May | June 2013

An effective source of dietary methionine for the turbot Psetta maxima

The International magazine for the aquaculture feed industry

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64 | InternatIonal AquAFeed | May-June 2013

The turbot Psetta maxima is animportant cultured flatfish speciesin Europe and now increasingly inChina.Dietaryformulationsforthis

species typically have been dependent onhigh levels of fishmeal inclusion. Increasingcosts and the decreasing availability offishmealhavenecessitatedloweringfishmeallevels and increasing plant proteins in feedformulations.

Insuchdietsmethioninecanbecomeoneof the first limiting amino acids and supple-mentation is frequently necessary tobalancedietsandachieveoptimumperformance.Thehydroxy analog of methionine, 2-hydroxy-4-methylthio butanoic acid (HMTBa) is asafe and effective source ofmethionine thathas been used to supplement methioninedeficient diets for livestock and aquaculturespecies.

HMTBa is structurally different fromL-methionine in that ithas ahydroxyl groupinstead of an amino group at theα-carbonposition, potentially reducing feed nitrogeninputs into grow out systems. It is passivelyabsorbed and is converted to L-methionineby D-hydroxy acid dehydrogenase andL-hydroxy acidoxidaseenzymeswhichhavebeen confirmed in tissues of shrimp andfish.Atwo-partstudywascarriedoutattheOceanUniversityofChina,Chinatoevaluate1. The response of turbot fed diets sup-plemented with either HMTBa (2-hydroxy-4-methylthio-butanoic acid)or L-methionine,and2.ThedynamicsofabsorptionofHMTBaandL-methionine.

Materials and methodsA 75-day growth trial was carried out

to evaluate the effects of HMTBa andL-methionine as dietary methionine sources

on the growth of juvenile turbot (initialweight5.6g.N=5tankspertreatment).Fivelevels(0.3,0.6,0.9,1.2and1.5%drymatter) of HMTBa (added as Mera™Met– 84 percent HMTBa; Novus InternationalInc., USA) and L-methionine were addedrespectivelytoapracticalbasaldiet,thatwaslimiting in methionine (0.59% methionine;0.42%cystine;1.01%totalsulfuraminoacids).This basal diet served as the control dietandcontained48percentcrudeproteinandapproximately12.5%crudelipid.

A crystallineL-amino acidpremix, whichwas devoid ofsulfur aminoacids was addedto the basal dietto approximatethe whole bodycompositionof the turbot.Different levelsofeitherHMTBaor L-methioninewere added tothe basal dietat the expenseof glutamic acidto give totalmethionine con-centrations rang-ing from 0.59- 2.09 and totalsulfuraminoacidconcentrationsranging from1.01to2.51%.

A secondstudy wasconducted to

evaluate the absorption of HMTBa andL-methionine from diets containing thesemethionine sources. The basal diet used inthesecondtrialwasgenerallysimilartothatused in the growth trial (0.75% methionineand 0.45% cystine; total sulfur amino acidconcentrationof1.20%).Thebasaldietwas

Figure 1: Weight gain and specific growth rate in juvenile turbot fed either HMTBa or L-methionine

An effective source of dietary methionine for the turbot Psetta maxima

by Rui Ma1, Huapeng Hou1, Wenbing Zhang1, Anant Bharadwaj2, Craig Browdy2 and Kangsen Mai1

38 | InternatIonal AquAFeed | May-June 2013

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May-June 2013 | InternatIonal AquAFeed | 39

supplementedwitheither0.75%HMTBaorL-methionine.Inthistriallargerfish(65g;45fish/tank;N=3)wereusedtofacilitatebloodsamplingattheendofthetrial.

Thebasalcontroldietandthetwoexperi-mental diets were fed to the turbot for aperiod of 14 days twice daily to apparentsatiation.Fishwere fasted12hprior to finalfeeding.Followingfeeding, fishweresampledat0,0.25,0.5,1,2,3,4,6,9,12,18and24hrespectivelyand theirbloodcollected.Bloodwas centrifuged and serum collected andstoredprior toanalysis. SerumwasanalyzedusingHPLCforHMTBaandfreemethionine.

ResultsThere were no significant differences in

survival between treatments. Fish fed the

basal diet (red bar)showed significantlylower weight gainthan all the othertreatments (Figure1). Weight gainin fish fed bothmethionine sourcesincreasedinaquad-ratic fashion withincreasing dietaryconcentration. Infish fed HMTBa(blue bars) maxi-mum response wasobserved at 0.9%supplementationand was significantly

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Figure 2a: Serum free-methionine concentrations in post-fed turbot fed the control, L-methionine and HMTBa diets

Figure 2b: Serum HMTBa concentrations in fish fed diets containing HMTBa

38 | InternatIonal AquAFeed | May-June 2013 May-June 2013 | InternatIonal AquAFeed | 39

FEATURE

higher than the response seen in fish sup-plemented 0.9% L-methionine. Generally, atdoses ranging from 0.6 to 1.5%, fish fedHMTBa showedequalorhigherweight gaincompared to fish fed L-methionine (greenbars). Similar responses were observed forother performance parameters such as finalweight, specific growth rate, feed efficiency,proteinefficiencyratioandproteinretention.

Results from the analysis of serumshow that the maximum serum concentra-tion of free-methionine (435.13µmol/L) inL-methionine-fed fish was observed at 9 h

after feeding (Figure 2a) whereas the maxi-mumserumHMTBa level (426.17µmol/L) inHMTBa-fed fish occurred 6 h after feeding(Figure2b).AppreciablequantitiesofHMTBawere observed in the serum of HMTBa-fedfish shortly after feeding as expected basedonpassivediffusionofHMTBaacrossthegutwallinfish.TwofreemethioninepeaksweremeasuredintheserumofHMTBa-fedfishat3-4 h and 12 h post-feeding. This suggestscontributionsfromeitherHMTBametabolismto L-methionine, tissue protein turnover ordigestion of intact dietary proteins. These

data suggest that HMTBa is absorbed intocirculationinfishshortlyafterfeedingandalsoshow that the maximum serum concentra-tionsofHMTBainHMTBa-fedfisharesimilarto serum free-methionine concentrations inL-methioninefedfish.

ConclusionsThe results of these trials confirm that

HMTBa is a safe and available source ofmethionine in practical diets for the turbotPsetta maxima. The HMTBa was absorbedefficiently into circulation at rates that aresimilarorbetterthanL-methionine.Dynamicsconfirmeddirectabsorptiondemonstratedinprevious livestock studies. Mera™Met canprovide a cost effective methionine sourcein reduced fishmeal formulations with 100percent bioavailability. HMTBa has beenshown in previous studies to improve feedattractability, providing an effective alterna-tive for optimizing performance and allow-ing for higher cost efficiencies through thereplacement of fishmeal by plant meals inaquaticfeeds.

More InforMatIon:1The Key Laboratory of Mariculture, Ocean University of China, ChinaEmail: wzhang@ouc.edu.cn

2Novus International Inc., USAEmail: craig.browdy@novusint.comWebsite: www.novusint.com

40 | InternatIonal AquAFeed | May-June 2013

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40 | InternatIonal AquAFeed | May-June 2013 May-June 2013 | InternatIonal AquAFeed | 41

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