title: Assessment of different sample processing procedures applied to the determination of melatonin in plants.
abstract: BACKGROUND
  Melatonin, an indoleamine well known in vertebrates and structurally related to other important substances such as tryptophan or indole-3-acetic acid, is also present in the plant kingdom although its specific function(s) remain to be established. The emerging field of melatonin studies in plants has progressed very slowly, mainly due to the problems associated with melatonin quantification in plants.
  
  
  OBJECTIVE
  Two commonly used procedures for plant samples are compared. The analytical characteristics of both procedures are quantitatively presented using different solvents and small amounts of fresh biological material, and the respective recovery rates and quantitative limits are presented. Some improvements are suggested.
  
  
  METHODS
  Two different sample extraction procedures were compared: a direct-sample extraction (DSE) and a homogenised- sample extraction (HSE). Melatonin was then determined in the respective plant samples by HPLC with fluorescence detection.
  
  
  RESULTS
  Using the DSE procedure, more than 94% melatonin was recovered from standard solutions, whereas levels higher than 93% were recovered from the spiked plant samples, with little difference between ethyl acetate and chloroform extractions. In the case of HSE, the recoveries of melatonin were approximately half and never higher than 55%. The ultrasonic treatment proposed in the DSE procedure showed different levels of efficiency (2-20%), depending on the sample.
  
  
  CONCLUSIONS
  This study has established that, with the direct sample extraction procedure, higher recovery rates are obtained both in standard solutions and in plant samples. The straightforwardness and reproducibility of the extraction procedure is accompanied by the high sensitivity obtained with fluorescence detection.
authors.affiliation: Department of Plant Physiology, Faculty of Biology, University of Murcia, 30100-Murcia, Spain. marino@um.es
authors.forename: Marino B
authors.initials: MB
authors.lastname: Arnao
authors.forename: Josefa
authors.initials: J
authors.lastname: Hernández-Ruiz
mesh_terms: D000975:Antioxidants / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D001467:Hordeum / Q000737:chemistry; D007202:Indicators and Reagents; D031297:Lupinus / Q000737:chemistry; D008550:Melatonin / Q000032:analysis*; D010936:Plant Extracts / Q000032:analysis; D010944:Plants / Q000737:chemistry*; D012015:Reference Standards; D012044:Regression Analysis; D012639:Seeds / Q000737:chemistry; D012997:Solvents; D013048:Specimen Handling; D013050:Spectrometry, Fluorescence
chemical_list: D000975:Antioxidants; D007202:Indicators and Reagents; D010936:Plant Extracts; D012997:Solvents; D008550:Melatonin