A: The powdered mucilage stains red with ruthenium red TSand lead acetate TS.

B: It meets the requirements of the test for Swell volume.

Internal standard solution— Transfer 5.0mLof n-propyl alcohol into a 500-mLvolumetric flask containing approximately 450mLof water.Dilute with water to volume,insert the stopper into the flask,and mix well.

Standard stock solution— Transfer 5.0mLof absolute alcohol at 20±2into a 500-mLvolumetric flask containing approximately 450mLof water.Dilute with water to volume,insert the stopper into the flask,and mix well.

Standard solution— Transfer 10.0mLof the Standard stock solutionand 10.0mLof Internal standard solutioninto a 100-mLvolumetric flask.Dilute with water to volume,insert the stopper into the flask,and mix well.

Test solution— Transfer 0.5g of Psyllium Hemicellulose,accurately weighed,into a 150-mLconical flask.Add about 90mLof water,insert the stopper into the flask,and stir rapidly for 3hours using a magnetic stirrer.Add 10.0mLof the Internal standard solution,and mix well.Pass the sample through a filter having a 0.45-µm porosity.

Chromatographic system (see Chromatography á621ñ)— The gas chromatograph is equipped with a flame-ionization detector and a 0.53-mm ×30-m fused silica analytical column coated with 3.0-µm G43stationary phase.A0.53-mm ×2-m fused silica guard column may be used.The chromatograph is programmed as follows.Initially,the column temperature is equilibrated at 40for 5minutes.The temperature is then increased at a rate of 10per minute to 230,and is maintained at 230for 3minutes.The injection port temperature is maintained at 250,and the detector is maintained at 300.The carrier gas is helium.The split flow ratio is about 10:1,and the flow rate is maintained at about 4.0mLper minute.Inject the Standard solution,and record the peak responses as directed for Procedure:the relative standard deviation for replicate injections is not more than 2%.

Procedure— Separately inject equal volumes (about 0.5µL)of the Standard solutionand the Test solution into the chromatograph,record the chromatograms,and measure the responses for all the peaks.Calculate the percentage of alcohol in the portion of Psyllium Hemicellulose taken by the formula: in which Cis the concentration,in mg per mL,of alcohol in the Standard stock solution;Wis the weight,in mg,of Psyllium Hemicellulose taken;and RUand RSare the ratios of the peak responses of alcohol to those of n-propyl alcohol from the Test solutionand the Standard solution,respectively:not more than 12.0%(w/w)is found.

Alcohol solution— Transfer 82.0mLof alcohol to a 100-mLvolumetric flask,dilute with water to volume,and mix.

Buffer solution— Dissolve 1.95g of 2-(N-morpholino)-ethanesulfonic acid and 1.22g of tris(hydroxymethyl)aminomethane in 170mLof water.Adjust with 6Nsodium hydroxide to a pHof 8.2,dilute with water to 200mL,and mix.[NOTE—It is important to adjust the pHto 8.2at 24.If the Buffer solutiontemperature is 20,adjust the pHto 8.3;if the temperature is 28,adjust the pHto 8.1.For deviations between 20and 28,adjust by interpolation.]

Acid solution— Prepare 0.561Nhydrochloric acid by dissolving 9.35mLof 6Nhydrochloric acid in 70mLof water.Dilute with water to 100.0mL,and mix.

Phosphate buffer— Prepare a pH6.0phosphate buffer (see Buffer Solutionsunder Reagents,Indicators,and Solutions).

Protease solution— Dissolve 5mg of protease in 0.1mLof Phosphate buffer.

Enzyme purity— To ensure the absence of undesirable enzymatic activities and the presence of desirable enzymatic activities,proceed as directed for Test preparations and Procedure using the substrates listed in the following table in place of Psyllium Hemicellulose. The enzyme preparation is suitable if more than 90%of the original weight of pectin,arabinogalactan,and b-glucan is recovered;not more than 2%of the original weight of casein and corn starch is recovered;and not more than 1%of the original weight of wheat starch is recovered.[NOTE—Test the enzyme purity of every new lot of enzyme and at 6-month intervals thereafter.]

Blank preparations— Using two 400-mLtall-form beakers,appropriately labeled,proceed as directed for Procedurewithout Psyllium Hemicellulose.

Test preparations— Weigh accurately,in duplicate,approximately 0.2g of Psyllium Hemicellulose,previously milled to very fine powder.[NOTE—Duplicates should differ by less than 1mg in weight.]Transfer duplicate samples to appropriately labeled 400-mL,tall-form beakers,and proceed as directed for Procedure.

Procedure— Treat each preparation in the following manner.Add 40mLof Buffer solutionto the beaker.[NOTE—For the Test preparation,stir until Psyllium Hemicellulose is completely dispersed.]Add 125µLof heat-stable a-amylase solution,and stir to ensure uniform mixing.Cover the beaker with aluminum foil,and incubate over a water bath maintained at 95to 100for 15minutes,with continuous agitation.[NOTE—Start timing once the water bath temperature reaches 95;a total time of 35minutes is usually sufficient.]Remove the beaker from the water bath,and cool to 60.Remove the aluminum foil,scrape any ring from inside the beaker,and disperse any gels in the bottom of the beaker with a spatula.Rinse the walls of the beaker and the spatula with 10mLof water,collecting the rinsings in the beaker.Add 500µLof Protease solution.Cover with aluminum foil,and incubate over a water bath maintained at 60±3for 30minutes with continuous agitation.[NOTE—Start timing when the bath temperature reaches 60.]Remove the foil,and transfer 5mLof Acid solution while stirring.Adjust,if necessary,with 1Nsodium hydroxide or 1Nhydrochloric acid to a pHof 4.28±0.07at 60.[NOTE—It is important to adjust the pHto 4.28while the solution in the beaker is maintained at 60,otherwise the pHwill increase at lower temperatures.]Add 150µLof amyloglucosidase solution while stirring.Cover with aluminum foil,and incubate over a water bath maintained at 60±3for 30minutes with constant agitation.[NOTE—Start timing once the water bath reaches 60.]Transfer approximately 40mLof the beaker contents to a 50-mLcentrifuge tube,and sonicate the tube contents for 3minutes.*Centrifuge at 10,000–14,000rpm for 10minutes.Carefully pour the supernatant into an appropriately labeled 600-mLtared beaker.Do not disturb any pellet in the bottom of the centrifuge tube.Add the remaining sample from the original 400-mLbeaker into the centrifuge tube still containing the pellet.Rinse the 400-mLbeaker with 15–20mLof water,and add the rinsing to the 50-mLcentrifuge tube.Centrifuge the sample at 10,000–14,000rpm for 10minutes.Carefully pour the supernatant into the 600-mLbeaker containing the first supernatant.Add 390mL(measured before heating)of alcohol at 60to the 600-mLbeaker.Cover the beaker,and allow to stand for at least 1hour to form a precipitate.