6. AOACSPIFANMethods-2018Awards

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1666 Butler-Thompson et al. : J ournal of AOAC I nternational V ol. 98, N o. 6, 2015

INFANT FORMULA AND ADULT NUTRITIONALS

Determination of Myo-Inositol in Infant, Pediatric, and Adult Formulas by Liquid Chromatography-Pulsed Amperometric Detection with Column Switching: Collaborative Study, Final Action 2011.18 Linda D. Butler-Thompson, Wesley A. Jacobs, and Karen J. Schimpf 1 Abbott Nutrition, 3300 Stelzer Rd, Columbus, OH 43219 Collaborators: J. Austad; L. Basumallick; W.U. Bolong; L. Chen; S. Christiansen; C. Domer; D. Ellingson; G. Lautenschlager; I. Malaviole; S. Purachaka; G. Wang; F. Xong

Received April 30, 2015. Accepted by SG June 9, 2015 The method was approved by the AOAC Official Methods Board as Final Action. See “Standards News,” (2014) Inside Laboratory Management , November/December issue. The AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN) invites method users to provide feedback on the Final Action methods. Feedback from method users will help verify that the methods are fit for purpose and are critical to gaining global recognition and acceptance of the methods. Comments can be sent directly to the corresponding author. 1 Corresponding author’s e-mail: karen.schimpf@abbott.com DOI: 10.5740/jaoacint.15-107 Requirements (SMPRs ® ) for free myo-inositol plus phosphatidylinositol bound myo-inositol for all the matrixes analyzed. SMPRs for repeatability were ≤5% RSD at myo-inositol concentrations of 2–68 mg/100 g ready-to-feed (RTF) liquid. SMPRs for reproducibility were ≤8% RSD in products with myo-inositol AOAC First Action Method 2011.18, Myo-Inositol (Free and Bound as Phosphatidylinositol) in Infant and Pediatric Formulas and Adult Nutritionals, was collaboratively studied. With this method free myo-inositol and phosphatidylinositol bound myo-inositol are extracted using two different sample preparation procedures, separated by ion chromatography using a combination of Dionex Carbo Pac PA1 and MA1 columns with column switching, and detected with pulsed amperometry using a gold electrode. Free myo-inositol is extracted from samples with dilute hydrochloric acid and water. Phosphatidylinositol is extracted from samples with chloroform and separated from other fats with silica SPE cartridges. Myo-inositol is then released from the glycerol backbone with concentrated acetic and hydrochloric acids at 120°C. During this collaborative study, nine laboratories from five different countries analyzed blind duplicates of nine infant and pediatric nutritional formulas for both free and phosphatidylinositol bound myo-inositol, and one additional laboratory only completed the free myo-inositol analyses. The method demonstrated acceptable repeatability and reproducibility and met the AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN) Standard Method Performance

concentrations ranging from 2 to 68 mg/100 g RTF liquid. During this collaborative study, repeatability RSDs ranged from 0.51 to 3.22%, and RSDs ranged from 2.66 to 7.55% for free myo-inositol plus phosphatidylinositol bound myo-inositol. M yo-inositol is a water-soluble hydroxylated, cyclic 6-carbon quasi-vitamin. Myo-inositol is regarded as a quasi-vitamin because it appears to satisfy the criteria of vitamin status for only a few species or only under certain conditions. It is the only one of the nine possible stereoisomeric forms of cyclohexitol with reported biological activity and is present in nearly all living cells. Myo-inositol occurs in foods mainly as free myo-inositol, phytic acid, and inositol- containing phospholipids. Plants seeds are the richest sources of myo-inositol, but it is present predominantly as phytic acid which is a poor source of myo-inositol because most mammals have little or no intestinal phytase activity. Myo-inositol is present in animal products such as milk, eggs, and meat primarily as free myo-inositol and phosphatidylinositol (1). Phosphatidylinositol is also present in soy flour and lecithin. Although the predominant inositol phosphate in foods is myo- inositol hexakisphosphate or phytic acid (InsP 6 ), other inositol phosphates such as myo-inositol tris- (InsP 3 ), tetrakis- (InsP 4 ), and pentakis- (InsP 5 ) phosphate may be present in some foods in relatively minor amounts (2). The AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN) defined myo-inositol as free myo-inositol and phosphatidylinositol, excluding methyl ethers, glycosides, phosphorylated forms, and phytate. Nutritional products are fortified with free myo-inositol, but they may contain some inherent free, phosphatidylinositol bound myo-inositol, inositol phosphates, and phytate. The myo-inositol SPIFAN working group developed Standard Method Performance Requirements (SMPRs ® ) that were then approved by SPIFAN (3). A reference method capable of determining only free myo-inositol and myo-inositol bound as phosphatidylinositol in infant, adult, and pediatric formula powders; ready-to- feed (RTF) liquids; and liquid concentrates was needed since there were no myo-inositol reference methods. Traditional microbiological methods are not precise and accurate enough to meet the nutrient specification requirements of infant,