title: Extraction and Quantification of Bioactive Tyrian Purple Precursors: A Comparative and Validation Study from the Hypobranchial Gland of a Muricid Dicathais orbita.
abstract: Muricidae are marine molluscs known for the production of Tyrian purple and bioactive precursor compounds. A validation study for the extraction and analysis of secondary metabolites found in the hypobranchial gland of the muricidDicathais orbitais reported, using high performance liquid chromatography-mass spectrometry (HPLC-MS) with diode array detector (DAD). Quantification of the dominant secondary metabolites fromD. orbitais described, followed by a comparison of solvent extraction procedures and stability studies. The intra- and inter-day relative standard deviation (RSD) for tyrindoxyl sulphate was 0.46% and 0.17%, respectively. The quantification was linear for standards murexine, 6-bromoisatin, and tyrindoxyl sulphate. The limits of detection were 0.03, 0.004, and 0.07 mg/mL, respectively, and the limits of quantification were 0.09, 0.01, and 0.22 mg/mL, respectively. The results showed that alcoholic solvents were better for extracting choline ester and indoxyl sulphate ultimate precursors, while chloroform was more suitable for the extraction of the intermediate precursors. Multivariate analysis revealed significant differences in extract composition according to the solvent used. Stability testing showed an increase of the oxidative compounds 6-bromoisatin and putative tyrindoxyl S-oxide sulphate in the ethanol extracts while more degradation products were seen in the chloroform extracts after months of cold storage. The validated method was found to be simple, reproducible, precise, and suitable for quantification of the secondary metabolites of muricid molluscs for dye precursor and nutraceutical quality control, as well as applications in marine chemical ecology.
authors.affiliation: Marine Ecology Research Centre, School of Environment, Science and Engineering, Southern Cross University, Lismore, NSW 2480, Australia. r.regino.10@student.scu.edu.au.
authors.forename: Roselyn
authors.initials: R
authors.lastname: Valles-Regino
authors.affiliation: Southern Cross Plant Science, Southern Cross University, Lismore, NSW 2480, Australia. peter.mouatt@scu.edu.au.
authors.forename: Peter
authors.initials: P
authors.lastname: Mouatt
authors.affiliation: Marine Ecology Research Centre, School of Environment, Science and Engineering, Southern Cross University, Lismore, NSW 2480, Australia. david.rudd@scu.edu.au.
authors.forename: David
authors.initials: D
authors.lastname: Rudd
authors.affiliation: Marine Ecology Research Centre, School of Environment, Science and Engineering, Southern Cross University, Lismore, NSW 2480, Australia. lachlan.yee@scu.edu.au.
authors.forename: Lachlan H
authors.initials: LH
authors.lastname: Yee
authors.affiliation: Marine Ecology Research Centre, School of Environment, Science and Engineering, Southern Cross University, Lismore, NSW 2480, Australia. kirsten.benkendorff@scu.edu.au.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Benkendorff
mesh_terms: D000818:Animals; D001934:Branchial Region / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D049851:Gastropoda / Q000378:metabolism*; D007211:Indoles / Q000737:chemistry*; D013058:Mass Spectrometry
chemical_list: C553805:6,6'-dibromoindigo; D002457:Cell Extracts; D007211:Indoles
keywords: brominated indole; liquid chromatography–mass spectrometry; marine mollusc; marine natural products; nutraceutical