Rhodiola rosea.Rosavin.Salidroside.Narrative History,Function,Uses and Application of Rhodiola rosea.Rhodiola Rosea Extract.

Article Content:

HPLC analysis of rosavins and salidroside.

The content of rosavins and salidroside were determined using a Waters System HPLC equipped with 996 Photodiode Array Detector, two model 515 Pumps, a Gradient Mixer Kit 051518, a Pump Control Module, a Bus SAT/IN Module, a model 7725I Injector with 20 ml loop, and a Millenium32 Chromatography Manager (Version 3.0). For all separations a RP-C18 analytical column C-18, 3.9 x 150 mm, 5 mm particle size was used (Symmetry, WATO27324, Waters Associates, Inc.). Two mobile solvents were used to develop a binary gradient, phase A: 0.16 M ammonium acetate solution in water (w/v) (pH is adjusted to 5.5 with acetic acid) and, phase B: methanol. The mobile phase flow rate was adjusted to 1.0 ml/min, and UV detection wavelength was set at 2 different wavelengths, 254 for rosavin, rosin and rosarin, and 280 nm for salidroside (Dubichev et al. 1991; Kurkin et al. 1986a; Ramazanov and Bernal Suarez, 1999). After 5 min holding the initial solvent mix, 66:34 (A: B, v/v), a linear ramp up to 60:40 (A: B, v/v) over 17 min was developed, followed by a return to the initial conditions over 5 min, and 5 min equilibration before the next analysis.

The HPLC reference standards of rosavin, rosin, rosarin and salidroside. Stock standard solutions were prepared in ethanol: water (85:15, v/v) to a concentration of 1 mg/ml. Four standard solutions containing both components in different concentrations, between 0.01 and 0.3 mg/ml, were injected. The calibration curve for each standard was linear in the described range with correlation coefficients of 0.99.

Qualitative Reactions of Rhodiola rosea:

Into a 20ml flask is put 1 gram of mill stock (see Section Quantitative Determination), then add 10ml of methyl alcohol and heat on water bath at 65 for 20 minutes with a reflux condenser. Extract is filtered through a paper filter. On the starting line of Silufol* plate UV-254 with a micropipette is applied 0.002ml of resultant filtrate. The plate with the applied sample is placed into a chamber which is previously saturated with solvents mixture (chloroform-methyl alcohol-water (26:14:3)) for not less than 24 hours and chromatograph by the ascending method.

When the solvent front passes about 13cm, the plate is taken out of the chamber, dries in air for 5 minutes and looked through UV light at 254nm wavelength. On the chromatogram must appear a dominating spot of violet color with R1 constituting about 0.4 (rosavine); it is possible to have available other spots.

The chromatogram is sprinkled with a 10% solution of sodium carbonate ...... at 110 for 2 minutes, then sprinkle with diazotized sulfacyl and heat at 110 for 2 minutes. On the chromatogram must appear a spot of rosy color with R1 constituting about 0.42 (salidrozide); it is possible to have available other spots.

Quantitative Determination:

A stock analytical sample up to the size of particles passing through a 2mm sieve. About 0.5 gram (precise weight) of mill stock is put into a 100ml flask, then add 10ml of water and heat on boiling water bath with a reflux condenser for 15 minutes.

Then the extract is filtered through a paper filter into a 50ml measuring flask avoiding getting stock particles on the filter. Extraction is repeated again 3 times by 10ml water, heating each time for 10 minutes and filtering into the same measuring flask.

To the cooled filtrate is added 6ml of 10% solution of lead acetate, 2ml of saturated solution of sodium sulfate, then thoroughly mix, bringing the volume of the solution with water up to the mark and filter through a paper filter. The first 15ml of filtrate is discarded.

Into a 25ml measuring flask is transferred 5ml of resultant filtrate, add 2.5ml of 2% solution of sodium carbonate, 2.5ml of diazotized sulfanyl, bringing the volume of the solution with water up to the mark, mix and in 5 minutes measure the optical density on a spectrophotometer at 486nm wavelength in a cuvette with the layer 10mm thick, using water as a reference solution.

The content of salidrozide in terms of absolutely dry stock in percentage terms (X) should be computed from the formula:

? 250 100
X =--------------------------,
253 m(100 - W)

where:
D:Optical density of solution under analysis;
253:Specific index of absorption (E...) of salidrozide;
m:Stock mass in grams;
W:Mass loss during stock drying in percentage terms.

Notes:1. Preparation of diazotized sulfanyl:7 grams of sodium sulfanyl is dissolved in 50ml of water in a 100ml measuring flask, add 9ml of concentrated hydrocarbon acid and bring the volume of the solution with water to the mark, then 1ml of resultant solution is put into a 100ml measuring flask, put it on ice, add 50ml of water, 0.2ml of 10% solution of sodium nitrite, mix and bring the volume of the solution with water to the mark. The solution should be used fresh.
Notes:2. Preparation of saturated solution of sodium sulfate:60 grams of sodium sulfate is filled with 100ml of water and leave, while frequently shaking, for 24 hours.

Reference:

1.Rhodiola rosea.Rosavin.Salidroside.Narrative History,Function,Uses and Application of Rhodiola rosea.Rhodiola Rosea Extract.

Rhodiola rosea.Rosavin.Salidroside.Narrative History,Function,Uses and Application of Rhodiola rosea.Rhodiola Rosea Extract.

Article Content:

HPLC analysis of rosavins and salidroside.

Reference:

last edit date:6th,May.2009.

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