In vivo absorption and metabolism of leptosperin and methyl syringate, abundantly present in manuka honey.

SCOPE: Manuka honey, which shows strong nonperoxide-dependent antibacterial activity, contains unique components, such as methyl syringate 4-O-ß-D-gentiobioside (leptosperin) and its aglycone, methyl syringate (MSYR). To determine the potential for biological activity evoked by the ingestion of leptosperin and MSYR, we investigated the absorption and metabolism of these components in manuka honey. METHODS AND RESULTS: The incubation of MSYR with liver microsomes or S9 fractions in vitro resulted in the formation of MSYR-glucuronide (MSYR-GA), MSYR-sulfate (MSYR-S), and syringic acid as metabolites. Then, manuka honey (15 g) was fed to healthy human volunteers. MSYR-GA, MSYR-S, and MSYR were detected in both plasma and urine. Within plasma, their levels were highest within 0.5 h to 1 h post-ingestion, and most metabolites disappeared within 3 h. In conjunction with the disappearances, a significant amount of metabolites along with trace leptosperin was excreted in urine within 4 h. To elucidate the detailed metabolisms of leptosperin and MSYR, each compound was separately administered to mice. In each case, MSYR-GA, MSYR-S, and MSYR were detected in both plasma and urine. CONCLUSION: This study shows the major molecular pathway for leptosperin and MSYR metabolism and could facilitate an understanding of biological functions of manuka honey post ingestion.

Characterization of a Monoclonal Antibody against Syringate Derivatives: Application of Immunochemical Detection of Methyl Syringate in Honey.

Syringic acid is one of the key skeletal structures of plant-derived chemicals. The derivatives of syringic acid have certain biological functions. In this study, a monoclonal antibody to syringic acid-based phytochemicals was prepared and characterized. The obtained antibody reacted with methyl syringate, syringic acid, and leonurine. Methyl syringate is a characteristic compound found in manuka honey, other honey varieties, and plants. Manuka honey was fractionated using HPLC, and the reactivity of the fractions with the antibody was examined. The antibody reacted with the fraction in which methyl syringate was eluted. The amount of methyl syringate in honeys as estimated by ELISA using the antibody had a good linearity compared with that estimated by HPLC. These results suggest that the antibody is applicable for the immunochemical detection of syringic acid derivatives in plants and foods.

Competitive immunochromatographic assay for leptosperin as a plausible authentication marker of manuka honey.

Manuka honey is known as one of the premium honeys because of its unique property: a potent antibacterial activity. Leptosperin, methyl syringate 4-O-ß-d-gentiobioside, has been specifically identified in manuka honey. Because leptosperin is relatively stable under warmer conditions, measuring leptosperin levels may be applied to authenticate manuka honey. In this study, an immunochromatographic separation and quantification of leptosperin techniques have been developed. The concentration of leptosperin measured by immunochromatography was significantly correlated with the concentration measured by high-performance liquid chromatography (HPLC) or enzyme-linked immunosorbent assay (ELISA). Because the immunochromatographic method is rapid and reliable, it could be applied to on-site quality control or inspection of honey samples by a beekeeper, a manufacturer, an inspector, a retailer, or a consumer.

Immunochemical authentication of manuka honey using a monoclonal antibody specific to a glycoside of methyl syringate.

Leptosperin, a novel glycoside of methyl syringate, is exclusively present in manuka honey derived from the Leptospermum species Leptospermum scoparium. Quantification of leptosperin might thus be applicable for authentication of honey. The concentration of leptosperin has high linearity with antibacterial activity. We established a monoclonal antibody to leptosperin and characterized the antibody in detail by a competitive enzyme-linked immunosorbent assay (ELISA), comparing the results with those of the high-performance liquid chromatography (HPLC) method for validation. The antigen in manuka honey was confirmed as leptosperin by HPLC fractionation with quantitation by an ELISA. Leptosperin contents of 50 honey samples were analyzed by an established ELISA, which can handle 20 samples (duplicate) on one 96-well plate. Significant coincidence with the chemical quantitation was observed. Immunochemical quantitation of leptosperin would be an economical and facile method for the possible authentication of manuka honey, allowing many honey samples to be processed and analyzed by an ELISA simultaneously.