Charantin content

REAGENTS & INSTRUMENT REQUIRED:

1. High Performance Liquid Chromatography
2. Water HPLC grade
3. Methanol LR grade
4. Charantin working standard.

PROCEDURE:

Standard Preparation :

Weigh accurately about 2.5mg of working standard in 10mL volumetric flask. Add about 5mL of Tetrahydrofuran, ultrasonicate till completely dissolved and make up the volume with Tetrahydrofuran.

Shake well and filter through 0.45µm filter paper.

Sample Preparation :

Weigh accurately about 100mg of sample of extract in 50mL volumetric flask. Add about 25mL of Tetrahydrofuran, ultrasonicate for five minutes and make up the volume with Tetrahydrofuran. Ensure that the concentration of active ingredient in sample solution is approximately similar to that of standard solution and Shake well and filter through 0.45µm filter paper.

Chromatographic Condition:

Mobile Phase               - Water (A) : Methanol (B) 

Conditions                    - 85% B increasing to 95% B over 15 minutes, then increasing to 100% B for                                                                                            

                                       5minutes.                                           

Column                        - Phenomenex luna 5µ, C18 (250mm X 4.6mm)

Mode                           – Gradient

Injection Volume         – 20µL

Flow rate                     – 1.0mL /minute

Detector wavelength   – 250 ± 50nm

Condition                    -  RT

Calculation:

                                             Area of sample  X  Concentration of std    X   purity of std

  % of  Charantin content =---------------------------------------------------------------------------------

                                                Area of standard    X   Concentration of sample