Belladonna Extract
»Belladonna Extract contains,in each 100g,not less than 1.15g and not more than 1.35g of the alkaloids of belladonna leaf.
PILULAR BELLADONNA EXTRACT
Prepare the extract by percolating 1000g of Belladonna Leaf,using a mixture of 3volumes of alcohol and 1volume of water as the menstruum.Macerate the drug for 16hours,and then percolate it at a moderate rate.Evaporate the percolate under reduced pressure and at a temperature not exceeding 60to a pilular consistency,and adjust the remaining extract,after assaying,by dilution with liquid glucose so that the finished Extract will contain,in each 100g,1.25g of the alkaloids of belladonna leaf.
POWDERED BELLADONNA EXTRACT
Prepare the extract by percolating 1000g of Belladonna Leaf,using alcohol as the menstruum.Macerate the drug for 16hours,and then percolate it slowly.Evaporate the percolate under reduced pressure and at a temperature not exceeding 60to a soft extract,add 50g of dry starch,and continue the evaporation,at the same temperature,until the product is dry.Powder the residue.The extract may be deprived of its fat by treating either the soft extract first obtained,or the dry and powdered extract,as directed under Extracts(seePharmaceutical Dosage Forms á1151ñ).Assay the powdered residue,and add sufficient starch,previously dried at 100,to obtain a finished Extract containing 1.25g of the alkaloids of belladonna leaf in each 100g.Mix the powders,and pass the Extract through a fine sieve.
Packaging and storage— Preserve in tight containers,at a temperature not exceeding 30.
Organic volatile impurities,Method IVá467ñ: meets the requirements.
Assay—
pH9.5Phosphate buffer— Dissolve 34.8g of dibasic potassium phosphate in 900mLof water,and adjust to a pHof 9.5,determined electrometrically,by the addition of 3Nhydrochloric acid or sodium hydroxide,with mixing.
Internal standard solution— Dissolve about 40mg of USP Homatropine Hydrobromide RS,accurately weighed,in about 25mLof dilute sulfuric acid (1in 350)in a 50-mLvolumetric flask,add the same dilute acid to volume,and mix.Prepare fresh on the day of use.
Standard preparation— Dissolve about 10mg of USP Scopolamine Hydrobromide RS,accurately weighed,in about 5mLof dilute sulfuric acid (1in 350)in a 10-mLvolumetric flask,add the same dilute acid to volume,and mix (Solution A).Dissolve about 20mg of USP Atropine Sulfate RS,accurately weighed,in about 25mLof dilute sulfuric acid (1in 350)in a 50-mLvolumetric flask,add 2.0mLof Solution A,and mix.Add dilute sulfuric acid (1in 350)to volume,and mix.Prepare fresh on the day of use.
Extraction blank— Place about 10mLof dilute sulfuric acid (1in 350)in a 60-mLseparator.Proceed as directed under Assay preparation,beginning with “then add 15mLof chloroform.”The blank chromatogram contains no significant interferences at the locus of atropine,scopolamine,or homatropine.
Assay preparation— Weigh accurately about 0.5g of Extract,transfer to a 125-mLconical flask,and add 40mLof dilute sulfuric acid (1in 350).Heat to a temperature not above 45,and stir to hasten solution.Filter the solution through filter paper into a 100-mLvolumetric flask.Wash the flask and the filter with two 20-mLportions of warmed dilute sulfuric acid (1in 350),and collect the washings in the 100-mLvolumetric flask.Add dilute sulfuric acid (1in 350)to volume,and mix.
Pipet 10mLof this solution into a 60-mLseparator.To the separator add 1.0mLof Internal standard solution,then add 15mLof chloroform,shake vigorously,allow the layers to separate,and discard the chloroform layer.(If emulsions are formed,a mixed solventconsisting of chloroform and isopropyl alcohol (10:3)may be substituted for chloroform throughout the extraction procedure.)Add another 15mLof chloroform,and extract again,discarding the chloroform phase.Add 15mLof pH9.5Phosphate bufferand sufficient 1Nsodium hydroxide to yield a final pHbetween 9.0and 9.5.Add 15mLof chloroform,shake vigorously,and allow the layers to separate.Filter the organic phase through 10g of anhydrous sodium sulfate (see Suitability for alkaloid assaysunder Sodium Sulfate,Anhydrous,in the section Reagents,Indicators,and Solutions),previously washed with chloroform and supported in a funnel with a small pledget of glass wool,into a suitable container.Extract again with two 15-mLportions of chloroform,again collecting the clarified organic phase.Wash the sodium sulfate and the tip of the funnel with 5mLof chloroform.Evaporate the combined organic phases under reduced pressure,at a temperature below 45,add 1mLof chloroform,and mix to dissolve the alkaloids,taking care to wet the sides of the container.
Standard curve— Prepare three Standard solutionsas follows.Pipet into three separate 60-mLseparators 1.0-,2.0-,and 3.0-mLportions,respectively,of Standard preparation,and add 9.0,8.0,and 7.0mL,respectively,of dilute sulfuric acid (1in 350).Proceed as directed under Assay preparation,beginning with “add 1.0mLof Internal standard solution.”
Chromatographic system— Under typical condition,the instrument contains a 1.2-m ×4-mm glass column packed with 3%G3on S1AB.The column may be cured and conditioned as specified under Gas Chromatography(see Chromatography á621ñ).The column is maintained at a temperature of about 215,and the injection port and detector block at about 240,and dry helium is used as a carrier gas at a flow rate of about 65mLper minute.
System suitability— Chromatograph six to ten injections of the solution,and record peak areas as directed for Procedure.The analytical system is suitable for conducting this assay if the relative standard deviation for the ratio,RA,calculated by the formula:
100×(standard deviation /mean ratio),
does not exceed 2.0%;the resolution,R,between aHand aAis not less than 3;and the tailing factor (the sum of the distances from peak center to the leading edge and to the tailing edge divided by twice the distance from peak center to the leading edge),measured at 5%of the peak height of aA,does not exceed 2.0.
Procedure— Inject a portion (about 5µL)of each Standard solutioninto a suitable gas chromatograph equipped with a flame-ionization detector.Measure the areas,aA,aH,and aS,of the atropine,homatropine,and scopolamine peaks,respectively,in each chromatogram,and calculate the ratios AAand ASby the formulas:
aA/aHand aS/aH.
Plot the Standard curvesof the values of RAand RSagainst the amounts,in mg,of atropine and scopolamine in the solutions.(The ratio of the molecular weight of atropine to that of anhydrous atropine sulfate is 0.8551,and the ratio of the molecular weight of scopolamine to that of anhydrous scopolamine hydrobromide is 0.7894.)Inject a portion of the Assay preparationinto the chromatograph,obtain the chromatogram area ratios,measure the peak areas,and calculate the area ratios,as with the Standard solutions.Record from the Standard curvethe quantities,in mg,of atropine and scopolamine in the volume of specimen taken.Add the quantity,in mg,of atropine and scopolamine,and multiply by 10to obtain the weight,in mg,of alkaloids in the portion of Extract taken.
Auxiliary Information— Staff Liaison:Gabriel I.Giancaspro,Ph.D.,Senior Scientist and Latin American Specialist
Expert Committee:(DSB)Dietary Supplements:Botanicals
USP28–NF23Page 223
Phone Number:1-301-816-8343