Furthermore, even if a reference standard for quantifying a food compound is currently commercially _FTble, the reagent manufacturer may terminate the supply because it becomes economically unviable. However, considerable effort is required to prepare purified or synthesized standards. Additionally, for naturally derived compounds such as food-related compounds, not all reference standards used for quantification are commercially available, requiring the isolation and purification, or chemical synthesis, of these compounds used as reference standards for quantification. Since the manufacturer guarantees these commercial standards' purity, it may not be metrologically accurate, resulting in questionable reliability of the obtained quantitative value. However, there are very few certified reference materials, and therefore quantification methods such as HPLC are typically used together with commercial standard products from reagent manufacturers as reference standards. An authentic and identical analyte standard, such as certified reference material (CRM) with proven identity and purity determined by valid metrological procedures, is thus required to obtain reliable analytical results for each analyte. However, the sensitivity of HPLC methods depends on the molar absorption coefficient of the specific analyte. Quantification is commonly accomplished by purifying a sample and analyzing it with chromatographic methods such as high-performance liquid chromatography (HPLC). It is crucial to ensure the quality and safety of food sold domestically and internationally using analytical evaluations, and thus there is a need to develop highly reliable analytical methods for various food-related compounds. The funder provided support in the form of salaries for TO but did not have any additional role in the study design, data collection, and analysis, decision to publish, or preparation of the manuscript.Д¬ompeting interests: The authors have declared that no competing interests exist. This research was also financially supported by the Nihon University Fund for Supporting Education and Research (TO). The funder had no role in study design, data collection, and analysis, decision to publish, or preparation of the manuscript. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Дata Availability: All relevant data are within the manuscript.ДЇunding: Takashi Ohtsuki (TO) was funded by JSPS KAKENHI ( ). Received: AugAccepted: NovemPublished: December 3, 2020Д¬opyright: В© 2020 Ohtsuki et al. PLoS ONE 15(12):Д®ditor: Walid Elfalleh, Higher Institute of Applied Sciences and Technology of Gabes University of Gabes, TUNISIA (2020) Development of an HPLC method with relative molar sensitivity based on 1H-qNMR to determine acteoside and pedaliin in dried sesame leaf powders and processed foods. Thus, the proposed RMS method is a useful and practical tool for determining acteoside and pedaliin in terms of the accuracy of quantitative values, the routine analysis, and the cost of reagents.Д¬itation: Ohtsuki T, Matsuoka K, Fuji Y, Nishizaki Y, Masumoto N, Sugimoto N, et al. Moreover, the proposed method yields analytical data with SI-traceability without the need for an authentic and identical analyte standard. In this study, the content of acteoside and pedaliin in several samples, such as dried sesame leaf powders and commercially processed foods, can be determined by the proposed RMS method and demonstrated in good agreement that obtained by a conventional method. Therefore, this method can quantify several analytes with metrological traceability to the International System of Units (SI) using the RMS and one alternative reference standard. Each RMS is also calculated from the ratio of each analyte's molar absorption coefficient to that of MHB after correcting the purities of the analytes and reference standard by 1H-qNMR. This method is used methyl 4-hydroxybenzoate (MHB) as an alternative reference standard. A high-performance liquid chromatography (HPLC) method with relative molar sensitivity (RMS) based on 1H quantitative NMR spectroscopy ( 1H-qNMR) has been developed for food ingredients such as acteoside (verbascoside) and pedaliin (pedalitin-6- O-glucoside) without requiring authentic and identical standards as the reliable analytical methods.

Hplc analysis procedure