METHODS FOR SYNTHESIZING AND PURIFYING AMINOALKYL TETRACYCLINE COMPOUNDS
Methods for the synthesis and purification of 9-amino alkyl tetracycline compounds are described.
1 . A method of purifying an alkylaminomethyl minocycline compound, comprising:
a) injecting a low pH aqueous solution of said alkylaminomethyl minocycline compound into a liquid chromatography device in a polar organic solvent gradient, and combining the product fractions;
b) adjusting the pH of said product fractions to 4.0-4.5;
c) washing said product fractions with a first non-polar organic solvent to form a first organic layer and a first aqueous layer, and discarding said first organic layer;
d) adjusting the pH of said first aqueous layer to 7.5-8.5; and
e) washing said first aqueous layer with a second non-polar organic solvent to form a second organic layer and a second aqueous layer, and discarding said second aqueous layer,
such that said alkylaminomethyl minocycline compound is purified.
2 . The method of claim 1 , wherein said low pH aqueous solution has a pH of 2-3.
3 . The method of claim 1 , wherein said low pH aqueous solution comprises methyl sulfonic acid.
4 . The method of claim 1 , wherein said polar organic solvent is acetonitrile.
5 . The method of claim 1 , wherein said pH in step b) or d) is adjusted with a base.
6 . The method of claim 5 , wherein said base is selected from the group consisting of metal hydroxide, metal carbonate, metal bicarbonate, ammonia, organic primary amine, organic secondary amine and organic tertiary amine
7 . The method of claim 6 , and wherein said metal is selected from the group consisting of lithium, sodium, potassium, calcium, magnesium and aluminum.
8 . The method of claim 6 , wherein said base is sodium hydroxide or ammonia.
9 . The method of claim 1 , wherein said first non-polar organic solvent is methylene chloride.
10 . The method of claim 1 , wherein said first organic layer comprises by-products, hydrophobic impurities and oxidative degradents of said alkylaminomethyl minocycline compound.
11 . The method of claim 1 , wherein said second non-polar organic solvent is methylene chloride.
12 . The method of claim 1 , wherein said second aqueous layer comprises by-products and β epimer of said alkylaminomethyl minocycline compound.
13 . The method of claim 1 , wherein an antioxidant is added.
14 . The method of claim 13 , wherein said antioxidant is ammonium sulfite, sodium sulfite, bisulfite or meta bisulfite.
15 . The method of claim 1 , wherein said alkylaminomethyl minocycline compound is:
wherein R A is alkyl, and R B is hydrogen or alkyl.
16 . The method of claim 15 , wherein R B is hydrogen.
17 . The method of claim 16 , wherein R A is alkyl.
18 . The method of claim 17 , wherein said alkyl is (CH 3 ) 3 CCH 2 —.
19 . The method of claim 1 , wherein said alkylaminomethyl minocycline compound is:
20 . The method of claim 1 , wherein hydrophobic impurities and oxidative degradents are removed from said alkylaminomethyl minocycline compound.
21 . The method of claim 1 , wherein by-products and β-C-4 epimer are removed from said alkylaminomethyl minocycline compound.
22 . The method of claim 1 , wherein said alkylaminomethyl minocycline compound is essentially free of hydrophobic impurities and oxidative degradents.
23 . The method of claim 1 , wherein said alkylaminomethyl minocycline compound is essentially free of by-products, β-C-4 epimer, hydrophobic impurities and oxidative degradents.
24 . The method of claim 1 , wherein said alkylaminomethyl minocycline compound comprises at least 50% α-C-4 epimer.
25 . The method of claim 24 , wherein said alkylaminomethyl minocycline compound comprises at least 95% α-C-4 epimer.
26 . The method of claim 25 , wherein said alkylaminomethyl minocycline compound comprises at least 99.9% α-C-4 epimer.
27 . The method of claim 1 , wherein said alkylaminomethyl minocycline compound comprises less than 7% β-C-4 epimer.
28 . The method of claim 27 , wherein said alkylaminomethyl minocycline compound comprises less than 3% β-C-4 epimer.
29 . A pharmaceutical composition comprising alkylaminomethyl minocycline compound purified by the method of claim 1 and a pharmaceutically acceptable carrier.