5. AOACSPDSMethods-2018AwardsV3

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B rown & Y u : J ournal of AOAC I nternational V ol . 96, N o . 1, 2013  17

Table 7. Reproducibility and HorRat values of the ginseng test materials

Test materials

Reproducibility

No. of laboratories

Mean total ginsenoside, mg/g

Repeatability RSD r , %

Matrix

GIN-GCS

RSD R

, % HorRat ( r,R )

P.q. powdered whole root P.q. powdered extract

1, 2 3, 4

12 12

45.5

1.03 1.16

4.80 4.39

1.5 1.6

100.8

P.q. powdered whole root tablet

5

12

16.5

12.80

–

3.5

P.g. powdered whole root

6, 7

12

60.0

2.22

4.72

1.6

P.g. powdered extract

8, 9

12

96.1

1.66

5.29

1.9

P.g. powdered whole root capsule

10

12

54.6

8.95

–

2.9

P.g. spiked matrix blank

11, 12

12

38.8

2.00

5.93

1.8

( 4 ) Record the average capsule fill weight. ( 5 ) Transfer the contents into a 50 mL conical tube and mix using a spatula to homogenize the samples. ( 6 ) Accurately weigh the sample into a 50 mL conical tube. Use the following weight (±0.1 mg) and code for this ginseng matrix: GC No. for P. ginseng/P. quinquefolius capsule (400 mg). ( 7 ) Follow the same procedure as indicated for powdered raw materials and finished products in ( a ) from steps ( 2 ) to ( 8 ), then ( 11 ). The basic hydrolysis step is not necessary for this sample type. ( c )  Tablets and caplets.— ( 1 ) Determine the average tablet weight by weighing out 20 tablets or caplets. The average weight is calculated using the following equation: Average tablet or caplet weight, g = (total weight)/20 ( 2 ) Grind the materials to make a homogeneous fine powder using a mortar and pestle, and then transfer the contents into a 50 mL conical tube. Mix using a spatula to homogenize the samples. ( 3 ) Accurately weigh the sample into a 50 mL conical tube. Use the following weight (±0.1 mg) and code for this sample type: GT No. for P. ginseng / P. quinquefolius tablets (1000 mg). ( 4 ) Follow the same procedure as indicated for powdered raw materials and finished products in Section ( a ) from steps ( 2 ) to ( 8 ), then ( 11 ). Note that basic hydrolysis is not necessary for this sample type. ( a )  System suitability test .—Equilibrate the HPLC system with the mobile phase for at least 5 min until a stable baseline is obtained. Inject 10 µL mixed standard solution eight times (e.g., Linearity 7, lowest concentration). An RSD r of ≤5% for peak area, an R s ≥ 1.5, and a T ≤ 2 are desirable for the system to be suitable for the analysis. ( b )  Calibration .—Make single 10 µL injections of calibration solutions from Linearity 7 through Linearity 1 (omitting Linearity 4a). Calculate the slope, y-intercept, and r 2 value for the calibration curves for Rg 1 , Re, Rb 1 , Rc, Rb 2 , and Rd. The r 2 value should be ≥0.995 for each analyte. Record these values along with peak areas on the appropriate raw data form. Expected retention times for each ginsenoside are listed in Table 4. Determination

the following weights (±0.1 mg) and codes for each individual ginseng matrix: GR No. for P. ginseng/P. quinquefolius whole root samples (400mg) andGENo. for P. ginseng/P. quinquefolius powdered extracts (200 mg). ( 2 ) Add 10 mL extraction solvent. Mix the sample tube using a vortex mixer for at least 1 min to ensure optimal mixing, and then sonicate for 25 min at room temperature. ( 3 ) Centrifuge the sample tubes at 3500 rpm for 5 min. ( 4 ) Decant the solvent into a prelabeled 25 mL volumetric flask and identically label the flask with its appropriate sample identification code. ( 5 ) Repeat steps ( 2 ) to ( 4 ) with a second 10 mL aliquot extraction solvent. ( 6 ) Repeat steps ( 2 ) to ( 4 ) a third time using 4 mL extraction solvent. Combine the extracts. ( 7 ) Rinse the extracted residue with approximately 1 mL extraction solvent and vortex mix for 5 s. Centrifuge at 3500 rpm for 5 min, decant into the same volumetric flask, and bring to a final volume of 25 mL using the extraction solvent. ( 8 ) Cap and invert the volumetric flask at least 10 times to ensure complete mixing of the extract solution. Note: For root samples, follow steps ( 9 ) to ( 11 ). For powdered extracts and finished products (GC and GT below) proceed directly to step ( 11 ). ( 9 ) Pipet 1 mL extract solution into a 2 mL microcentrifuge tube and add 100 µL base hydrolyzing agent (5% KOH, w/v). Mix the solution using a vortex mixer. Cover and allow the mixture to sit for 2 h at room temperature protected from direct light. ( 10 ) Neutralize the solution by adding 100 µL neutralizing agent (14% KH 2 PO 4 , w/v) solution. Mix the solution using a vortex mixer. ( 11 ) Transfer an aliquot of the solution into an HPLC vial using a syringe fitted with a 0.45 µm PTFE filter. ( b )  Hard-filled capsules.— ( 1 ) Determine the total capsule content weight (C) by weighing 20 capsules. Record the weight. ( 2 ) Cut open the 20 capsules with a suitable cutting instrument. ( 3 ) Empty and combine the capsules’ contents in a conical tube. Record the weight of the capsules’ contents, as well as the weight of the empty capsule shells (S). The average capsule fill weight is calculated using the following equation: Average capsule fill weight, g = (C – S)/20