Molecular Brain Research 88 (2001) 155–160 www.elsevier.com / locate / bres Research report Presence of morphine and morphine-6-glucuronide in the marine mollusk Mytilus edulis ganglia determined by GC / MS and Q–TOF–MS Starvation increases opiate alkaloid levels a b a a,c c Wei Zhu , Geert Baggerman , Yannick Goumon , Federico Casares , Bruce Brownawell , a, * George B. Stefano a Neuroscience Research Institute, State University of New York, College at Old Westbury, Old Westbury, NY 11568, USA b Zoological Institute, Catholic University of Leuven, 3000 Leuven, Belgium c Marine Sciences Research Center, State University of New York at Stony Brook, Stony Brook, NY 11794-5000, USA Accepted 30 January 2001 Abstract Morphine and morphine-6-glucuronide, a morphine metabolite, have been identified and quantified in Mytilus edulis pedal ganglia at a level of 2.6760.44 and 0.9860.14 ng / ganglia, respectively by high performance liquid chromatography coupled to electrochemical detection. These opiate alkaloids were further identified by both gas-chromatography mass spectrometry and nanoflow electrospray ionization double quadrupole orthogonal acceleration Time of Flight mass spectrometry. In animals that were starved, the morphine level rose to 6.3860.88 ng / ganglion and the morphine 6-glucoronide rose to a level of 23.063.2 ng / ganglion after 30 days. These studies demonstrate that opiate alkaloids are present as naturally occurring signal molecules whose levels respond to stress, i.e., starvation. Opiate alkaloids were not found in the animal’s incubation media or food, demonstrating their synthesis occurred in the respective tissue. These new method of opiate alkaloid detection, conclusively proves that morphine and morphine-6-glucuronide are present in animal tissues.  2001 Elsevier Science B.V. All rights reserved. Theme: Neurotransmitters, modulators, transporters and receptors Topic: Transmitters in invertebrates Keywords: HPLC; Opiates; Ganglia; GC / MS; Q–TOF; Morphine; Morphine 6 glucuronide 1. Introduction tography (HPLC) coupled to electrochemical detection [13,16,18,20]. Recently, via GC / MS morphine and mor- Endogenous morphine has been identified in mamma- phine 6 glucuronide (M6G) has also been identified in a lian neural tissues by various techniques, including gas- parasitic worm and marine bivalve [9,10]. chromatography mass spectrometry (GC/MS) [2,3,5– In the present report we identify morphine by another 7,11,12,23,24]. Recently, we have extended these studies more sensitive methodology, namely, nanoflow electro- to include invertebrate molluscan ganglia [16,18,20]. In spray ionization double quadrupole orthogonal acceleration regard to invertebrate morphine identification, its charac- Time of Flight mass spectrometry (Q–TOF MS). We also terization has been by way of immunocytochemistry, demonstrate that morphine and M6G are present within the radioimmunoassay and high performance liquid chroma- pedal ganglia of the marine mollusk Mytilus edulis. Furthermore, starvation results in a statistically significant increase in endogenous ganglionic opiate alkaloid levels, *Corresponding author. Tel.: 11-516-876-2732; fax: 11-516-876- demonstrating that morphinergic signaling is involved in 2727. E-mail address: gstefano@li.net (G.B. Stefano). the organism’s stress response. 0169-328X / 01 / $ – see front matter  2001 Elsevier Science B.V. All rights reserved. PII: S0169-328X(01)00048-1