SPDS Aloe Vera Method Book

Date Issued: Supersedes:

Page:

4 of 7

Procedure Title: Determination of Organic Acid Contents in Aloe vera Fresh Leaf and Concentrated Juice Powder by HPLC Method

N/A

Effective Date:

QCL

SOP #:

Revision #:

Prepared By: Yuehong Zhang, Zhichao Bao Department: R&D Applies To: Herbalife International

1.2.1.6.6 Standard Level 6: Pipette 0.5 mL of the Mixed Organic Acid Standard Stock Solution into a 500 mL volumetric flask and dilute to volume with ultrapure water. Mix well.

1.3

Sample Preparation 1.3.1

Raw Material (Fresh Leaf and Concentrated Juice Powder) 1.3.1.1 Preparation of fresh leaf samples: for whole leaf sample, cleaned by water and cut the leaf tip, butt, barbs and edges with a knife. Grind the entire aloe leaves in a high speed blender until the blend was flowing smoothly and no large chunks remained. Squeeze the juice out of the leaf residue and lyophilize the juice to concentrated whole leaf juice powder. For inner leaf, filet the cleaned leaf with knife to separate the rind from the inner leaf and lyophilize inner leaf to concentrated inner leaf juice powder. 1.3.1.2 Weigh ~250 mg of the concentrated juice powder sample into a 50 mL volumetric flask. Dissolve and dilute to volume with ultrapure water. Vortex the sample for 60 seconds and sonicate for 10 minutes. Filter sample liquid through a 0.45μm Hydrophilic PTFE filter into a glass HPLC vial. Finished Product (powder and liquid form) 1.3.2.1 Weigh ~250 mg of the powder raw material sample into a 50mL volumetric flask, dissolve and dilute to volume with ultrapure water, vortex the sample for 60 seconds, and sonicate for 10 minutes. For liquid raw material, weigh ~25 g of the raw material sample into a 50mL volumetric flask; Dissolve and dilute to volume with ultrapure water. Vortex the sample for 60 seconds, and sonicate for 10 minutes. Filter sample liquid through a 0.45μm Hydrophilic PTFE filter into a glass HPLC vial. Chromatograph the three calibration standards. The Correlation Coefficient (R) for the oxalic acid, tartaric acid, formic acid, malic acid, isocitric acid, lactic acid, acetic acid, citric acid, succinic acid, fumaric acid and isocitric acid lactone curve(s) should be >0.998. Run a standard check after every six sample injections and at the end of the run. Retention time deviation of the standards should be no more than 0.5 minute, and the Relative Standard Deviation (RSD) for standard check injections and calibration standard should be no more than 8.0%. The theoretical plates of the organic acids and isocitric acid lactone peaks should be >10000. Calculations Obtain organic acid standards calibration curve(s) by plotting standards concentrations (μg/mL) versus standards peak areas. Interpolate the sample concentration (μg/mL) from the standard curve. Calculate the amounts of isocitric acid, isocitric acid lactone and other organic acid per the formulas below: 1.5.1 Correction of Isocitric Acid Lactone Standard Concentration: Isocitric acid lactone is partially hydrolyzed into hydrolyzate isocitric acid in aqueous solution. The actual System Suitability 1.4.1 1.4.2 1.4.3 1.4.4 1.4.5 The tailing factor for all three peaks should be <2.0.

1.3.2

1.4

1.5

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