of at least one of the quaternary ammonium salt conjugated compound or the therapeutically acceptable salt, the solvate, the composition, the enantiomer and the isotope substitution or the double salt thereof according to claim 11.

30. A berberine montelukast double salt composition, wherein the berberine montelukast double salt composition is prepared into a clinically acceptable pharmaceutical preparation by taking a double salt of the quaternary ammonium salt conjugated compound according to any one of claims claim 11 as an effective ingredient and adding appropriate adjuvants and carriers.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0081] FIG. 1 is a comparison diagram of anti-inflammatory effects of different concentrations of double salt compounds in Example 3.

[0082] FIG. 2 is a comparison diagram of anti-inflammatory effects of double salts and monomers thereof in Example 4.

DETAILED DESCRIPTION OF THE INVENTION

[0083] In order to illustrate the present invention in more detail, the following examples are given, but the scope of the present invention is not limited thereto.

Example 1: Preparation of Montelukast Berberine Double Salt (Double Salt 1)

[0084] ##STR00019##

1. Synthesis of 1-(9,10-dimethoxy-5,8-dihydro-6H-[1,3]dioxacyclo[4,5-g]isoquinolinyl[3,2-a]isoquinolin-8-yl) propan-2-one

[0085] ##STR00020##

[0086] At room temperature, a sodium hydroxide (5 M, 6 mL) aqueous solution was added dropwise to an acetone solution (2 mL) in which 9,10-dimethoxy-5,6-dihydro-[1,3]dioxocyclo[4,5-g]isoquinolinyl[3,2-a]isoquinolin-7-ammonium chloride (1.00 g, 2.70 mmol) was dissolved. After dropwise addition was completed, the mixture was reacted at room temperature for 4 h. At the end of the reaction, the reaction solution was directly concentrated to obtain a crude product. The crude product was pulped with methanol (30 mL) for 30 min and then filtered, and a filter cake was collected and dried to obtain a yellow solid compound 1-(9,10-dimethoxy-5,8-dihydro-6H-[1,3]dioxacyclo[4,5-g]isoquinolinyl[3,2-a]isoquinolin-8-yl)propan-2-one (830 mg, 2.11 mmol, yield: 78.1%).

[0087] Product NMR: .sup.1H NMR (400 MHz, DMSO-d.sub.6) ?7.25 (s, 1H), 6.86 (d, J=8.4 Hz, 1H), 6.76 (s, 1H), 6.72 (d, J=8.4 Hz, 1H), 6.00 (s, 1H), 5.99 (s, 2H), 5.22-5.19 (m, 1H), 3.76 (d, J=2.4 Hz, 6H), 3.30-3.26 (m, 1H), 3.20-3.19 (m, 1H), 2.96-2.90 (m, 1H), 2.78-2.76 (m, 2H), 2.33-2.29 (m, 1H), 2.03 (s, 3H).

2. Synthesis of (R,E)-2-(1-(((1-(3-(2-(7-chloroquinolin-2-yl)vinyl)phenyl)-3-(2-(2-hydroxypropyl-2-yl)phenyl) propyl)thio)methyl)cyclopropyl)acetic acid and 9,10-dimethoxy-5,6-dihydro-[1,3]dioxocyclo[4,5-g]isoquino[3,2-a]isoquinolin-7-onium conjugate

[0088] ##STR00021##

[0089] A compound 1-(9,10-dimethoxy-5,8-dihydro-6H-[1,3]dioxane[4,5-g]isoquinolinyl[3,2-a]isoquinolin-8-yl)propan-2-one (791 mg, 2.01 mmol) and a compound (R,E)-2-(1-(((1-(3-(2-(7-chloroquinoline-2-yl)vinyl)phenyl)-3-(2-(2-hydroxypropyl-2-yl)phenyl)propyl)thio)methyl)cyclopropyl)acetic acid (1.18 g, 2.01 mmol) were added in a sealed tube containing methanol (8 mL) and water (0.1 mL), and then the reaction solution was heated to 80? C. and reacted for 2 h. At the end of the reaction, a solvent was removed from the reaction solution to obtain a yellow solid compound (11.90 g, 2.06 mmol, crude product). Then, the crude product compound (600 mg, 0.65 mmol) was added to a mixed solvent (chloroform (6.5 mL), methanol (3.5 mL) and water (0.1 mL)), heated to 80? C. until the crude product was completely dissolved, and stirred at 80? C. for 1 h. At the end of the reaction, the reaction solution was naturally cooled to room temperature, and the reaction solution was directly concentrated to remove the solvent to obtain (R,E)-2-(1-(((1-(3-(2-(7-chloroquinoline-2-yl)vinyl)phenyl)-3-(2-(2-hydroxypropyl-2-yl)phenyl)propyl)thio)methyl)cyclopropyl)acetic acid and 9,10-dimethoxy-5,6-dihydro-[1,3]dioxocyclo [4,5-g]isoquino[3,2-a]isoquinolin-7-onium conjugate (double salt 1, 600 mg, 0.65 mmol, yield: 100%/), retention time: 1.271 min. LCMS: [M+H].sup.+ 336.1. Retention time: 1.853 min. LCMS: [M+H].sup.+ 586.3.

[0090] Product NMR: .sup.1H NMR (400 MHz, DMSO) ?9.89 (s, 1H), 8.93 (s, 1H), 8.40 (d, J=8.8 Hz, 1 H), 8.20 (d, J=9.2 Hz, 1H), 8.03 (d, J=2.4 Hz, 1H), 8.01-7.94 (m, 3H), 7.89 (d, J=16.4 Hz, 1H), 7.80 (s, 1H), 7.73 (s, 1H), 7.62-7.57 (m, 2H), 7.51 (d, J=16.4 Hz, 1H), 7.40-7.33 (m, 3H), 7.11-7.00 (m, 4H), 6.17 (s, 2H), 4.94-4.91 (m, 2H), 4.08 (d, J=10.0 Hz, 6H), 4.02 (t, J=8.0 Hz, 1H), 3.25-3.19 (m, 2H), 3.13-3.06 (m, 1H), 2.75-2.65 (m, 2H), 2.54 (s, 1H), 2.29-2.20 (m, 1H), 2.10-2.06 (m, 2H), 1.95-1.91 (m, 1H), 1.45 (d, J=8.4 Hz, 6H), 0.41-0.30 (m, 2H), 0.25-0.14 (m, 2H).

Montelukast

Example 2: Preparation of Montelukast Berberine Analogue Double Salts (Double Salts 2-15)

[0091]

TABLE-US-00001 Nos. of Compounds Structure Double salt 2 [00022] Double salt 3 [00023] Double salt 4 [00024] Double salt 5 [00025] Double salt 6 [00026] Double salt 7 [00027] Double salt 8 [00028] Double salt 9 [00029] Double salt 10 [00030] Double salt 11 [00031] Double salt 12 [00032] Double salt 13 [00033] Double salt 14 [00034] Double salt 15 [00035]

Example 3: Evaluation on Anti-Inflammatory Effect of Test Compound Through Hypertonic Inflammatory Cell Model

3.1 Cells and Culture

[0092]

TABLE-US-00002 Cell line Culture method Human 12-SV40 corneal DMEM/F12 + 10% FBS + 1% ITS epithelial cell line Primary human corneal DMEM/F12 + 5% FBS + 10 ng/ml epithelial cell EGF + 1% ITS + 0.5% DMSO + 0.5 ?g/ml Hydrocortison + 50 ?g/m1 Gentamicin + 1.25 ?g/m1 amphotericin B

3.2 Reagents, Instruments and Consumables

[0093]

TABLE-US-00003 Reagents and consumables Manufacturers PBS Gibco FBS Gibco DMSO SIGMA L-glutamine Gibco DMEM/F12 Gibco Human epidermal growth factor Gibco Primers Invitrogen Centrifuge tube Falcon Inc., USA Cell culture plate Falcon Inc., USA Hanks Gibco Mycillin solution (100?) Gibco Insulin, Transferrin, Selenium, Ethanolamine Gibco Solution (ITS-X), 100? 0.25% EDTA-containing trypsin Gibco NaCl powder Sigma CCKS kit (cell counting kit-8) DOJINDO Ophthalmic scissors, ophthalmic tweezers 66 VISION TECH Hydrocortison Solarbio Gentamicin Solarbio Amphotericin B Sigma

3.3 Experimental Method and Process

[0094] SV40 cell culture: SV40 cells were cultured in a DMEM/F12 medium containing 10% fetal bovine serum, and added with insulin and a human epidermal growth factor at the same time, so that a final concentration of insulin in the medium was 5 ?g/mL and a final concentration of the human epidermal growth factor was 10 ng/mL. The medium was routinely subcultured in a cell culture incubator with a temperature of 37? C., CO2 content of 5% and a humidity of 90%. The cells were grown to a fused state of 70%-90% for an experiment.

[0095] Primary human cell culture: the remaining limbus of the donor cornea used in a surgery was treated with an ophthalmic instrument into a tissue block carrying corneal stem cells, the tissue block was pasted at the bottom of a cell culture dish, cultured in an SHEM medium, and placed in a cell culture incubator with a temperature of 37? C., CO2 content of 5% and humidity of 90%; primary human corneal epithelial cells crawling out of the limbal stem cells can be seen after 3-5 days; and the cells were grown to a fused state of 70%-90% for an experiment.

[0096] Preparation of solution: (1) preparation of hypertonic liquid (500 mOsm): 90 mmol/L NaCl solution was added to a serum-free medium. (2) Preparation of a tested drug solution: 10 mg of the above prepared product was dissolved in 1 mL of 37? C. DMSO solution, prepared into 104 ?g/mL mother liquor, and stored in a ?20? C. refrigerator for later use; and the mother liquor was diluted with a serum-free medium to prepare drug solutions having final concentrations of 1 ?mol/L and 10 ?mol/L for an experiment.

[0097] Establishment and administration of hypertonic inflammatory cell model: the old medium was discarded, and a hypertonic medium containing 500 mOsm hypertonic liquid and a normal isotonic medium were supplemented; after four hours of cell culture, a drug group was replaced with a drug solution, and the normal isotonic medium is replaced for control groups (including a hypertonic model control and a normal cell control); and materials were sampled after four hours of continuous culture.

[0098] Real-time PCR (RT-PCR): mRNA expressions of different groups of IL-6, IL-1?, IL-17A, IL-18, TNF-?, and NLRP3 were detected using RT-PCR. SV40 and primary human corneal epithelial cells inoculated in a 12-well plate at different time points (after modeling of a hypertonic inflammatory cell model and after four hours of administration) were collected for later use. The same experiment was repeated for three times. RNA was extracted from the cells according to an instruction of an RNA extraction kit, and a total amount of RNAs required to synthesize a cDNA template was calculated based on a measured concentration. cDNA was synthesized by an M-MLV reverse transcription kit, and the cDNA obtained by reverse transcription was stored at ?20? C. for subsequent PCR amplification. Gene primer sequences were as follows:

TABLE-US-00004 Gene Serial Serial name number Forwardprimers number Reverseprimers NLRP3 1 5-CGTGAGTCCCATTAAGATG 2 5-CCCGACAGTGGATATAGAA GAGT-3 CAGA-3 IL-6 3 5-TGAGAGTAGTGAGGAACA 4 5-CGCAGAATGAGATGAGTT AG-3 G-3 IL-17A 5 5-TGTCACTGCTACTGCTGCT 6 5-GGTGAGGTGGATCGGTTGT GAG-3 AGT-3; TNF-? 7 5-CCTCTCTCTAATCAGCCCT 8 5-GAGGACCTGGGAGTAGAT CTG-3 GAG-3; GAPDH 9 5-ATGTTCGTCATGGGTGTGA 10 5-GGTCCTAAGCAGTTGGTG A-3 GT-3.

[0099] A SYBR Green fluorescent dye method was used for RT-PCR detection. A relatively quantified mRNA expression quantity of a target gene was calculated. Each group was provided with three replicate wells, and a final result was taken as an average of the three times.

[0100] A t-test was used for comparison between the two groups, and P<0.05 indicated a significant difference

3.4 Results

[0101] The experimental results showed (FIG. 1: NC: normal control, HS: hypertonic model, HS-1: hypertonic model+1 ?mol/L montelukast berberine double salt drug treatment group, HS-10: hypertonic model+10 ?mol/L montelukast berberine double salt drug treatment group), in a hypertonic model of primary human corneal epithelial cells and SV40 corneal epithelial cells, the treatments with drugs containing montelukast having concentrations of 1 ?mol/L and 10 ?mol/L could significantly reduce various inflammation-related cytokines (FIG. 1: A-H, HS group vs. HS-1 and HS-10 groups, Ps<0.01). It was indicated that the montelukast berberine double salt had a good anti-inflammatory effect and thus a therapeutic potential for related diseases.

Example 4: Evaluation Comparison of Anti-Inflammatory Effects of a Test Compound and Monomers Thereof Through Hypertonic Inflammatory Cell Model

4.1 Experimental Method and Process

[0102] The reagents, instruments and consumables of this example were the same as those of Example 3, and contrasts drug used were berberine base (Aladdin, B414323), and montelukast (Aladdin, M421902).

[0103] Primary human cell culture: the remaining limbus of the donor cornea used in a surgery was treated with an ophthalmic instrument into a tissue block carrying corneal stem cells, the tissue block was pasted at the bottom of a cell culture dish, cultured in an SHEM medium, and placed in a cell culture incubator with a temperature of 37? C., CO2 content of 5% and humidity of 90%; primary human corneal epithelial cells crawling out of the limbal stem cells can be seen after 3-5 days; and the cells were grown to a fused state of 70%-90% for an experiment.

[0104] Preparation of solution: (1) preparation of hypertonic liquid (500 mOsm): 90 mmol/L NaCl solution was added to a serum-free medium. (2) Preparation of a tested drug solution: 10 mg of the above prepared product was dissolved in 1 mL of 37? C. DMSO solution, prepared into 104 ?g/mL mother liquor, and stored in a ?20? C. refrigerator for later use; and the mother liquor was diluted with a serum-free medium to prepare a drug solution having a final concentration of 0.5 ?mol/L for this experiment. (3) Preparation of a drug solution of berberine and montelukast: 10 mg of berberine or montelukast was dissolved in 1 mL of 37? C. DMSO solution, respectively prepared into 104 ?g/mL mother liquor, and stored in a ?20? C. refrigerator for later use; and the mother liquor was diluted with a serum-free medium to prepare a drug solution having a final concentration of 0.5 ?mol/L for this experiment.

[0105] Real-time PCR (RT-PCR): mRNA expressions of different groups of IL-6, IL-1?, IL-17A, IL-18, TNF-?, and NLRP3 were detected using RT-PCR. SV40 and primary human corneal epithelial cells inoculated in a 12-well plate at different time points (after modeling of a hypertonic inflammatory cell model, and four hours after administration) were taken, and cells were collected for later use. The same experiment was repeated for three times. RNA was extracted from the cells according to an instruction of an RNA extraction kit, and a total amount of RNAs required to synthesize a cDNA template was calculated based on a measured concentration. cDNA was synthesized by an M-MLV reverse transcription kit, and the cDNA obtained by reverse transcription was stored at ?20? C. for subsequent PCR amplification. A gene primer sequence was the same as that in Example 3.

[0106] A t-test was used for comparison between the two groups, and P<0.05 indicated a significant difference

4.2 Results

[0107] The experimental results showed (FIG. 3: UT: normal control, HS: hypertonic cell model, HS-0.5: hypertonic cell model+0.5 ?mol/L montelukast berberine double salt drug treatment group, A: hypertonic cell model+0.5 ?mol/L montelukast, B: hypertonic cell model+0.5 ?mol/L berberine base), in a hypertonic model of primary human corneal epithelial cells, compared with a single drug of berberine and montelukast of the same concentration, the resulting double salt drug could significantly reduce various inflammation-related cytokines (FIG. 3: A-D, HS-0.5 group vs. groups A and B, Ps<0.01). It was indicated that the anti-inflammatory effect of the montelukast berberine double salt was superior to that of a montelukast or berberine monomer.

Example 5: Stability Comparison of Montelukast Berberine Double Salt with Montelukast and Berberine Base in High Humidity Environment

5.1 Experimental method and process

[0108] In this example, a 1/10,000 electronic balance (Sartorius, Germany) was used, and contrast drugs used were the berberine base (Aladdin, B414323), and montelukast (Aladdin, M421902).

[0109] Three parts of 1 g of montelukast berberine double salt (double salt 1), three parts of 1 g of montelukast and three parts of 1 g of berberine were taken respectively, respectively placed in a flat dish, and weighed precisely; a test sample was placed in a constant humidity closed container, placed under conditions of 25? C., RH90%?5% for 10 days, and weighed precisely on Day 5 and Day 10; and weighing results were recorded. A moisture absorption gain ratio was calculated.

5.2 Results

[0110] In a high humidity environment, a moisture absorption weight gain ratio of the montelukast berberine double salt was less than that of montelukast and berberine base in a high humidity environment. The compound was stored conveniently.

TABLE-US-00005 Weight gain Weight gain percentage percentage Compound of Day 5 (%) of Day 10 (%) Montelukast berberine 3.3 4.9 double salt Montelukast 4.5 6.3 Berberine base 4.2 6.7

Example 6: Solubility Comparison of Montelukast Berberine Double Salt with Montelukast and Berberine Base in a Solvent

6.1 Experimental Method and Process

[0111] Preparation of solvent: 1) dissolution of carboxymethyl cellulose CMC (MACKLIN: C889437): a 2 L beaker was taken, about 1500 mL of ultrapure water at 80? C. was measured, and carboxymethyl cellulose CMC (3.75 g, 0.25%) was added slowly while stirring with an LED overhead stirrer, stirred for about 7 h and continued to be added with other adjuvants. 2) Addition of hydroxypropyl-?-cyclodextrin HPBCD (Bidepharm: BD44359): hydroxypropyl-?-cyclodextrin HPBCD (82.5 g, 5.5%) was added slowly and stirred for about 1 h until dissolved. 3) Adjustment of a pH value: a pH value was measured before adjustment of the pH value, and then 1 mol/L NaOH solution or 1 mol/L HCl solution was added to adjust the pH value to about 7.5-8.

[0112] Preparation of a reference substance: 1) montelukast berberine salt reference solution: an appropriate amount of montelukast berberine salt was weighed precisely and placed in a measuring flask, dissolved with methanol and diluted to a desired scale. 2) Berberine base reference solution: an appropriate amount of berberine reference substance was weighed precisely and placed in a measuring flask, dissolved with methanol and diluted to a desired scale. 3) Montelukast reference substance solution: 3) a montelukast reference substance was weighed precisely and placed in a measuring flask, dissolved with DMSO and diluted to a desired scale. 4) Test solution: the montelukast berberine salt, the berberine base, and the montelukast were respectively added to a solvent to be supersaturated, and filtered, and pH was adjusted to 7.5-8 with a sodium hydroxide or hydrochloric acid solution.

[0113] Chromatographic conditions: 1) chromatographic conditions of montelukast berberine salt: a chromatographic column used was Agilent ZORBAX SB-C18 (4.6?250 mm, 5 ?m); a mobile phase: 0.01 mol/L ammonium dihydrogen phosphate solution (pH was adjusted to 2.8 with phosphoric acid)-acetonitrile (75:25); a detection wavelength: 345 nm; an injection volume: 10 ?L; a column temperature: 25? C.; and a flow rate: 1.0 mL/min. A test solution and a reference solution of montelukast berberine salt were measured precisely and injected into a liquid chromatograph respectively. 2) Chromatographic conditions of montelukast: a chromatographic column used was Agilent Eclipse XDB-C18 (4.6?150 mm, 5 ?m); a mobile phase A: 3.85 g of ammonium acetate was weighed and dissolved with 1000 mL of water, added with 1 mL of triethylamine, adjusted for a pH value to 5.5 with glacial acetic acid, filtered, and ultrasonically treated; a mobile phase B: methanol; a detection wavelength: 240 nm; an injection volume: 20 ?L; a column temperature: 25? C.; and a flow rate: 1.0 mL/min. Determination method: a montelukast reference solution was measured precisely and injected into a liquid chromatograph; and various test solutions of montelukast were appropriately diluted and injected into the liquid chromatograph. 3) Chromatographic conditions of berberine base: a chromatographic column used was Agilent ZORBAX SB-C18 (4.6?250 mm, 5 ?m); a mobile phase: 0.01 mol/L ammonium dihydrogen phosphate solution (pH was adjusted to 2.8 with phosphoric acid)-acetonitrile (75:25); a detection wavelength: 345 nm; an injection volume: 10 ?L; a column temperature: 25? C.; and a flow rate: 1.0 mL/min. A test solution and a reference solution of berberine base were measured precisely and injected into a liquid chromatograph respectively.

[0114] A t-test was used for comparison between the two groups, and P<0.05 indicated a significant difference

6.2 Results

[0115]

TABLE-US-00006 Compound Concentration in solvent (?g/ml) Montelukast berberine double salt 31.51 Montelukast Not detected for very small amount Berberine base 24.45

[0116] The results showed that the solubility of the montelukast berberine double salt in this solvent was superior that of the berberine base, and significantly superior to that of montelukast monomer (Ps<0.05).

Example 7: Stability Comparison of Montelukast Berberine Double Salt with Montelukast and Berberine Base in the Solvent

7.1 Experimental Method and Process

[0117] The reagents, instruments and consumables in this example were the same as those of Example 6, because the solubility of montelukast in the solvent was extremely poor, so montelukast was not be detected out. Therefore, the stability of the montelukast in the solvent was compared by using a sodium salt (Aladdin: M129586) and the montelukast berberine double salt.

[0118] Preparation of solvent: 1) dissolution of carboxymethyl cellulose CMC (MACKLIN: C889437): a 2 L beaker was taken, about 1500 mL of ultrapure water at 80? C. was measured, and carboxymethyl cellulose CMC (3.75 g, 0.25%) was added slowly while stirring with an LED overhead stirrer, stirred for about 7 h and continued to be added with other adjuvants. 2) Addition of hydroxypropyl-?-cyclodextrin HPBCD (Bidepharm: BD44359): hydroxypropyl-?-cyclodextrin HPBCD (82.5 g, 5.5%) was added slowly and stirred for about 1 h until dissolved. 3) Adjustment of a pH value: a pH value was measured before adjustment of the pH value, and then 1 mol/L NaOH solution or 1 mol/L HCl solution was added to adjust the pH value to about 7.5-8.

[0119] Preparation of a reference substance: 1) montelukast berberine salt reference solution: an appropriate amount of montelukast berberine salt was weighed precisely and placed in a measuring flask, dissolved with methanol and diluted to a desired scale. 2) Berberine base reference solution: an appropriate amount of berberine reference substance was weighed precisely and placed in a measuring flask, dissolved with methanol and diluted to a desired scale. 3) Montelukast sodium reference substance solution: an appropriate amount of montelukast sodium reference substance was weighed precisely and placed in a measuring flask, dissolved with DMSO and diluted to a desired scale. 4) Test solution: the montelukast berberine salt, the berberine base, and montelukast sodium were respectively added to a solvent to be supersaturated, and filtered, and pH was adjusted to 7.5-8 with a sodium hydroxide or hydrochloric acid solution.

[0120] Chromatographic conditions: 1) chromatographic conditions of montelukast berberine salt: a column used was Agilent ZORBAX SB-C18 (4.6?250 mm, 5 ?m); a mobile phase: 0.01 mol/L ammonium dihydrogen phosphate solution (pH was adjusted to 2.8 with phosphoric acid)-acetonitrile (75:25); a detection wavelength: 345 nm; an injection volume: 10 ?L; a column temperature: 25? C.; and a flow rate: 1.0 mL/min. A test solution and a reference solution of montelukast berberine salt were measured precisely and injected into a liquid chromatograph respectively. 2) Chromatographic conditions of montelukast sodium: a chromatographic column used was Agilent Eclipse XDB-C18 (4.6?150 mm, 5 ?m); a mobile phase A: 3.85 g of ammonium acetate was weighed and dissolved with 1000 mL of water, added with 1 mL of triethylamine, adjusted for a pH value to 5.5 with glacial acetic acid, filtered, and ultrasonically treated; a mobile phase B: methanol; a detection wavelength: 240 nm; an injection volume: 20 ?L; a column temperature: 25? C.; and a flow rate: 1.0 mL/min. Determination method: a montelukast sodium reference solution was measured precisely and injected into a liquid chromatograph; and various test product solutions of montelukast sodium were appropriately diluted and injected into the liquid chromatograph. Chromatographic conditions of berberine base: a chromatographic column used was Agilent ZORBAX SB-C18 (4.6?250 mm, 5 ?m); a mobile phase: 0.01 mol/L ammonium dihydrogen phosphate solution (pH was adjusted to 2.8 with phosphoric acid)-acetonitrile (75:25); a detection wavelength: 345 nm; an injection volume: 10 ?L; a column temperature: 25? C.; and a flow rate: 1.0 mL/min. A test solution and a reference solution of berberine base were measured precisely and injected into a liquid chromatograph respectively.

[0121] Experimental process: 1) photostability experiment: a test solution of montelukast berberine salt at 0 h, a test solution of berberine base at 0 h, and a test solution of montelukast at 0 h were taken respectively, and the above solutions were placed under an illumination condition; samples were taken at 8 h, Day 5 and Day 10 respectively; and test solutions of various substances under various conditions were obtained and compared with a standard solution. 2) Phototability experiment: a test solution of montelukast berberine salt at 0 h, a test solution of berberine base at 0 h, and a test solution of montelukast at 0 h were taken respectively, and the above solutions were placed under conditions of 40? C. (in dark place) and 60? C. (in dark place); samples were taken at 8 h, Day 5 and Day 10 respectively; and test solutions of various substances under various conditions were obtained and compared with a standard solution.

[0122] A t-test was used for comparison between the two groups, and P<0.05 indicated a significant difference.

7.2 Results

[0123]

TABLE-US-00007 C(?g/ml) Purity Montelukast 0:00 31.51 94.53% berberine salt Illumination 40? C. 60? C. C(?g/ml) Purity C(?g/ml) Purity C(?g/ml) Purity 8:00 24.68 92.80% 26.8 94.22% 28.81 93.01% Day 5 15.17 85.74% 26.57 93.19% 28.41 93.00% Day 10 9.75 79.56% 12.27 92.68% 13.70 92.90% C(?g/ml) Purity Berberine 0:00 24.45 97.35% base Illumination 40? C. 60? C. C(?g/ml) Purity C(?g/ml) C(?g/ml) Purity C(?g/ml) 8:00 21.84 97.62% 22.62 97.17% 22.45 97.17% Day 5 14.70 88.52% 22.41 97.02% 21.31 96.96% Day 10 7.02 74.89% 10.35 96.86% 9.68 95.99% C(?g/ml) Purity Montelukast 0:00 3706.643 98.33% sodium Illumination 40? C. 60? C. C(?g/ml) Purity C(?g/ml) C(?g/ml) Purity C(?g/ml) 8:00 1444.936 62.47% 3587.359 97.77% 3448.598 96.72% Day 5 126.667 23.07% 1388.78 94.61% 534.2 86.43% Day 10 51.213 20.11% 724.31 77.10% 215.12 68.13%

[0124] The results showed that the overall photothermal stability of the montelukast berberine double salt in this solvent of common eye drops at three time points of 8 h, Day 5 and Day 10 was superior to that of montelukast (Ps<0.05), and the photothermal stability was equivalent to that of the berberine base (Ps>0.05).

[0125] Embodiments of the present invention will be described above. However, the present invention is not limited to the above embodiments. Thus, any modification, equivalent replacement, improvement and so on made within the spirit and principle of the present invention shall be encompassed by the protection scope of the present invention.