Process for preparation of labisia pumila extract

Abstract

A process for the preparation of Labisia pumila extract from Labisia pumila plant material and to obtain an optimized and reproducible chemical profile by high performance liquid chromatography.

1 . A process for obtaining optimized and reproducible chemical profile of Labisia pumila extract comprising the steps of:
 a) centrifuging a mixture of water and dried extract obtained by extracting dried  Labisia pumila  plant material with water in a ratio of 1:6 of dried Labisia pumila plant material:water to form a water-soluble extract and drying the extract obtained;   b) diluting the supernatant obtained from (a) to 10 ml, with shaking;   c) filtering the diluted supernatant obtained in (b) with a 0.45 pm PTFE membrane filter;   d) subjecting the filtered supernatant obtained in (c) to high performance liquid chromatography analysis; and   e) obtaining a high performance liquid chromatography profile which contains the marker compound.   
     
     
         2 . The process according to  claim 1 , wherein the dried  Labisia pumila  plant material in (a) is prepared by drying at a temperature of 40° C. for 3 days. 
     
     
         3 . The process according to  claim 1 , wherein the extracting in (a) is carried out at 80° C. for 3 hours and with continuous stirring. 
     
     
         4 . The process according to  claim 3 , wherein the extracting is repeated and the ratio of  Labisia pumila  plant material:water is 1:6. 
     
     
         5 . The process according to  claim 1 , wherein the  Labisia pumila  extract in (a) is dried and concentrated by spray-drying. 
     
     
         6 . The process according to  claim 5 , wherein the spray-drying is performed using a spray tower having a tower inlet and outlet, and wherein tower inlet temperature of the tower inlet and outlet are is 185° C. and wherein tower outlet temperature is 107° C., respectively. 
     
     
         7 . The process according to  claim 1 , a consistent yield of water-soluble extract powder of 4-5% is obtained. 
     
     
         8 . The process according to  claim 1 , wherein 50-60 mg dried extract in 10 ml water is centrifuged at 20,000 rpm for 15 mins. at room temperature. 
     
     
         9 . The process according to  claim 1 , wherein the marker compound obtained is 3,4,5-trihydroxybenzoic acid. 
     
     
         10 . The process according to  claim 9 , wherein the marker compound 3,4,5-trihydroxybenzoic acid appears as a single peak absorbing at 254 nm between 15 to 17 minutes in the high performance liquid chromatography or chromatogram. 
     
     
         11 . The process according to  claim 9 , wherein the standardized extract contains 35 to 45 μg/ml of the marker compound 3,4,5-trihydroxybenzoic acid. 
     
     
         12 . A process for isolating a marker compound from  Labisia pumila  extract comprising
 subjecting the extract to semi-preparative or preparative reverse-phase high performance liquid chromatography,   wherein the extract is obtained by extracting dried  Labisia pumila  plant material with water in a ratio of 1:6 of dried  Labisia pumila  plant material:water to form a water-soluble extract and drying the extract obtained.   
     
     
         13 . The process according to  claim 12 , wherein the dried  Labisia pumila  plant material is prepared by drying at a temperature of 40° C. for 3 days. 
     
     
         14 . The process according to  claim 12 , wherein the extracting is carried out at 80° C. for 3 hours and with continuous stirring. 
     
     
         15 . The process according to  claim 14 , wherein the extracting is repeated and the ratio of  Labisia pumila  plant material:water is 1:6. 
     
     
         16 . The process according to  claim 12 , wherein the  Labisia pumila  extract is dried and concentrated by spray-drying. 
     
     
         17 . The process according to  claim 16 , wherein the spray-drying is performed using a spray tower having a tower inlet and outlet, and wherein tower inlet temperature of the tower inlet and outlet are is 185° C. and wherein tower outlet temperature is 107° C., respectively. 
     
     
         18 . The process according to  claim 12 , wherein a consistent yield of water-soluble extract powder of 4-5% is obtained. 
     
     
         19 . The process according to  claim 12 , wherein reverse-phase high performance liquid chromatography is run at ambient temperature at 254 nm wavelength using 99% (0.1% H 3 PO 4 ): 1% (CH 3 Cl) as a solvent in an isocratic system. 
     
     
         20 . The process according to  claim 12 , wherein the marker compound obtained is 3,4,5-trihydroxybenzoic acid. 
     
     
         21 . The process according to  claim 20 , wherein the marker compound 3,4,5-trihydroxybenzoic acid appears as a single peak absorbing at 254 nm between 15 to 17 minutes in the high performance liquid chromatography or chromatogram. 
     
     
         22 . The process according to  claim 20 , wherein the standardized extract contains 35 to 45 μg/ml of the marker compound 3,4,5-trihydroxybenzoic acid. 
     
     
         23 . A process for elucidating the structure of a marker compound from  Labisia pumila  extract comprising
 subjecting the isolated marker compound 3,4,5-trihydroxybenzoic acid to any one of the following methods: nuclear magnetic resonance spectrometry, liquid chromatography mass spectrometry and ultraviolet spectrophotometry,   wherein the marker compound 3,4,5-trihydroxybenzoic acid is isolated from the extract obtained by extracting dried  Labisia pumila  plant material with water in a ratio of 1:6 of dried  Labisia pumila  plant material:water to form a water-soluble extract and drying the extract obtained.   
     
     
         24 . The process according to  claim 23 , wherein the dried  Labisia pumila  plant material is prepared by drying at a temperature of 40° C. for 3 days. 
     
     
         25 . The process according to  claim 23 , wherein the extracting is carried out at 80° C. for 3 hours and with continuous stirring. 
     
     
         26 . The process according to  claim 25 , wherein the extracting is repeated and the ratio of  Labisia pumila  plant material:water is 1:6. 
     
     
         27 . The process according to  claim 23 , wherein the  Labisia pumila  extract is dried and concentrated by spray-drying. 
     
     
         28 . The process according to  claim 27 , wherein the spray-drying is performed using a spray tower having a tower inlet and outlet, and wherein tower inlet temperature of the tower inlet and outlet are is 185° C. and wherein tower outlet temperature is 107° C., respectively. 
     
     
         29 . The process according to  claim 23 , wherein a consistent yield of water-soluble extract powder of 4-5% is obtained. 
     
     
         30 . The process according to  claim 23 , wherein nuclear magnetic resonance spectrometry is run using deuterated dimethyl sulfoxide (DMSO-d 6 ) as solvent. 
     
     
         31 . The process according to  claim 23 , wherein the marker compound obtained is 3,4,5-trihydroxybenzoic acid. 
     
     
         32 . The process according to  claim 31 , wherein the marker compound 3,4,5-trihydroxybenzoic acid appears as a single peak absorbing at 254 nm between 15 to 17 minutes in the high performance liquid chromatography or chromatogram. 
     
     
         33 . The process according to  claim 31 , wherein the standardized extract contains 35 to 45 μg/ml of the marker compound 3,4,5-trihydroxybenzoic acid. 
     
     
         34 . An optimized and reproducible chemical profile of  Labisia pumila  extract according to  claim 1  wherein the marker compound obtained is 3,4,5-trihydroxybenzoic acid.