Chromatographic fractions of a rough extract of echinoid spawn (REES) is demonstrated to efficiently induce oocyte maturation in aspidochirote holothuroids. The method is so efficient that it is currently used in holothuriculture to obtain fertilised eggs even outside the reproductive period of the aquacultivated species. In the present study, we isolated and identified from echinoid spawns the molecule responsible for the induction of the oocyte maturation in the Mediterranean holothuroid Holothuria tubulosa and the Indo-Pacific Holothuria scabra. The use of proteinase K and dialysing membranes suggests that the active molecule is a protein with a molecular weight superior to 12,000 kDa. The active molecule has been isolated on G-100 Sephadex chromatography column. Active chromatographic fractions include seven proteins identified with nanoLC-MS/MS technique. One of them, identified as a thioredoxin-2 and to which the name Trx-REES was given, gave levels of maturation similar to those obtained with the REES and with a commercial thioredoxin extracted from Escherichia coli. Occurrence of thioredoxin in REES was confirmed by immunoblot analysis, and the maturation-inducing properties of thioredoxin were positively checked using anti-thioredoxin antibodies. A peptide of 6 AAs corresponding to the active site of Trx-REES, composed of WCNPCK, was synthesised and its efficiency in holothuroid oocyte maturation was tested. At some concentrations, the isolated peptide was 1.2 times more active than the REES. This short peptide may be used as an effective tool for artificial induction of oocyte maturation in the sea cucumber aquaculture context.