Claims
- 1. A method for preparing sphingosine-1-phosphate essentially free of L-threo isomer comprising
- (1) protecting the C1 and C3 hydroxyl groups of sphingosine (SPN);
- (2) deprotecting the C1 hydroxyl group of SPN;
- (3) phosphorylating the C1 hydroxyl group of SPN; to produce sphingosine-1-phosphate essentially free of L-threo isomer; and
- (4) deprotecting the C3 hydroxyl group of SPN.
- 2. A method for chemically synthesizing sphingosine-1-phosphate comprising:
- (A) esterifying the C-3 hydroxyl group of compound 1' to give compound 2' ##STR1## wherein X represents a moiety that protects the amino group formed in step (B),
- (B) selectively deprotecting the C-1 hydroxyl group of said compound 2' to give compound 3' ##STR2## (C) phosphorylating the C-1 hydroxyl group of said compound 3', followed by, hydrolyzing to give compound 4' ##STR3## (D) deprotecting the C-3 hydroxyl group and the amino group, respectively, of said compound 4' to give compound 1, which is said sphingosine-1-phosphate ##STR4##
- 3. The method of claim 2, wherein:
- (1) X represents N-tert-butyloxycarbonyl;
- (2) said esterifying step (A) comprises treating said compound 1' with pivaloyl chloride in dry pyridine;
- (3) said deprotecting step (S) comprises treating said compound 2' with p-toluenesulfonic acid in methanol;
- (4) said phosphorylating and hydrolyzing step (C) comprises treating said compound 3' with:
- (a) phosphorous oxychloride in the presence of triethylamine and CH.sub.2 Cl.sub.2, and then
- (b) 1N HCl in CHCl.sub.3 ; and
- (5) said deprotecting step (D) comprises treating said compound 4' with:
- (a) tetrabutylammonium hydroxide in aqueous dioxane, AMBERLITE IR-120, water, and then
- (b) trifluoroacetic acid and CH.sub.2 Cl.sub.2.
- 4. A method for chemically synthesizing N, N-dimethyl-sphingosine-1-phosphate comprising:
- (A) esterifying the C-3 hydroxyl group of compound 1' to give compound 2' ##STR5## wherein X represents a moiety that protects the amino group formed in step (B),
- (B) selectively deprotecting the C-1 hydroxyl group of said compound 2' to give compound 3' ##STR6## (C) eliminating the protecting moiety X from said compound 3' and then subjecting the product to reductive methylation to give compound 3.sup.a ##STR7## (D) phosphylating the C-1 hydroxyl group of said compound 3.sup.a followed by hydrolyzing to give compound 3.sup.b ##STR8## (E) deprotecting the C-3 hydroxyl group to give compound 2, which is said N,N-dimethyl-sphingosine-1-phosphate ##STR9##
- 5. The method of claim 4, wherein:
- (1) X represents N-tert-butyloxycarbonyl,
- (2) said esterifying step (A) comprises treating said compound 1' with pivaloyl chloride in dry pyridine;
- (3) said deprotecting step (B) comprises treating said compound 2' with p-toluenesulfonic acid in methanol;
- (4) said eliminating and reductive methylation step (C) comprises treating said compound 3' with:
- (a) trifluoroacetic acid in CH.sub.2 Cl.sub.2, and then
- (b) 37% CH.sub.2 O and NaCNBH.sub.3 in sodium acetate aqueous buffer;
- (5) said phosphylating and hydrolyzing step (D) comprises treating said compound 3.sup.a with:
- (a) phosphorous oxychloride in the presence of triethanolamine and CH.sub.2 Cl.sub.2, and then
- (b) 1N HCl in CHCl.sub.2 ; and
- (6) said deprotecting step (E) comprises treating said compound 3.sup.b with tetrabutylammonium hydroxide in aqueous dioxane.
- 6. A method for chemically synthesizing N,N,N-trimethyl-sphingosine-1-phosphate comprising:
- (A) esterifying the C-3 hydroxyl group of compound 1' to give compound 2' ##STR10## wherein X represents a moiety that protects the amino group formed in step (B),
- (B) selectively deprotecting the C-1 hydroxyl group of said compound 2' to give compound 3' ##STR11## (C) eliminating the protecting moiety X from said compound 3' and then subjecting the product to reductive methylation to give compound 3.sup.a ##STR12## (D) permethylating said compound 3.sup.a followed by treating with a basic anion exchange resin to give compound 3.sup.h ##STR13## (E) phosphylating the primary hydroxyl group of said compound 3.sup.h followed by hydrolyzing and then deprotecting the C-3 hydroxyl to give compound 3, which is said N,N,N-trimethyl-sphingosine-1-phosphate ##STR14##
- 7. The method of claim 6, wherein:
- (1) X represents N-tert-butyloxycarbonyl,
- (2) said esterifying step (A) comprises treating said compound 1' with pivaloyl chloride in dry pyridine;
- (3) said deprotecting step (B) comprises treating said compound 2' with p-toluenesulfonic acid in methanol;
- (4) said eliminating and reductive methylation step (C) comprises treating said compound 3' with:
- (a) trifluoroacetic acid in CH.sub.2 Cl.sub.2, and then
- (b) 37% CH.sub.2 O and NaCNBH.sub.3 in sodium acetate aqueous buffer;
- (5) said permethylating step (D) comprises treating said compound 3.sup.a with:
- (a) CH.sub.3 I in NaHCO.sub.3 and CHCl.sub.3, and then
- (b) DOWEX 1.times.2 (Cl.sup.-), and
- (6) said phosphorylating, hydrolyzing, and deprotecting step (E) comprises treating said compound 3.sup.h with:
- (a) POCl.sub.3 in triethyl amine and CH.sub.2 Cl.sub.2, then
- (b) 1N HCl and CHCl.sub.3, and then
- (c) tetrabutylammonium hydroxide in aqueous dioxane.
- 8. A method for chemically synthesizing N-acyl and N-acetylsphingosine-1-phosphate comprising:
- (A) esterifying the C-3 hydroxyl group of compound 1' to give compound 2' ##STR15## wherein X represents a moiety that protects the amino group formed in step (B),
- (B) selectively deprotecting the C-1 hydroxyl group of said compound 2' to give compound 3' ##STR16## (C) eliminating the protecting moiety X, and then acylating or acetylating the unprotected amino group to give compound 3.sup.i ##STR17## wherein Ac represents acyl or acetyl, (D) phosphylating the C-1 hydroxyl group of said compound 3.sup.i, followed by hydrolyzing to give compound 3.sup.j ##STR18## (E) deprotecting the C-3 hydroxyl group to give compound 4, which is said N-acyl or N-acetylsphingosine-1-phosphate ##STR19##
- 9. The method of claim 8, wherein:
- (1) X represents N-tert-butyloxycarbonyl;
- (2) said esterifying step (A) comprises treating said compound 1' with pivaloyl chloride in dry pyridene;
- (3) said deprotecting step (B) comprises treating said compound 2' with p-toluenesulfonic acid in methanol;
- (4) said eliminating and acylating or acetylating step (C) comprises treating said compound 3' with:
- (a) trifluoroacetic acid in CH.sub.2 Cl.sub.2, and then
- (b) CH.sub.3 (CH.sub.2).sub.n COCl, wherein n represents 0 to 22, in aqueous 55% K.sub.2 CO.sub.3 tetrahydrofuran;
- (5) said phosphylating and hydrolyzing step (D) comprises treating said compound 3.sup.i with:
- (a) phosphorous oxychloride in the presence of triethylamine and CH.sub.2 Cl.sub.2, and then
- (b) 1N HCl in CHCl.sub.3 ; and
- (6) said deprotecting step (E) comprises treating said compound 3.sup.j with tetrabutylammonium hydroxide in dioxane.
Parent Case Info
This is a divisional of application Ser. No. 07/863,179 filed Apr. 3, 1992 U.S. Pat. No. 5,260,288.
US Referenced Citations (1)
Number |
Name |
Date |
Kind |
5110987 |
Liotta et al. |
May 1992 |
|
Non-Patent Literature Citations (2)
Entry |
"Enzymatic Quantificatin of Sphingosine in the Picomole Range in Cultured Cells", Van Veldhoven, et al., Analytical Biochemistry 183, 177-189 (1989). |
Chemical Abstracts 1980, 92(5), No. 41295; Bushnev, A. S. et al. Bioorg. Khim. 1979, 5(9), 1381-1384. |
Divisions (1)
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Number |
Date |
Country |
Parent |
863179 |
Apr 1992 |
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