Preparation of thiabendazole standards for use in applications of the benomyl/carbendazim rapid assay

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GENERAL COMMENTS:
This procedure describes the preparation of thiabendazole (TBZ) standards used in applications of the Benomyl/Carbendazim RaPID Assay. Standard concentrations of 10, 50 and 250 ng/mL (ppb) TBZ and a control concentration of 100 ppb TBZ are prepared by making appropriate dilutions of a concentrated TBZ solution in the Benomyl/Carbendazim Sample Diluent. The standards and control can be used up to 12 months after preparation when stored at refrigerated temperatures (2 - 8°C).

MATERIALS:
Reagents: Thiabendazole (2-(4-thiazolyl) benimidazole) PestanalÒ, Crescent. Chemical Co., Inc., Hauppauge, NY 11788. N,N-Dimethylformamide (DMF), HPLC grade. Five bottles of Benomyl/Carbendazim Sample Diluent (SDI part no. A00095, 100 mL each).
Equipment: Analytical balance capable of accurately measuring milligrams (± 1 mg), appropriate size volumetric flasks (10 and 100 mL) and graduated cylinders, appropriate size volumetric or precision pipets for measuring 0.1, 0.5, 1 and 2.5 mL, graduated cylinders.

PREPARATION OF TBZ STOCKS:
Stock I (1 mg/mL): Weigh 10 mg of thiabendazole (using an analytical balance) into a 10 mL volumetric flask. Fill flask to volume with DMF. Stopper the flask and invert repeatedly (minimum 20 to 30 times) until dissolved. Label flask as 'Stock I' and store refrigerated for up to 1 month.
Stock II (10 mg/mL): Measure approximately 90 mL of Sample Diluent into graduated cylinder. Add this amount of Sample Diluent to a clean 100 mL volumetric flask. Transfer 1 mL of Stock I solution to the flask using a 1 mL volumetric or precision pipet. Fill flask to final volume with Sample Diluent. Stopper flask, swirl and invert several times to ensure thorough mixing of solution. Label flask as 'Stock II' and store refrigerated for up to 1 month.

PREPARATION OF STANDARD SOLUTIONS: Prepare the following standard solutions:
Standard 1 (0.01 mg/mL): Measure approximately 90 mL of Sample Diluent into graduated cylinder. Add this amount of Sample Diluent to a clean 100 mL volumetric flask. Transfer 0.1 mL of Stock II solution to the flask using a 0.1 mL volumetric or precision pipet. Fill flask to final volume with Sample Diluent. Stopper flask, swirl and invert several times to ensure thorough mixing of solution. Label flask as 'TBZ Standard 1, 10 ppb” and store refrigerated.
Standard 2 (0.05 mg/mL): Measure approximately 90 mL of Sample Diluent into graduated cylinder. Add this amount of Sample Diluent to a clean 100 mL volumetric flask. Transfer 0.5 mL of Stock II solution to the flask using a 0.5 mL volumetric or precision pipet. Fill flask to final volume with Sample Diluent. Stopper flask, swirl and invert several times to ensure thorough mixing of solution. Label flask as 'TBZ Standard 2, 50 ppb' and store refrigerated.
Standard 3 (0.25 mg/mL): Measure approximately 90 mL of Sample Diluent into graduated cylinder. Add this amount of Sample Diluent to a clean 100 mL volumetric flask. Transfer 2.5 mL of Stock II solution to the flask using a 2.5 mL volumetric or precision pipet. Fill flask to final volume with Sample Diluent. Stopper flask, swirl and invert several times to ensure thorough mixing of solution. Label flask as 'TBZ Standard 3, 250 ppb' and store refrigerated
Control (0.10 mg/mL): Measure approximately 90 mL of Sample Diluent into graduated cylinder. Add this amount of diluent to a clean 100 mL volumetric flask. Transfer 1.0 mL of Stock II solution to the flask using a 1.0 mL volumetric or precision pipet. Fill flask to final volume with diluent. Stopper flask, swirl and invert several times to ensure thorough mixing of solution. Label flask as 'TBZ Control, 100 ppb' and store refrigerated.
Zero Standard: Label remaining Sample Diluent as “TBZ Zero Standard”.

PROCEDURAL NOTES:
Benomyl and carbendazim are approximately fifty times more reactive in the Benomyl/Carbendazim RaPID Assay than TBZ. Therefore, a TBZ application is not recommended if residues of benomyl and carbendazim are expected to be present. Accuracy in weighing small amounts (milligrams) of thiabendazole and accuracy in making dilutions of Stock Solutions is critical for preparing standard solutions with accurate concentrations of thiabendazole. Note: A TBZ procedure must be installed on the RPA-1 analyzer so the TBZ concentration units are applied correctly. Installation of the TBZ procedure on the RPA-1 can be obtained from SDI Technical Service at the number shown below.

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