【Snow Brand Bean Stark】
The 3rd National Breast Milk Survey Developed a method for measuring pyrroloquinoline quinone (PQQ), a functional ingredient contained in breast milk
Snow Mark Bean Stark The third national breast milk survey Development of a method for measuring pyrroloquinoline quinone (PQQ) functional components contained in breast milk We announced at the 2019 Annual Meeting of the Japanese Society of Agricultural Chemistry ………………………………………………………………………………………… Snow Brand Bean Stark Co., Ltd. (Head office: Shinjuku-ku, Tokyo President & Representative Director: Mr. Inaba) has jointly conducted the third nationwide breast milk survey with Snow Brand Meg Milk Co., Ltd. As part of these surveys, we developed a method for measuring pyrroloquinoline quinone (PQQ), which is one of functional components contained in breast milk, in collaboration with Tohoku University. The following is an oral presentation of this research result at the 20th Annual Meeting of the Japan Society for Agro-Chemical Chemistry, held from March 24 at the Tokyo University of Agriculture Setagaya Campus and others.
◆ Summary summary ・ Pyrroloquinoline quinone (PQQ) is a water-soluble quinone compound that has been discovered as one of the coenzymes of oxidoreductases and is attracting attention as a vitamin B-like substance. Recent research on PQQ has reported functions such as brain function improvement, antioxidative action and antidiabetic action, and it is a functional ingredient that is attracting attention as a health food material. ・ PQQ has been reported to be contained in milk and human breast milk, but because there are few analysis examples and different extraction methods and measurement principles, quantitative discussions have been difficult. -As a collaborative research between Tohoku University and Snow Mark Bean Stark, a new method for analyzing PQQ in breast milk by combining an extraction method using internal standard substances with a highly sensitive and highly selective LC-MS / MS method Developed. -From the measurement results, it was found that PQQ, which is hardly contained in milk, is present in human breast milk, and a part is present as an amino acid adduct of PQQ. ・ From now on, we will conduct analysis of various human milks with different lactation period to clarify the physiological significance in human milk.
◆ Japanese Society of Agricultural Chemistry 2019 presentation summary Title LC-MS / MS analysis of pyrroloquinoline quinone (PQQ): Measurement of milk and human milk Presenter ○ Kato Capital Tax 1, Kawai Shoko 1, Kobayashi Shunjiro 2, Ito Satoshi 1, Nagatsuka Takahiro 1, Nakagawa Kiyotaka 1 1. Tohoku University, Agricultural and Functional Molecular Analysis 2. Snow Brand Bean Stark Co., Ltd. Product Development Department Date of presentation: March 26, 2019 (Tuesday) Venue Tokyo University of Agriculture Setagaya Campus (1-1-1 Sakuragaoka, Setagaya-ku, Tokyo)
◆ Summary of research presentation contents
Pyrroloquinoline quinone (PQQ) is a water-soluble quinone substance known as a bacterial coenzyme and is present in an experiment using PQQ as a coenzyme in PQQ deficiency symptoms (growth failure and decreased birth rate) and in mice. Was reported to be considered as a new vitamin B-like substance. Furthermore, it is reported that PQQ is contained in milk and human breast milk in a considerable amount, and its physiological significance is attracting attention. However, quantitative analysis is difficult because the PQQ analysis examples of milk and breast milk are still limited to the number method and the extraction method and measurement principle are different. In addition, PQQ and adducts of various amino acids (IPQ-R: R = amino acid side chain) are suggested to be present in human breast milk, but there is no analysis example of milk. Therefore, in this study, a quantitative method of PQQ and IPQ-R was constructed by combining extraction method using internal standard substance and highly sensitive and highly selective LC-MS / MS method, and analyzed milk and human milk. PQQ and IPQ-R I tried to clarify the existence and quantity.
【Method and result】
The ionization conditions of MS / MS were optimized using PQQ2Na standard and IPQ (prepared by incubating a mixture of PQQ2Na and various amino acids) as a reference compound. LC was applied to MS / MS, separation was performed in a reverse phase system using an ion pair agent, and detection was performed in Multiple Reaction Monitoring (MRM) mode that allows selective analysis. As a result, PQQ and IPQ-R (17 types except Cys, Pro, IPQ derived from Trp) were successfully detected, PQQ quantification of ng / mL level by external standard method, and qualitative analysis of IPQ-R were possible. became. The analysis of a sample containing PQQ reliably (PQQ-producing culture medium supernatant; provided by Mitsubishi Gas Chemical Co., Ltd.) with this method can detect the peaks of PQQ and several types of IPQ-R, confirming the usefulness of this method. The Therefore, we decided to work on the analysis of several types of milk with different manufacturing processes and human milk of 1 month lactating period. Internal standard for these samples
C-PQQ (Mitsubishi Gas Chemical) is added and subjected to Oasis WAX, and the resulting eluate is subjected to ethyl acetate / acetonitrile extraction under strongly acidic conditions for LC-MS / MS analysis. did. As a result, PQQ and IPQ-R in milk were almost below the detection limit, while human breast milk contained PQQ, and its concentration was 1-3 ng / mL. In addition, human milk containing at least a glycine-derived IPQ was also identified. From the above results, it is inferred that PQQ, which is hardly contained in milk, is present in human breast milk, and part of it is present as IPQ. In the future, together with simultaneous determination of PQQ and IPQ-R, we plan to carry out analysis of various human milks with different lactation stages in order to clarify their significance in human milk.
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