Gluten Exorphin A5

The effect of exogenous opioid peptides, gluten exorphins A5 and B5, which were isolated from the enzymatic digest of wheat gluten, on the postprandial insulin level were examined in rats. The oral administration of gluten exorphin A5 at a dose of 30 mg/kg w. potentiated the postprandial plasma insulin level and the effect was reversed by naloxone. The administration of gluten exorphin B5 showed a similar effect at a higher dose (300 mg/kg w). Furthermore, intravenous administration of gluten exorphin A5 at a dose of 30 mg/kg w. also stimulated the postprandial insulin release. The fact that orally and intravenously administered gluten exorphin A5 stimulates insulin release suggests that it modulates pancreatic endocrine function by the action after the absorption rather than within the the gastrointestinal tract.

Central effects of gluten exorphin A5 (Gly-Tyr-Tyr-Pro-Thr), a fragment from wheat gluten, were studied on the pain-inhibitory system, emotionality and learning/memory processes in mice. Orally administered gluten exorphin A5 produced neither an antinociceptive effect nor an effect on morphine analgesia. Intracerebroventricularly (i.c.v.) administered gluten exorphin A5 produced mild but significant antinociception in a dose-depepndent manner, while not affecting the morphine analgesia. On the other hand, oral gluten exorphin A5 suppressed the endogenous pain-inhibitory system, i.e., antinociception induced by socio-psychological- (PSY-) stress (SIA) using a communication box; intraperitoneal gluten exorphin A5 abolished both footshock- (FS-) stress-induced antinociception (SIA) and PSY-SIA; and i.c.v. gluten exorphin A5 suppressed FS-SIA, but rather potentiated PSY-SIA. This peptide given by these routes was without effect on forced swim-SIA. In addition, oral gluten exorphin A5 tended to prolong the retention time on open arms in the elevated plus-maze test. Finally, oral gluten exorphin A5 when given during the post-training period of learning/memory processes significantly increased the latency into the dark compartment in the one-trail step-though type passive avoidance test, indicating that the peptide also facilitates the acquire/consolidation process of learning/memory. Thus, gluten exorphin A5 has been found to produce various effects not only in the peripheral nervous systems but also in the central nervous system.

In the present work, for the first time, a method for the quantification of the alimentary opioid peptide Gluten Exorphin A5 (GE-A5; Gly-Tyr-Tyr-Pro-Thr) in cerebrospinal fluid (CSF) was developed. Aliquots (5 microL) of CSF were injected into a liquid chromatography-mass spectrometry (LC-MS) instrument equipped with a reversed-phase C18 column at a flow-rate of 0.4 mL/min. The mobile phase consisted of Eluent A water with 0.6% acetic acid as an ion-pairing reagent and Eluent B acetonitrile/methanol (75:25, v/v). The LC-MS system was programmed to divert column flow to waste for 4 min after injection, after which time flow was directed into the mass spectrometer that operated in positive ion mode. No significant interfering peaks were detected at the retention times of GE-A5 in CSF blanks. The lower limit of detection and the lower limit of quantitation values for GE-A5 in CSF were established at 0.60 and 1.50 ng/mL, respectively. The intra- and inter-day precision values were <5% relative standard deviation. The intra- and inter-day accuracy were 99.6-102.8% and 100.0-101.9%, respectively. The reported assay employs extremely small volumes of CSF, thus allowing the analysis of GE-A5 from both small and large animal models.