Corn Products International Standard Methods of Analysis Method A76Date: Supersedes: Page: 10/04 4/98 1 of 3 Title: ANISIDINE VALUE - (IN OIL) 1 PRINCIPLE Spectrophotometric determination of aldehydes (principally 2-alkenals and 2,4-dienals) in vegetable fats and oils by reaction in an acteic solution of the aldehydic compounds in an oil and the p-anisidine (Note 6.1). The p-anisidine value is defined as 100 times the optical density measured at 350 nm in a 1 cm cell of a solution containing 1.00 g of the oil in 100 mL of a mixture of solvent and reagent according to the method. 2 SCOPE This method applies to all crude and refined oils (Note 6.2). 3 APPARATUS 3.1 Spectrophotometer: Single or double beam digital instrument, capable of operating in the visible wavelength region (350-720 nm), and equipped with 1 X 1 cm cells; such as Shimadzu Model UV-1201 or Perkin Elmer Lambda 3B or equivalent. Follow instrument instruction or guide to accomodate different size cells. 1.00(±0.01) cm, paired, glass cells. 3.2 4 REAGENTS 4.1 Acetic Acid, Glacial 99.7% acetic acid (HC2H3O2, sp g 1.05). (Note 6.3). 4.2 Iso-octane (2,2,4-trimethyl pentane) optically clear. (Note 6.4) Iso-octane is flammable and a fire risk. Explosive limits in air are 1.1-6.0%. Be sure to use an operating fume hood when working with this solvent.. 4.3 p-Anisidine (Note 6.5) p-Anisidine is an irritant handled with care in a fume hood. It is an aromatic amine, a class of toxic and possibly carcinogenic chemicals. TLV is 0.1 ppm. (p-Anisidine, continued) Analytical reagent grade 0.25 g/100 mL solution in glacial acetic acid. Authorization Prepared by: ______________________________________________________Job Title Laboratory Chemist Approved by: _____________________________________________________Job Title Analytical Services Director and any variation in test procedure may cause variations in results.Ab) Sample Weight.2 . p-anisidine reacts with aldehydic compounds in oils or fats. g 5. By means of an auto-pipette.2 As . The intensity of color of the yellowish reaction products formed depends on both quantity and structure of these compounds.1 In the presence of acetic acid.001 g. 2-alkenals and dienals will contribute substantially.1 5.(IN OIL) 5 PROCEDURE 5.Corn Products International Standard Methods of Analysis Method A76 Date: Supersedes: Page: 10/04 4/98 2 of 3 Title: ANISIDINE VALUE . 6. Determine the p-anisidine value (p-AV) by: 1) Calculation Anisidine Value (AV) = 25 (1. 6 NOTE 6.5 5. Pipette exactly 5 mL of the fat solution into one 10-mL test tube with ground glass stopper or teflon lined screw cap and exactly 5 mL of solvent into another. Ab = Absorbance of the solution of the oil or fat. using the reference cell filled with solvent as a blank.2 The sample should be perfectly clear and dry. Measure the absorbance (Ab) of the solution at 350 nm in a cell with the spectrophotometer. add exactly 1 mL of the p-anisidine reagent to each tube and mix. Dissolve and dilute to volume with iso-octane.4 5. After about 10 minutes measure the absorbance (As) of the solvent in the first test tube in the spectrophometer cell at 350 nm. using the solution from the second test tube as a blank in the reference cell. It has been found that a double bond in the carbon chain conjugated with the carbonyl double bond increases the molar absorbance four to five times.6 Where: As = Absorbance of the fat solution after reaction with the panisidine reagent. This method is highly empirical.5-4. Weigh 0. into a 25 mL volumetric flask.3 5.0 g of the sample to the nearest 0. edited by Dr. Bestfoods Analytical Methods. measured in a 1 cm cell between 300 and 380 nm.1 American Oil Chemists' Society: Official Methods And Recommended Practices Of The American Oil Chemists' Society. Method Cd 18-90. must be nearly nil. Champaign.1%. IL.4 6. 7.2 . In most cases n-hexane can be substituted for iso-octane as solvent. Crystals should be stored at 0-4°C in a dark bottle and with care should last about 1 year. 1998. Commercial product can be freed ffrom absorptive material by percolating it through a 100 cm long glass column filled with silica gel. Fifth Edition. the acetic acid must not be used for this analyses and the label changed to show the moisture level. BFM Oa-10 (1991). it is essential that the highly hygroscopic acetic acid is checked for moisture. oils containing high levels of oxidized fatty acids will not dissolve completely in nhexane. Solutions must be freshly made as needed (daily). Reagent solutions having absorbance greater than 0.(IN OIL) 6.5 7 REFERENCE 7. 6. Firestone. However. D.200 when measured in the 1 cm cell at 350 nm against iso-octane or n-hexane as a blank should be discarded.3 Since the reaction is dependant on the formation of water. p-Anisidine crystals and solutions will discolor with time. The absorbance of the solvent used.Corn Products International Standard Methods of Analysis Method A76 Date: Supersedes: Page: 10/04 4/98 3 of 3 Title: ANISIDINE VALUE . If the content exceeds 0.