Single-Laboratory Validation Study of a Rapid TD-NMR Method for Quantitation of Total Fat in Sunflower Oil Powder.

BACKGROUND: A rapid total fat quantitation method for sunflower oil powder was developed using time-domain nuclear magnetic resonance (TD-NMR). Currently, industry has three major methods for the total fat quantitation: gravimetric analysis after ether extraction (AOAC Methods 933.05 and 989.05), gas chromatography with flame ionization detector (GC-FID; AOAC Method 996.06), and High-resolution NMR. The gravimetric analysis method takes a day using highly flammable solvents, and the GC-FID method takes two days requiring harsh chemicals for hydrolyzation, extraction, and methylation. The High-resolution NMR spectroscopy method requires simpler sample preparation and shorter analysis time compared to the other two methods. Often, the only required sample preparation step is to dissolve a sample in a solvent. The acquisition time depends on types of analyzing nuclei and sample. The vegetable oil analysis by 13C NMR takes about 4 h per sample. 1H NMR usually takes less time to analyze. In contrast, the TD-NMR relaxometry method takes only 1 h to prepare and analyze samples if the test is for total fat only. The acquisition time is 40 s per sample, and samples are analyzed "as is". A rapid analysis method in a quality control laboratory is very crucial for laboratory efficiency in releasing products. In this paper, a single-laboratory validation study is described for a rapid TD-NMR method to quantitate total fat in sunflower oil powder. OBJECTIVE: This validation work is to provide documented evidence for the method validity as well as the method performance. METHOD: The method used a Bruker minispec mq-20 NMR analyzer® with minispec plus® software. A Hahn echo pulse program was used in the method to collect spin echo signal to determine total fat content. RESULTS: The linearity/range result from 10 standards (0, 21, 42, 63, 83, 92, 100, 108, 117, and 125%) has coefficients of determination (R2) of 1.0000. The 100% level is 1.2 g-fat in 2.5 g sample, which is targeted fat content in a sunflower oil powder raw material. The method is specific for the quantitation of total fat in sunflower oil powder with no background interference from the matrix. The precision result of the 6 replicate samples at 100% level is 0.3% RSD. The accuracies measured from triplicate analysis of 80, 100, and 120% sample matrices are 100, 100, and 100% average recoveries, respectively. The ruggedness of the test method is 0.4% RSD of 12 analysis from 2 analysts (6 results from each analyst) on the different days. CONCLUSIONS: The test method is proven to be specific, linear, precise, accurate, rugged, and suitable for the intended use of quantitative analysis for total fat in sunflower oil powder. HIGHLIGHTS: Traditional methods of gravimetric or GC-FID for total fat analysis of raw materials require lengthy sample preparation and experiment time. Laboratory needs to spend a day to perform gravimetric analysis following ether extraction method and 2 days for the GC-FID method. In addition, these test methods use highly flammable and harsh chemicals that generate hazardous chemical wastes. These hazardous wastes are harmful to analysts and environments. In contrast, the TD-NMR method is safe, environmentally friendly, and fast. Therefore, TD-NMR is a preferred method for quality control laboratories.

Single-Laboratory Validation Study of a Proton NMR Method for the Determination of L-Arginine, L-Citrulline, and Taurine Contents in Dietary Supplements.

BACKGROUND: A quantitative NMR (qNMR) method can provide rapid analysis compared to chromatographic methods. Sample preparation steps are relatively simpler and run time is shorter. Rapid analysis methods for release tests in quality control laboratories are very important for laboratory efficiency. Here, we describe a single-laboratory validation study for a rapid qNMR analysis of L-arginine, L-citrulline, and taurine in powdered and tablet dietary supplement products. OBJECTIVES: This validation work is to provide documented evidence for the qNMR method validity as well as method performance. METHODS: The method used Bruker 400 MHz high-resolution proton NMR spectroscopy for simultaneous determination of L-arginine, L-citrulline, and taurine contents in dietary supplement product 1 (powder) and dietary supplement product 2 (tablet). The absolute NMR quantitation is based on a principle of universal proton response intensity correlation with the number of protons in each target analyte (amino acids) vs. that of a reference standard (maleic acid). RESULTS: The test method performance was validated with dietary supplement-1 (powder) and dietary supplement-2 (tablet). The linearity of the method was studied from about 360 mg/g to about 675 mg/g of L-arginine; from about 15 mg/g to about 30 mg/g of L-citrulline; and from about 20 mg/g to about 40 mg/g of taurine in dietary supplement-1, and from about 15 mg/g to about 30 mg/g of taurine in dietary supplement-2. The coefficients of determination (R2) are 1.0000 for L-arginine, 0.9967 for L-citrulline, and 0.9995 for taurine in dietary supplement-1 and 0.9903 for taurine in dietary supplement-2. The accuracies measured from the sample matrices are 102%, 101%, and 100% average recoveries for 80%, 100%, and 120% concentration levels of L-arginine, 105%, 105%, and 103% average recoveries for 80%, 100%, and 120% concentration level of L-citrulline, and 101%, 102%, and 100% average recoveries of taurine for 80%, 100%, 120% concentration levels in dietary supplement-1; and 95, 98%, and 93% average recoveries of taurine for 80%, 100%, 120% concentration levels in dietary supplement-2, respectively. The precisions (RSD) are 1% for L-arginine, 5% for L-citrulline, and 2% for taurine in dietary supplement -1, respectively; and 4% for taurine in dietary supplement-2. The ruggedness of the test method is within 2%, 4%, and 2% for L-arginine, L-citrulline, and taurine for dietary supplement -1, respectively, and within 4% for dietary supplement-2. The method is specific for the quantitation of each nutrient with no background interference from the matrix for the proton peaks of L-arginine, L-citrulline, taurine, and maleic acid (standard). CONCLUSIONS: The test method is proven to be specific, precise, accurate, rugged, and suitable for intended quantitative analysis of L-arginine, L-citrulline, and taurine in powdered and tablet finished products. HIGHLIGHTS: The simultaneous determination of all three nutrients of L-arginine, L-citrulline, and taurine using proton NMR provides rapid analysis for quality control release tests that is more efficient versus that of two HPLC methods. Previously, our laboratory was using one HPLC method to analyze L-arginine and L-citrulline while using a second HPLC method to analyze taurine. That approach required two HPLC instruments and two analysts for parallel analysis that takes 2 days using volatile and flammable solvents for extraction and chemical derivatization. This rapid NMR method can analyze the sample "as is" with results obtained in less than 4 h, and is efficient, safe, and environmentally friendly. The initial higher NMR instrument investment versus two HPLC instruments is rewarded with high returns for continued quality control tests.