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Received 14 February 2011Received in revised form 6 June 2011Accepted 30 August 2011Available online 19 September 2011

Keywords:

into food and beverage products. The use of lipases for selective concentration of EPA and DHA, or for

from sh oil. The main bioactive omega-3 fatty acids are cis-5,8,11,15,17-eicosapentaenoic acid (EPA) and cis-4,7,10,13,16,19-docosahexaenoic acid (DHA) (Fig. 1). EPA and DHA are essentialcomponents of healthy nutrition and have been shown clinicallyto decrease the risk of coronary heart disease, partly through an abil-ity to reduce serum triglyceride levels and help prevent secondaryheart attack. EPA and DHA are precursors to anti-inammatory

but have very low levels of EPA and DHA. The major health benetof the consumption of ALA is that it converts to EPA and DHA.However, in the human body ALA is converted to EPA and DHAat an efciency of only 510% for EPA and 15% for DHA (Davis &Kris-Etherton, 2003). Most sh oils do not contain more than a30% combined level of EPA and DHA. For example, the primarysources of most commercially used omega-3 sh oil is anchovy(Engraulis ringens) and sardine (Sardinops sagax sagax) oils thatcontain1522% of EPA and 915% of DHA. This natural oil is normallyknown as 1812 triglyceride (TG) sh oil. However, oils of

Corresponding author.

Food Chemistry 131 (2012) 639644

Contents lists available at

he

lseE-mail address: cbarrow@deakin.edu.au (C.J. Barrow).1. Introduction

Omega-3 fats are long chain polyunsaturated fats containingmethylene-separated double bonds starting from the third carbonatom counted from the methyl-terminus. The presence of bis-allylic methylene groups and all double bonds being in the cis-con-guration makes these molecules prone to structural changes, par-ticularly oxidation, isomerisation and polymerisation.

Omega-3 fats are ingredients used in dietary supplements,healthy foods, and pharmaceutical products. These bioactive fattyacidshavewell establishedhealthbenets andareprimarilyderived

mediators and have demonstrated benets for the prevention ofinammatory mediated disorders including allergy, diabetes,Alzheimers disease and related neurodegenerative diseases (Lavie,Milani, Mehra, & Ventura, 2009).

2. The production of omega-3 concentrates from sh oil

Fish oil is the most abundant and the cheapest source of EPAand DHA. The best omega-3 terrestrial sources (seeds of ax,perilla, kiwifruit and chia) are very rich in a-linolenic acid (ALA)MicroencapsulationComplex coacervationOmega-3 fatsEPADHALipaseFunctional food0308-8146/$ - see front matter Crown Copyright 2doi:10.1016/j.foodchem.2011.08.085re-esterication reactions, is important in the production of omega-3 concentrates. Enzymatic strategiesrequire robust enzymes that can be immobilised and multiply re-used. Novel and mild processing meth-ods are particularly important for providing oils with good sensory properties, which are required for suc-cessful use as functional food ingredients. Although in some cases good quality oils can be used directly insome foods, such as margarine, many foods require that microencapsulated and stabilised omega-3 oilsbe used. This is particularly important when the oils are preconcentrated. There are a number of indus-trially used microencapsulation methods, but the most widely used are complex coacervates and spraydried emulsions. Fish oil is still the most widely used source of long-chain omega-3 fatty acids for addi-tion to food, although algal oil is the primary source of DHA for infant formula use in North America. Algaloil is still signicantly more expensive than sh oil for most applications, although many groups areimproving both the cost and quality of omega-3 oil from algal sources. In particular, Thraustochytridand Schizochytrid strains are a promising source of both DHA and EPA, and with further improvementcould be used to provide varying ratios of these omega-3 fats. In this short review we will describe someof the current research in omega-3 fat concentration and microencapsulation, with particular emphasison the use of lipases for concentration and complex coacervation for microencapsulation.

Crown Copyright 2011 Published by Elsevier Ltd. All rights reserved.Article history: Technology continues to evolve for the concentration and stabilisation of omega-3 fatty acids for deliveryA review of the progress in enzymatic coof omega-3 rich oil from sh and microb

Jaroslav A. Kralovec a, Shuocheng Zhang a, Wei ZhanaOcean Nutrition Canada Ltd., 101 Research Drive, Dartmouth, NS, Canada B2Y 4T6b School of Life and Environmental Sciences, Deakin University Geelong, VIC 3217, Austr

a r t i c l e i n f o a b s t r a c t

Food C

journal homepage: www.e011 Published by Elsevier Ltd. All rentration and microencapsulationl sources

Colin J. Barrow b,

SciVerse ScienceDirect

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vier .com/locate / foodchemights reserved.

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hemconcentration of 5090%, with controlled ratios of EPA and DHA,are preferred for many supplement and pharmaceutical applica-tions. Therefore, various groups have developed strategies for themanufacturing of highly concentrated EPA and DHA products.

Fish oil is a mixture of fatty acids (FA) in the TG form with EPAand DHA as the major long-chain components. Most oils containonly one EPA or DHA fatty acid per TG molecule. If we assume eachTG molecule contains only one DHA or EPA in an 1812 TG, then thefatty acids must be removed from the glycerol backbone to enableconcentration beyond 33% of EPA and DHA. Chemical ethanolysisresults in complete removal of all fatty acids from the glycerolbackbone and the formation of ethyl esters (EE), which can thenbe concentrated using fractional distillation or urea complexation.Ethanolysis converts the TG molecules to EEs reducing the averagemolecular size to one third and consequently the boiling points arereduced by about fty percent. For instance, the estimated boilingpoint of DHA-TG at atmospheric pressure is 907 65 C, whereasthe boiling point of DHA-EE is only 444 24 C (calculated usingAdvanced Chemistry Development (ACD/Labs) Software Solaris V4.67). The lower boiling point means that EEs are more easily dist-iled than are TGs. This together with the separation of fatty acidsfrom the TG glycerol means that EEs are used as starting materialsfor concentration of EPA and DHA.

There have been reports that EE concentrates are less bio-avail-able than TG concentrates and there is an argument that TG formsare natural whereas EE forms are not (Lawson & Hughes, 1988).Therefore, some regulatory environments demand that nutritionalsupplements contain oil in TG form, and so the production of TGconcentrates was achieved early on by converting EE concentratesback to the TG form by chemical reactionwith glycerol. This transe-sterication was carried out using traditional chemistry involvingthe use of strong bases and harsh reaction conditions, resulting inextensive formation of side-products. The use of lipases as catalystsoffers a milder re-esterication method, resulting in less byprod-ucts and better quality oils. It is well known that lipases catalyse

cis-4,7,10,13,16,19-D

cis-5,8,11,14,17-Eico

Fig. 1. Chemical stru

640 J.A. Kralovec et al. / Food Chydrolysis of lipids at the water lipid interface. However, it is lessknown that under anhydrous conditions lipases also effectivelycatalyse synthesis of esters (Zaks & Klibanov, 1985). Lipases arefrequently used in the production of pharmaceuticals. Examplesinclude the use of Candida rugosa lipase in the synthesis of serumcholesterol reducer lovastatin, Seratia marcescens for the produc-tion of coronary vasodilator diltiazem, and Candida antarctica lipaseB (CALB) for the production of anti-inammatory urbiprofen andpain killer baclofen. In addition, lipases are biodegradable and havea negligible biological oxygen demand in the waste stream.

Ocean Nutrition Canada Ltd. (ONC) has successfully convertedEE to TG both directly and via an intermediate hydrolysis stepthrough the free fatty acid (FFA) form (EE-FFA rst and thenFFA-TG) using immobilised CALB. Re-esterication from FFA con-centrate was signicantly faster than that from EE and could becarried out at lower temperatures and gave products in higheryields. After routinely achieving 90% conversion from FFA to TGin the laboratory with multiple re-use of the biocatalyst, the reac-tion was gradually scaled-up to manufacturing using proprietarypacked enzyme bed reactors. A plant assembly of four reactors al-lows us to manufacture up to 7500 kg of re-esteried TG per day(unpublished results). Other researchers have used lipases toincorporate EPA and DHA into food grade oils. For example, Ha-mam and Shahidi used a variety of lipases, including CALB, to suc-cessfully incorporate levels of between about 30% and 60%, EPA,DPA or DHA into high-laurate canola oil (Hamam & Shahidi,2006). These types of structured lipids containing both medium-chain and omega-3 fatty acids could become tailored functionalfood ingredients with specic bioactivity, bioavailability and sta-bility proles. A recent study shows that correctly immobilised li-pases can be used with organic solvents to improve specicity. Bycross-linking immobilised Rhizomucor miehei lipase (RML) withpolyfunctional polymers these researchers were able to used 2-propanol to increase EPA to DHA selectivity of this enzyme, result-ing in a product with 22:1 ratio of EPA to DHA (Fernandez-Lorenteet al., 2011).

Despite success with enzymatic EE to TG conversion, there havebeen questions about how much structural resemblance there-esteried TG share with the original natural 1812 TG oils. Unfor-tunately, CALB is not regioselective and not surprisingly a regio-specic analysis demonstrated differences between natural 1812TG and its re-esteried (1812 TG) counterpart (Fig. 2). That is, nat-ural 1812 TG has more DHA at position 2 with a close to equaldistribution of EPA across all three positions of glycerol, whilere-esteried concentrate has a statistical distribution of all fattyacids across the three positions on glycerol. Of course, all our TGconcentrates made by re-esterication of EE concentrates contain60% or more EPA and DHA and thus there will always be differ-ences between natural 1812 TG oils and TG concentrates such as4020 TG or 0555 TG, simply because these oils contain twice asmuch of DHA and EPA, so that these long-chain fatty acids replacealmost a third of FFA residues in the starting oil. Shahidi and

sahexaenoic Acid

entaenoic AcidO

O

(DHA)

(EPA)

OH

OH

es of EPA and DHA.

istry 131 (2012) 639644co-workers have recently shown that chemical randomisation ofEPA, DPA and DHA in both seal blubber oil and menhanden oilresults in more even distribution of these omega-3 fats amongthe terminal sn-1,3 and middle sn-2 position versus the naturaloils. This in term appears to lead to decreased oil stability, with in-creased omega-3 at the more exposed sn-1,3-positions, althoughresults were partly confounded due to modication in levels ofa-tocopherol antioxidant particularly in the seal oil (Wang,Reyes-Suarez, Kralovec, & Shahidi, 2010).

To conserve the original positional distribution of FA residues inthe natural triglycerides we developed an alternative concentra-tion process. In the rst step the starting 1812 TG oil was hydroly-sed in the presence of Thermomycetes lanuginosus lipase. The goalwas to remove shorter chain FAs while retaining EPA and DHAon the glycerol backbone. Using this strategy the degree of hydro-lysis was in 4555% range. After hydrolysis and stripping FFA offusing distillation, the EPA and DHA content of the glyceride portion

form

hemwas increased to 23% and 15%, respectively. In the second step thegenerated free hydroxyl groups of the produced diglycerides werere-esteried with a number of suitable sources of EPA and DHA inthe presence of CALB. For instance, 4020 FFA concentrate whenused as EPA and DHA donor boosted the levels of EPA and DHAto 28% and 17%, respectively, thus increasing EPA and DHA contentof the starting 1812 TG by about 50%. Both, the hydrolytic andesterication steps were monitored and analysed using severalstandard and modied methods. In the case of reactions consum-ing or generating FFA, a simple determination of acid values byalkalimetric titration was used. The positional distribution of theoriginal oil remained conserved after hydrolysis followed by re-assembly, without concentration, as assessed by NMR.

Although the importance of positional distribution of fatty acidresidues in omega-3 oils and their concentrates is a matter ofdebate, we developed a strategy for retention of positional distri-bution based on saturate removal, saturate removal followed byre-esterication. The oils obtained by selective enzyme hydrolysiswere shown to have superior stability, superior sensory proles,minimum polymer levels, trans-isomers and migration of doublebonds. However, it should be noted that the positional retentionstrategy has to date produced concentrates with signicantly low-er EPA and DHA levels than those made by EE to TG conversion

Fig. 2. 13C NMR spectra of starting 1812 TG oil and the oil after hydrolysis by lipasein the presence of immobilised CALB (Novozym 435) under anhydrous conditions.J.A. Kralovec et al. / Food C(45% vs. 60% EPA and DHA level). The lower levels of EPA andDHA may be an important reason for better stability and sensoryproles. The use of lipases has the potential to enable the produc-tion of more stable oils with good sensory properties, and havingFA distributions more representative of the natural TG oils.

3. Microbial sources of omega-3 concentrates

Two important issues have been raised with regard to the use ofsh oil as a source of EPA and DHA. Firstly, some sh oils containman-made pollutants such as methyl mercury, polychlorinatedbiphenols (PCBs) and dioxins. Secondly, some sh stocks fromwhich sh oil is derived, such as tuna, might not be sustainabledue to declining sh stocks. In contrast, microbial oils are sustain-able sources of EPA and DHA and their stability can also be morecontrollable than that of sh oils. The microbial oils are often lesscomplex than sh oils, exhibiting a simpler fatty acid proles.These oils also can have varying ratio of EPA to DHA and often havenatural antioxidants present that can help protection the oils fromoxidative damage during processing.

Various microbial strains could serve as producers of specialtyoils thatwouldmeetdesiredspecications. In1990s, somemicrobialsources of omega-3 oilswere developed, particularly for the produc-tion of DHA for infant formula. DHA and arachidonic acid (20:4 n-6,AA) are the dominant long-chain fatty acids in breast milk and playvital roles in neonatal development. As the ratio of AA:EPA:DHA inmothers milk (2.0:0.2:1.0, w/w) is completely different from thatin sh oils, with certain exceptions such as tuna oil, these sh oilsare unsuitable for the use in infant formulae where high DHA andlowEPA is preferred. Somemicrobial oils are rich inDHAandcontainlittle EPA and so can be combinedwith microbial produced AA to trequirements for infant formulae. For example, Martek developedfermentation processes for Crypthecodinium cohnii to commerciallyproduce microbial oil rich in DHA (4050%), and in collaborationwithDSMproducedmicrobial oils rich inAAusingMortierella alpina.Recently DSM has purchased Martek and so Martek is now a fullyowned subsidiary of DSM. Currently, Marteks microbial oils areadded tomost infant formula in the USA, and are being sold in morethan 60 countries. The use ofmicroalgae oils as sources of fatty acidsfor infant formula has been extensively reviewed elsewhere (Beh-rens & Kyle, 1996; Wright, Coverston, Tiedeman, & Abegglen,2006). The company also uses a Schizochytrium species to commer-cially producemicroalgal oil high in DHA. Lonza Group, Switzerlandis also a very important player in this eld, as it commercially pro-ducesmicroalgal DHAoil (4550%) using aUlkenia species (Kiy, Rus-

Thermomycetes lanuginosus (TL 100) followed by reassembling back to triglyceride,

istry 131 (2012) 639644 641ing, & Fabritius, 2005). Commercial production processes arecurrently under development in Australia, China, India, Japan, Spain,Norway and Canada (Raghukumar, 2008).

Traditional strain improvement techniques using mutagenesishave successfully been applied in enhancing microbial PUFA oilproductivity and developing specialty oil strains. Using thesemethods, Martek improved the lipid productivity of their DHAstrain by 242% in a 6-yr period. Similarly, Shimizus group suc-ceeded in breading mutant strains of M. alpina for the commercialproduction of specialty oils rich in DGLA (C20:3n-6) and other fattyacids, which also helped to elucidate the PUFA biosynthesis path-way in this microorganism (Sakuradani & Shimizu, 2009). Impor-tantly, strains developed via classic methods are not consideredas GMO. We have recently developed effective methods to ef-ciently trigger mutations and to change the FA proles of Thrausto-chytrid strains. Using classical strain improvement method, we aredeveloping modied of Thraustochytrid strains with unique fea-tures such as elevated PUFA productivity, the capability to produceEPA, and the capable of utilising cheap carbon sources such asglycerol.

We believe that the traditional methods will continue to play avital role in various aspects of strain improvement regardingspecialty oils and productivity improvement. However, many

using modern sequencing technologies, genetic pathway engineer-ing has become a robust technique for target-specic microbial

hemstrain development. Recently, two transformation systems havebeen developed for genetic manipulation ofM. alpina. One is basedon using a gene gun, and the other one is based on using Agrobate-rium tumefaciens as gene delivery system. The latter resulted inremarkably enhanced transformation efciency and transgene sta-bility. Using these methods, the elongase gene (EL2), a speed limitenzyme in AA biosynthesis, was overexpressed in M. alpina, whichenhanced AA productivity. These genetic tools enable the develop-ment ofM. alpina strains for specialty oils rich in n-9, n-7, n-6, n-4,n-3 or n-1 series PUFAs (Sakuradani & Shimizu, 2009). ThroughPUFA pathway engineering, DuPont has recently developed a yeaststrain for commercial production of oils rich in EPA (55% EPA) andthe company is hoping to develop various strains for the produc-tion of DHA, ARA or GLA (Sharpe et al., 2009). Additionally, usinga similar strategy, Microbia, Inc. recently has genetically engi-neered this oleaginous yeast into a carotenoid host for commercialantioxidant production (Grenfell-Lee, 2009).

At ONC, an isolation programme targeting Thraustochytridsfrom various Atlantic locations in Canada has been carried out.We have isolated and characterised a new strain ONC-T18 ofThraustochytrium sp., evolutionally most closely related to T. stria-tum T91-6. Our strain can not only produce up to 80% oil by dryweight, of which more than 31% are DHA, but also signicantamounts of carotenoid such as -carotene, astaxanthin, zeaxanthinand canthaxanthin. When compared to commercial DHA-product-ing microbial strains, our strain has one of the highest DHA produc-tivities, and has considerable potential as a commercial source ofhigh DHA oil (Burja, Radianingtyas, Windust, & Barrow, 2006).We and others have explored Thraustochytrids as producers ofPUFA microalgal oils. Since Thraustochytrids are effective highyielding lipid producers, efforts are being made to develop trans-formation systems aimed at engineer strains to improve theirDHA productivity and to enable them to produce high levels ofEPA and carotenoids. Recently, two genetic transformation systemshave been developed for a related Schizochytrium sp. One is basedon using electroporation and the other on using gene gun DNAdelivery method (Ono, Aki, & Kawamoto, 2006; Roessler, Mat-thews, Ramseier, & Metz, 2007). The PUFA biosynthetic pathwaysin this Schizochytrium sp. were elucidated by successfully knockingout PKS and FAS genes. Moreover, compared with the difculty ofdeveloping oleaginous yeast into EPA or carotenoid productionhosts through multiple steps of genetic engineering, Thraustochy-trid strains have natural PUFA and carotenoid biosynthetic path-ways with high activity, making them ideal hosts for channelingmetabolic uxes through genetic engineering. Overexpression ofthe carotene synthase gene indeed increased carotenoid productiv-ity in a Schizochytrium sp. by about 16 times (Weaver, Metz, Kuner,& Overton, 2006). Enhancing the levels of antioxidants such ascarotenoids in PUFA producing Thraustochytrid strains may alsosimplify the microalgal oil extraction procedures and extend theshelf life of microbial oils rich in PUFA. However, transformationsystems for other Thraustochytrid strains have not been demon-strated. We have been developing transformation systems for ournovel Thraustochytrid strain and have had some success using agene gun.

4. Stabilisation and delivery of omega-3 fatty acid rich oils intofoodstraditional methods are time-consuming and labour-intensive. Asmicrobial genomes can be sequenced in a straightforward way

642 J.A. Kralovec et al. / Food CEPA and DHA are unstable and will oxidise quickly leading tothe formation of unpleasant smelling and tasting aldehydes andketones. Various types of antioxidants have been used for thechemical stabilisation of EPA and DHA, but these are not adequateto enable sensory stabilisation of these oils for addition to manyfoods and beverages. The use of antioxidants to improve the stabil-ity of omega-3 containing oils has been reviewed recently (Shahidi& Zhong, 2010a, 2010b). A large body of work has been done to de-velop improved antioxidant systems for stabilisation of omega-3oils in food products. Mechanisms of oxidation are complex andvary depending upon the food matrix. This is particularly true forlipid emulsions, where antioxidants are normally exposed directlyto the food matrix components. The mechanisms of lipid oxidationin food systems has recently been reviewed (Waraho, McClements,& Decker, 2011) as have methods for evaluating the efcacy ofantioxidants in these systems (Mickael et al., 2010). Lipid emul-sions have seen some success in beverage products in particular,such as milk, where emulsication is a key step in the product pro-cess. Even in short shelf-life products such as milk, salad dressingand yoghurt it is necessary not only to optimise antioxidant com-binations for each specic application (Let, Jacobsen, & Meyer,2007; Nielsen, Klein, & Jacobsen, 2009), but additives, such as che-lators are also often required (Hu, McClements, & Decker, 2004). Toensure adequate protection of a wide range of food and beverageproducts against oxidation microencapsulation is required.

Microencapsulation is a unique process that has been used notonly to convert liquids to solids, but also to add functionalities orimproved oxidative stability to ingredients. Its advantages include:masking the unpleasant avours and odours of the microencapsu-lated ingredients; protecting ingredients from oxidation and otherunwanted reactions and therefore extending shelf life; controlledrelease of ingredients to improve the functionality of food addi-tives and expanding the application range of food ingredients(Desai & Park, 2005; Gharsallaoui, Roudaut, Chambin, Voilley, &Saurel, 2007; Gibbs, Kermasha, Alli, & Mulligan, 1999; Gouin,2004; Madene, Jacquot, Scher, & Desobry, 2006; Thies, 2001).Microencapsulation techniques can be divided into three classes:physical processes such as spray drying, spray chilling/coating,extrusion or uidised bed coating; chemical processes such asmolecular inclusion or interfacial polymerisation; and physico-chemical techniques such as single- or multi-core coacervationand liposome encapsulation. In recent years, microencapsulationhas increasingly become an important technology for the deliveryof numerous nutraceuticals and avour ingredients into food ma-trix. Table 1 summarises omega-3 concentration and delivery tech-nologies applied to food and beverage applications.

Spray drying is a widely used microencapsulation technique inthe food industry (Drusch, Benedetti, Scampicchio, & Mannino,2008; Jafari, Assadpoor,He,&Bhandari, 2008). It involves conversionof liquid oils andavours, in the formof emulsions, into dry powdersusing proteins and/or carbohydrate as the matrix materials. How-ever, this technology has limitations that the oil loading level islow, the level of surface (extractable) oil is high, and air inclusionin the emulsication process is difcult to avoid leading to particleinationduring the spray drying stage (Keogh et al., 2001). A leadingcommercialised spray-dried emulsion product was developed by agroup at CSIRO in Australia. Additional stabilisation and oil loadingwas achieved through the use of a casein-based Maillard reactionproduct (Kosaraju, Weerakkody, & Augustin, 2009).

Complex coacervation technology was rst developed usinggelatin and gum arabic. This technology was used to make carbon-less paper for the printing industry several decades ago. At ONC wehave developed a novel variation of this technology that involvesthe addition of a controlled agglomeration step and the formationof an outer shell that surrounds the agglomerations. This technol-

istry 131 (2012) 639644ogy has been scaled to manufacturing and is widely used commer-cially to stabilise and delivery omega-3 oils into foods andbeverages (Barrow, Nolan, & Jin, 2007). The key features of the

powder products from this technology are its high oil loading level,normally at least 60%, and its low level of surface oil, normally lessthan 0.1%. These levels compare favourably with spray dried pow-der products which have an oil loading level of typically less than30% and a surface oil level between 0.2% and 1%. A high loading le-

sensory prole for all applications. Alternative shell materials areneeded in some food and beverage applications. In recent years,a number of studies have been reported on complex coacervationsystems using non-gelatin material such as globular proteins withanionic polysaccharides, for example, b-lactoglobulin, bovineserum albumin, egg albumin, soy proteins, pea proteins and wheyproteins with gum Arabic, carrageenan and pectin (De Kruif,Weinbreck, & De Vries, 2004).

We have developed novel whey protein-gum arabic sh oil sys-tems as a viable option for the commercial production of non-gel-atin vegetarian microcapsules (Yan, Zhang, Jin, & Barrow, 2008;Zhang, Yan, May, & Barrow, 2009). These microcapsules were pre-pared using a combination of complex coacervation and thermalcrosslinking processes, including spray drying to form a free-ow-ing powder. Chemical or enzymatic crosslinking of whey proteinmicrocapsules was not required, as it is for gelatin. Instead, a ther-mal crosslinking process was developed to solidify the microcap-sules and prevent their dissolution, essentially by coagulation ofthe whey protein. Three types of the microcapsules were producedwith different processing sequences resulting in powder productswith distinguishable features. An example of the cross-section ofone type of these microcapsules is shown in Fig. 3. We found thatprocessing parameters determined the nal characteristics of thethree types of microcapsules, such as morphology, surface proper-ties and particle integrity, as well as their performances in sensory

Fig. 3. SEM pictures of the cross-section of a whey gum microcapsule with amagnication of X2000.

erag

J.A. Kralovec et al. / Food Chemistry 131 (2012) 639644 643vel means that less shell material is needed to deliver EPA and DHAso that cost and sensory impact on food are reduced to a minimallevel. The surface oil is essentially unprotected oil and is vulnerableto rapid oxidation and sensory deterioration. A low level of surfaceoil is critical to maintaining the sensory properties of the productduring both powder storage and addition and storage of the forti-ed food products.

A major disadvantage of complex coacervation technology isthe limitation in the selection of the shell materials. Gelatin hasbeen the material of the choice for use in complex coacervationdue to its unique gelation properties. However, gelatin has somelimitations, including it not being vegetarian and kosher versionsbeing relatively expensive. Also, gelatin does not have an ideal

Table 1Some current technologies for omega-3 concentration and delivery into food and bevEnzymaticprocessingtechnology

Purpose Lipase used P

EE or FFA to TG conversion Candida Antarctica lipase B(CALB)

F

EE or FFA to TG conversion CALB FEPA, DPA and DHA enrichedstructured lipids.

Various SD

Partial Concentration of EPAand DHA.

Thermomyceteslanuginosus lipase

UD

Separation of EPA and DHA Rhizomucor miehei lipase(RML)

E

Delivery technology Shell materials Loading of EPA/DHA (mg)per gram of powder

P

Spray-driedemulsion

Caseinate, dextrose, glucosesyrup

69143, Tuna oil 4

Spray-driedemulsion

Cornstarch, gelatin, sucrose 90, Algal oil 1m

Spray-driedemulsion

Carbohydrate, protein,antioxidant

170195, Sardine oil

Spray-driedemulsion

Modied starch, soy protein 100, Menhaden oil 1

Gravity ow dryblending

Maltodextrin 65155, Sardine oil >

Complexcoacervation

Gelatin, polyphosphate 140180, Sardine oil 6evaluations. Sensory stability was similar for each of the threetypes of microcapsules, and was determined using an in-housesensory panel evaluation over a 10 month period.

It is also worth noting that these whey protein and gum arabicmicrocapsules performed extremely well in a shelf life study in 2%ultra high UHT milk, as compared to gelatin microcapsules. UHTmilk treatment involves an ultra-high temperature process.Though it only lasts a few seconds, this type of heat can be verydestructive to certain types of microcapsules. The fact that thewhey protein and gum arabic microcapsules survived this severetreatment indicates an ability to resist the negative impact of heaton the structure and the quality of the microcapsules themselves.It suggests that since these microcapsules had undergone a

e applications.

roduct information Oil type Producer

ood grade concentrate Sardine/Anchovy Ocean NutritionCanada

ood grade concentrate Sardine/Anchovy Pronovatructured Lipids. 3060% EPA,PA or DHA

High-laurate canolaoil

Hamam and Shahidi(2006)

p to 50% increase in EPA andHA levels.

Sardine/Anchovyand Tuna oils.

Ocean NutritionCanada

PA/DHA ratio of 22:1 Sardine/Anchovy oil. Fernandez-Lorenteet al. (2011)

article size Shelf life Producer

585190 lm 612 months Clover/Nu-Mega

00% through 20 mesh, >85% 40esh,

thermal crosslink process when they were manufactured, someadditional heat treatment can enhance rather than decrease theirstructural stability, instead of thermally damaging the shell mate-rial. Also, we anticipate that the good performance of these new

ongoing and future research and development in the area of ome-

Gibbs, B. F., Kermasha, S., Alli, I., & Mulligan, C. N. (1999). Encapsulation in the foodindustry: A review. International Journal of Food Science and Nutrition, 50,

Grenfell-Lee, D. G. (2009). A fermentation-based process for carotenoid synthesis inYarrowia lipolytica. In SIM Annual Meeting Proceedings. Industrial Microbiologyand Biotechnology (p. 74), Toronto, Canada.

Hamam, F., & Shahidi, F. (2006). Synthesis of structured lipids containing medium-

644 J.A. Kralovec et al. / Food Chemistry 131 (2012) 639644213224.Gouin, S. (2004). Microencapsulation: Industrial appraisal of existing technologies

and trends. Trends in Food Science & Technology, 15, 330347.ga-3 oils.

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5. Future directions

Lipases are successfully used industrially for omega-3 partialconcentration and re-esterication. Improving the EPA and DHAselectivity of lipases would broaden the range of concentrates thatcould be made using enzymatic processing. A major future target isthe development of EPA and DHA selective lipases. Low cost novelmicrobial sources of DHA and EPA is another major research targetwithin the omega-3 eld. We and others are focused on the meta-bolic engineering of Thraustochytrid strains used, to manipulatethe concentration and ratios of EPA and DHA for food, supplementand pharmaceutical applications.

Liquid and spray-dried emulsions are being used successfully insome food and beverage applications for the delivery of omega-3oils. Complex coacervation has also been shown to be a commer-cially useful method. Most future work is focused on improvingoil stability to enable incorporation into a wider range of foodproducts. Also, decreasing the cost of microencapsulation technol-ogies is important to enable commercial success in low cost perserving food products. We believe that novel and mild processingtechnologies, new microencapsulation technologies, and alterna-tive sources of EPA and DHA to sh oil are all important areas ofchain and omega-3 fatty acids. Journal of Agricultural & Food Chemistry, 54,43904396.

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A review of the progress in enzymatic concentration and microencapsulation of omega-3 rich oil from fish and microbial sources1 Introduction2 The production of omega-3 concentrates from fish oil3 Microbial sources of omega-3 concentrates4 Stabilisation and delivery of omega-3 fatty acid rich oils into foods5 Future directionsReferences