Title of Invention

"AN IMPROVED PROCESS FOR THEPREPARATION OF 4,4-DIMETHOXYTRITYL CHLORIDE (DMTR CL)

Abstract

An improved process for the preparation of 4,4'-dimethoxytrityl chloride which comprises : (i) Mixing benzotrihalide with lewis acid catalyst at room temperature, (ii) cooling the resultant reaction mixture to a temperature in the range of 0 to 30 C, (iii) adding anisole dropwise over a period in the range of 1 to 2 hours while stirring the reaction mixture at room temperature for a period in the range of 30 to 60 min, (iv) refluxing the resulting mixture for a period in the range of 2 to 3 hours, (v) cooling of the resulting mixture, (vi) hydrolyzing the Lewis acid complex formed by known methods, (vii) extracting the 4,4'-dimethoxytritanol with a hydrocarbon solvent followed by washing of hydrocarbon phase with an alkaline solution and water, respectively. (viii) separating the organic phase, drying over anhydrous sodium sulphate followed by concentrating the organic phase on a rotary evaporator under reduced pressure to yield the 4,4'-dimethoxytritanol, (ix) converting the 4,4'-dimethoxytritanol so obtained to 4,4'-dimethoxytrityl chloride by addition of an organic acid chloride in the presence of a hydrocarbon solvent followed bv cooling to a temperature in the range of 4 to 6 C, (x) recovering the 4,4'-dimethoxytrityl chloride by washing with an organic solvent under nitrogen atmosphere and drying under vacuum.

Full Text

This invention relates to an improved process for the preparation of 4,4/-dimethoxytrityl chloride (DMTr CI). The 4,4'-dimethoxytrityl chloride prepared by the process of the present invention is an important reagent for oligonucleotide synthesis. Trityl chlorides are useful reagents and are known for a long time in carbohydrate chemistry for the selective protection of primary hydroxyl groups in the presence of secondary and tertiary hydroxyl groups. Because of their chemical selectivity, they have great potential in chemical synthesis of various molecules of biological or abiological importance. The use of these reagents in nucleic acids chemistry was first reported by Khorana and coworkers for the selective protection of 5'-hydroxyl function of ribdse or 2'-deoxyribose in nucleosides. Since then, these are in use as protective groups in oligonucleotide synthesis. The synthesis of 4,4'-dimethoxytrityl chloride (DMTr-Cl), reported in the literature, involves the preparation of two equivalents of Grignard reagent from p-bromoanisole and magnesium turnings in an anhydrous solvent followed by its reaction with ethyl or methyl benzoate (E.G. Khorana, J.Am. Chem. Soc, 1962, 84, 430). The procedure yields 4,4'-dimethoxytritanol, which is then converted into 4,4'-dimethoxytrityl chloride, DMTr-Cl using acetyl chloride in benzene. The main drawbacks of this method are, (i) final product is obtained in the range of 55-60%, (ii) strictly anhydrous conditions are required for the reaction and (iii) incomplete conversion of Grignard reagent may lead to the formation of side products. The main objective of the present invention is to provide an improved process for the synthesis of 4,4'-dimethoxytrityl chloride. Another objective of the present invention is to provide an improved process for the synthesis of 4,4'-dimethoxytrityl chloride using inexpensive reagents and solvents under normal laboratory conditions. Yet another objective of the present invention is to provide a process for the synthesis of 4,4'-dimethoxytrityl chloride, which could be upscaled for large scale synthesis of the same. Still another objective of the present invention is to provide a simple, rapid and economical process for the preparation of 4,4'-dimethoxytrityl chloride. The present invention is primarily based on Frie-del-Craft Arylation reaction employing Lewis Acid as a catalyst. The process of the present invention comprises the formation of a complex between Lewis acid and benzo-trihalide which facilitates the reaction with anisole. The resulting 4,4'-dimethoxytritanol obtained is then converted to dimethoxytrityl chloride by the reaction of acetyl chloride in a hydrocarbon. Accordingly, the present invention provides an improved process for the preparation of 4,4'-dimethoxy- trityl chloride (DMTr CI) which comprises : (i) Mixing benzotrihalide with lewis acid catalyst at room temperature, (ii) cooling the resultant mixture to a temperature in o the range of 0 to 30 C, (iii) adding anisole dropwise over a period in the range of 1 to 2 hours while stirring the reaction mixture at room temperature for a period in the range of 30 to 60 min, (iv) refluxing the resulting mixture for a period in the range of 2 to 3 hours, (v) cooling of the resulting mixture, (vi) hydrolyzing the Lewis acid complex formed by known methods, (vii) extracting the 4,4'-dimethoxytritanol with a hydrocarbon solvent followed by washing of hydrocarbon phase with an alkaline solution and water, respectively, (viii) separating the organic phase, drying over anhydrous sodium sulphate followed by concentrating the organic phase on a rotary evaporator under reduced pressure to yield the 4,4'-dimethoxytritanol, (ix) converting the 4,4'-dimethoxytritanol so obtained to 4,4'-dimethoxytrityl chloride by addition of an organic aci-d chloride in the presence of a hydrocarbon solvent followed by cooling to a temperature in the o range of 4 to 6 C, (x) recovering the 4,4/-dimethoxytrityl chloride by washing with an organic solvent under nitrogen atmosphere and drying under vacuum, The solvent system employed for the preparation of dimethoxytrityl chloride may be such as carbon disulfide, nitrobenzene, nitrobenzaldehyde. The catalyst used may be such as Aiei , AlBr , The dropwise addition of anisole may be effected preferably within a period of lh to 1.5h. The alkaline solution used in step (vii) may be such as sodium bicarbonate, potassium bicarbonate, cesium bicarbonate. The organic acid chloride used in step (ix) may be such as acetyl chloride, propionyl chloride, oxalyl chloride. The hydrocarbon solvent used in step (ix) may be such as benzene, toluene, xylene. The organic solvent used in step (x) may be such as hexane, pentane, petroleum ether. The invention is described in detail in the following examples itfhich are given by way of illustration and should not be construed to limit the scope of the present invention. EXAMPLE 1 1 Mol of benzotrichloride was mixed with 2.2 mol of anisole in a 2 liters of carbon disulfide at room temperature in a three necked round bottom flask fitted with a condenser, a gas inlet tube for nitrogen gas and a device for the addition of powdered aluminium chloride. The solution was cooled to 5 C and added 1.0 mol of anhydrous aluminium chloride in small portions. After complete addition of aluminium chloride, the mixture was stirred at room temperature for 30 min. Then it was allowed to reflux for 2h. Again it was cooled in an ice bath and the complex was destroyed by the addition of aqueous saturated ammonium chloride (250 ml). The mixture so obtained was then partitioned between benzene and water. The organic layer was separated and concentrated on an evaporator under reduced pressure to obtain 4,4'-dimethoxytritanol, which was converted into its chloride form by treating it with acetyl chloride in benzene (i.e. activation) at 4 C. The crystallized product, 4 ,4'--dimethoxy-trityl chloride, was recovered in a sintered glass column under nitrogen atmosphere, washed with petroleum ether (2 1) and dried under vacuum to obtain 4,4'-dimethoxytrityl chloride in 80% yield. The product was characterized spectroscopically and found to be identical with the authentic sample prepared by the procedure of Khorana, H.G., J.Am. Chem. Soc., 1962, %4, 430. EXAMPLE 2 1 Mol of benzotrichloride was mixed with 2.2 mol of anisole in a 2 liters of nitrobenzene at room temperature in a three necked round bottom flask fitted with a condenser, a gas inlet tube for argon and a device for the addition of powdered aluminium chloride. The solution was cooled to 5 C and added 1 mol of anhydrous aluminium chloride in small portions. After complete addition of aluminium chloride, the mixture was stirred at room temperature for 30 min. Then it was allowed to reflux for 2h. Again it was cooled in an ice bath and the aluminium chloride complex was destroyed by the addition of aqueous saturated ammonium chloride (500 ml). The mixture so obtained was then partitioned between benzene and water. The organic layer was separated and concentrated on a rotary evaporator under reduced pressure to obtain 4,4'-dimethoxytritanol, which was reacted with acetyl chloride in benzene to obtain 4,4/-dimethoxytrityi chloride in 75% yield. The product was characterized by spectroscopic techniques and found to be identical to the authentic sample prepared by the procedure of Khorana,H.G., J.Am. Chem. Soc, 1962, 84, 430. EXAMPLE 3 Benzotrichloride (1 mol) was mixed with 0.5 mol of aluminium chloride in 1 liter of carbon disulfide at room temperature in a three necked round bottom flask fitted with a condenser, a gas inlet tube for nitrogen and a pressure equalizing funnel for addition of ani-sole. The solution was cooled to 5 C and added 2.1 mol of anisole mixed in carbon disulfide (500 ml) dropwise over a period of lh. After complete addition of anisole, the mixture was stirred at room temperature for 20 min. Then it was allowed to reflux for 3h. Again it was cooled in an ice bath and reaction mixture was poured into cone, sulfuric acid-ice mixture. The mixture so obtained was then partitioned between benzene and water, the organic layer separated and concentrated on an evaporator under reduced pressure to obtain 4,4,-dimethoxytritanol. Finally, conversion of 4,4/-dimethoxytritanol to 4,4'-dimethoxytrityl chloride was carried out by the treatment acetyl chloride in benzene. The final product, 4,4'-dimethoxytrityl chloride, was obtained in 86% yield. The product was characterized spectroscopically and found to be identical to the authentic sample prepared by the procedure of Khorana,H.G., J.Am. Chem. Soc, 1962, 84, 430. EXAMPLE 4 Benzotrichloride (1 mol) was mixed with 0.5 mol of aluminium chloride in nitrobenzene (1 1) at room temperature in a three necked round bottom flask fitted with a condenser, a gas inlet tube for nitrogen and a pressure equalizing funnel for addition of anisole. The solution was cooled to 20 C and added slowly 2.2 mol of anisole mixed with nitrobenzene (500 ml) dropwise over a period of 1.5h. After complete addition of anisole, the mixture was stirred at room temperature for 30 min. Then it was allowed to reflux for 2h. Again it was cooled in an ice bath and the complex was destroyed by addition of aqueous saturated ammonium salt. The mixture so obtained was then partitioned between benzene and water, the organic layer separated and concentrated on an evaporator under reduced pressure to obtain 4,4'-dimethoxytritanol. Finally, conversion of 4,4'-dimethoxytritanol to trityl chloride was effected by adding acetyl chloride in benzene solution. After crystallization, the product, 4,4'~dimethoxytrityl chloride, was obtained in 91% yield. The product was characterized by spectroscopically and found to be identical to the authentic sample prepared by the procedure of Khorana,H.G., J.Am. Chem. Soc, 1962, 84, 430. EXAMPLE 5 Ferric chloride (0.75 mol) and benzotrichloride (1 mol) were mixed in carbon disulfide (1 1) at room temperature in a three necked round bottom flasX fitted with a condenser, a gas inlet tube for nitrogen and a pressure equalizing funnel for addition of anisole. The reaction mixture was cooled to 5 C and added anisole (2.2 mol) in carbon disulfide (500 ml) dropwise over a period of lh. After complete addition of anisole, the mixture was allowed to stir at room temperature for 20 min. Then it was refluxed for 3h. Again it was cooled in an ice bath and added aqueous saturated ammonium chloride. The mixture so obtained was partitioned between benzene and water. The organic layer is separated out and concentrated on rotary evaporator under reduced pressure to get 4,4/-dimethoxytritanol. In the final step, dimethoxytrityl chloride was obtained by treating 4,4'-dimethoxytritanol in benzene solution with acetyl chloride. After crystallization at 4C, the product, 4,4'-dimethoxytrityl chloride, was obtained in 85% yield. The product was characterized by spectroscopically and found to be identical to the authentic sample prepared by the procedure of Khorana,H.G., J.Am. Chem. Soc, 1962, 84, 430. EXAMPLE 6 1 mol of benzotrichloride was mixed with 0.5 mol of aluminium chloride at room temperature in a three necked round bottom flask fitted with a condenser, a gas inlet tube for nitrogen and a pressure equalizing funnel for addition of anisole. The solution was cooled to 15 C and added 2.2 mol of anisole dropwise over a period of lh. After complete addition of anisole, the mixture was stirred at room temperature for 20 min. Then it was refluxed on a heating mantle for 3h. Again it was cooled in an ice bath and added aqueous saturated ammonium salt. The mixture so obtained was then partitioned between benzene and water, the organic layer separated out and concentrated on a rotavapor under reduced pressure. Final conversion of 4,4'-dimethoxytritanol to 4,4/-dimethoxytrityl chloride was effected by adding acetyl chloride in benzene solution. The product, 4,4'-dimethoxytrityl chloride crystallized at 4C was then washed with about 2 liters of petrol and dried in a vacuum desiccator to obtain in 93% yield. The product was characterized by spectroscopically and found to be identical to the authentic sample prepared by the procedure of Khorana,H.G., J.Am. Chem. Soc, 1962, 84, 430. Advantages of the invention 1. The method is cheaper and reproducible. 2. The method does not require scrupulously anhydrous conditions hence laboratory grade solvents can be employed. 3. The yields are obtained between 90-95% from the starting material. We claim 1. An improved process for the preparation of 4,4'- dimethoxytrityl chloride which comprises : (i) Mixing benzotrihalide with lewis acid catalyst at room temperature, (ii) cooling the resultant reaction mixture to a tem- o perature in the range of 0 to 30 C, (iii) adding anisole dropwise over a period in the range of 1 to 2 hours while stirring the reaction mixture at room temperature for a period in the range of 30 to 60 min, (iv) refluxing the resulting mixture for a period in the range of 2 to 3 hours, (v) cooling of the resulting mixture, (vi) hydrolyzing the Lewis acid complex formed by known methods, (vii) extracting the 4,4/-dimethoxytritanol with a hydrocarbon solvent followed by washing of hydrocarbon phase with an alkaline solution and water, respectively, (viii) separating the organic phase, drying over anhydrous sodium sulphate followed by concentrating the organic phase on a rotary evaporator under reduced pressure to yield the 4,4/-dimethoxytritanol, (ix) converting the 4,4'-dimethoxytritanol so obtained to 4,4'-dimethoxytrityl chloride by addition of an organic acid chloride in the presence of a hydrocarbon solvent followed by cooling to a temperature in the range of 4 to 6 C, (x) recovering the 4,4'-dimethoxytrityl chloride by washing with an organic solvent under nitrogen atmosphere and drying under vacuum. 2. An improved process as claimed in claim 1 wherein the reaction in step (i) is carried out in the presence of organic solvent such as carbon disulfide, nitrobenzene. 3. An improved process as claimed in claims 1 & 2 wherein the Lewis acid catalyst used is such as alumin-lum Kchloride, aluminiumbromide, zinc chloride, boron trichloride, boron trifluoride. 4. An improved process as claimed in claims 1-3 wherein the alkaline solution used in step (vii) is used such as sodium bicarbonate, potassium bicarbonate, cesium bicarbonate. 5. An improved process as claimed in claims 1-4 wherein the organic acid chloride used in step (ix) is selected from acetyl chloride, propionyl chloride, oxalyl chloride. 6. An improved process as claimed in claims 1-5 wherein the hydrocarbon solvent used in step (ix) is such as benzene, toluene, xylene. 7. An improved process as claimed in claims 1-6 wherein the organic solvent used in step (x) for washing is such as hexane, pentane, petroleum ether. 8. An improved process for the preparation of 4,4'-dimethoxytrityl chloride substantially as herein described with reference to the examples.

Documents:

2339-del-1996-abstract.pdf

2339-del-1996-claims.pdf

2339-del-1996-correspondence-others.pdf

2339-del-1996-correspondence-po.pdf

2339-del-1996-description (complete).pdf

2339-del-1996-form-1.pdf

2339-del-1996-form-2.pdf

2339-del-1996-form-4.pdf