COMPARATIVE STUDY ON THE POSSIBILITIES OF INCORPORATING OLIVE OIL AND NATURAL FENNEL EXTRACT IN FERMENTED MILKS

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1 319 Bulgrin Journl of Agriculturl Science, 23 (No 2) 2017, Agriculturl Acdemy OMPARATIV STUD ON TH POSSIBILITIS OF INORPORATING OLIV OIL AND NATURAL FNNL XTRAT IN FRMNTD MILKS MIHALA IVANOVA * ; GORGI KOSTOV; TATNA BALABANOVA; RADKA VLASVA; GALINA UZU- NOVA; MAGALI POIRIUX University of Food Technologies, BG-4002 Plovdiv, Bulgri Astrct Ivnov, M., G. Kostov, T. Blnov, R. Vlsev, G. Uzunov nd M. Poirieux, omprtive study on the possiilities of incorporting olive oil nd nturl fennel extrct in fermented milks. Bulg. J. Agric. Sci., 23 (2): It is well-known tht olive oil consumption is ssocited with numer of helth enefits to humns due to its ftty cid composition. Becuse of these chrcteristics, olive oil cn e suitly included in diry products in order to improve their ftty cid composition. The present study investigted the possiilities of incorporting olive oil in the mnufcture of fermented milk with set cogulum, directly in the form of n emulsion or encpsulted (with nd without the ddition of n ntioxidnt). A significntly higher content (p < 0.05) of unsturted ftty cids nd lower peroxide vlue of extrcted ft ws estlished in the smples supplemented with 2 g.10 2 g 1 encpsulted oil nd n ntioxidnt in n mount of 0.03 cm dm 3. The sensory profile of the fermented milks contining encpsulted olive oil ws etter in comprison with the fermented milks with directly dded olive oil. At the end of the shelf-life, the chemicl indictors ctive nd titrtle cidity, nd the microiologicl chrcteristics lctocilli nd streptococci counts remined comprle to those of the control. Key words: functionl diry products; yoghurt; unsturted ftty cids; olive oil; encpsultion Introduction Nturlly-occurring fts re essentil to humn nutrition. They perform numer of functions: energy supplying, constructive nd regultive, which hve specific physiologicl significnce to the humn ody (Joshi, 2010). The high clorie fctor of fts encourge consumers to choose products with low ft content nd trditionl products with improved ftty cid profile, contining omeg ftty cids. A numer of clinicl trils hve een crried out tht prove the enefits of replcing milk ft with vegetle oils rich in omeg ftty cids (Dwczynski et l., 2009, 2010). Vegetle oils re source of essentil polyunsturted ftty cids (Hunter, 1989). Linoleic nd lph-linolenic cids cnnot e synthesized in niml cells (Belitz et l., 2009), nd re therefore identified s n essentil ingredient for the humn ody. It hs een shown tht the consumption of oils with high oleic cid content fvourly reduces LDL cholesterol nd lood pressure (Lopez-Huertz, 2010). Furthermore, s source of essentil ftty cids, vegetle oils provide the humn ody with oil-solule vitmins, tocopherols, phytosterols, lecithin, pigments nd other unsponifile components present in trce mounts (Gunstone, 2004). Olive oil contins lrge mounts of oleic cid ( 18:1 ) (50 83 g.10 2 g 1 extrcted ft) nd polyunsturted ftty cids (7 16 g.10 2 g 1 extrcted ft). The fvourle unsturted to sturted ftty cid rtio nd the presence of ll vitmins nd out 100 trce elements needed y humns increse the nutritionl nd iologicl vlue of olive oil (Kiritskis et l., 1998). Lipid oxidtion of fts found in foods is technologicl prolem tht hs significnt effect on the shelf-life nd orgnoleptic properties of products (Min nd Lee, 1999). *orresponding uthor: mihel_18g@yhoo.com

2 320 Mihel Ivnov; Georgi Kostov; Ttyn Blnov; Rdk Vlsev; Glin Uzunov; Mglie Poirieux Given this fct, possiilities re incresingly sought to limit lipid oxidtion in products. ncpsultion of essentil oils is one possiility of limiting the contct of oil with oxygen, while the ddition of nturl ntioxidnt further limits the oxidtion processes (Bermúdez-Aguirre nd Bros- ánovs, 2011). The im of this study ws to crry out comprtive chrcteristic etween the possiilities of incorporting olive oil in the form of emulsion or in encpsulted form during the mnufcture of fermented milks, nd the influence of the dded nturl ntioxidnt fennel extrct. Mterils nd Methods Fermented milk ws prepred with rw whole nd skimmed cow s milk in complince with the requirements of Regultion 853/2004 of the uropen ommission, nd crem mde from rw cow s milk under Regultion 853/2004, in the mounts given in Tle 1. Strter composition for Bulgrin yoghurt: Lctocillus delrueckii susp. ulgricus nd Streptococcus thermophilus, exceeding g -1 in n mount of 20 cm 3.dm -3, mnufctured y Lctin LTD. The emulsion ws prepred with glycerol monosterte s emulsifier in n mount of 1 g.dm -3 ccording to Hsenhuettl nd Hrtel (2008) nd commercilly ville olive oil in n mount given in Tle 1. Tle 1 xperimentl vrints of different yoghurt smples *Smple Ft content of product, Olive оil mount, Milk ft mount, Antioxidnt mount, g.10 2 g 1 g.10 2 g 1 g.10 2 g 1 cm dm ЕA * control smple (yoghurt); control smple (yoghurt) with dded ntioxidnt (fennel extrct); yoghurt with directly dded olive oil; A yoghurt with directly dded olive oil nd ntioxidnt (fennel extrct); yoghurt with encpsulted olive oil; yoghurt with encpsulted olive oil nd ntioxidnt (fennel extrct) The lginte eds were prepred with: medium viscosity sodium lginte supplied y Sigm-Aldrich. For the purposes of the experiment, solution with concentrtion of 1.37 g.10-2 cm -3 (Kostov et l., 2016) ws prepred. The lginte ws soked in pproprite quntity of distilled wter for 24 h until complete dissolution, nd then ws sterilized in microwve oven (800 W for 5 min) to ensure microiologicl stility of the solution. Olive oil in concentrtion of 30 cm cm 3 ws dded to the finished lginte solution nd the emulsion ws homogenized t rpm. The homogenized emulsion ws dded dropwise to 2 g.10 2 cm 3 l 2 solution using peristltic pump. Afterwrds, the gel eds remined for further 30 min in the gelling solution to chieve the required mechnicl stility. The resulting cpsules were wshed with distilled wter nd left for hours to dry under refrigertion conditions nd were then dded to the product. The ntioxidnt ws fennel extrct used in n mount of 0.03 cm dm 3 with known chrcteristics (Nenov et l., 2013). The experimentl vrints re given in Tle 1. Preprtion of fermented milk The experimentl smples were otined y the clssicl method for production of fermented milk with set cogulum (Tmime nd Roinson, 2007). The ntioxidnt in the control smple ws dded during normliztion of milk. The direct ddition of olive oil nd the ddition of olive oil with n ntioxidnt to the milk were performed prior to psteuriztion in the form of emulsion. mulsions were prepred y mixing the oil phse in the dispersion medium under constnt stirring (speed rpm) for 5 min using IKA T18 digitl Ultr-Turrx lortory homogenizer t until the emulsifier ws completely dissolved (Vlsev et l., 2014). The ddition of olive oil cpsules nd olive oil nd ntioxidnt cpsules to the milk ws performed during inocultion. Titrtle cidity The cidity of the smples ws determined y Thorner s method (BNS ), nd presented s percentge of lctic cid. Active cidity ph of the smples ws mesured y ph meter (model MS 2000, Mycrosist, Plovdiv, Bulgri) with glss electrode (Sensorex, Grden Grove, USA) stndrdized t 20 over the rnge of Active cidity (рн), potentiometriclly mesured with 7110 WTW (Germny) ph-meter. Microiologicl nlysis Totl lctocilli nd streptococci count smple preprtion ws conducted ccording to IDF Stndrd 122:1996. The suitle dilutions were inoculted into selective grs М17 nd MRS, s descried in IDF Stndrd 117B:1997.

3 omprtive Study on the Possiilities of Incorporting Olive Oil nd Nturl Fennel xtrct Peroxide vlue Ft extrction for determintion of the peroxide vlue ws performed y the method of Bligh nd Deyer, The iodometric method sed on the interction of ctive peroxides or hydroperoxides with oxygen in the presence of iodide cetic cid (ISO, N 3960:2007) ws used. Ftty cid composition The preprtion of methyl esters of ftty cids ws in ccordnce with ISO 5509:2000. The nlysis of the methyl esters of the ftty cids ws crried out y gs chromtogrphy (ISO 5508:2004). Ftty cid composition of the otined yoghurts ws determined fter extrction of ft y the method of Schmidt-Bonzynsky-Rtzloff (ISO 1735 IDF 5:2004). Orgnoleptic ssessment ws conducted ccording to BNS The evlution criteri included the following indictors: flvour nd rom 35 points, density nd frcture of cogulum 30 points, texture 25 points, ppernce 10 points (mximum overll score 100 points). The results were equted to 10-point evlution scle with view to eing presented in spider digrm. Sttisticl nlysis omputer processing of the results ws performed using the progrm Microsoft xcel 2010 (ANOVA). Multiple comprisons were mde y the LSD method. The results re presented s men vlue ± SD (n = 4). Results nd Discussion hemicl indictors of fermented milks The chnges in the indictors ph nd lctic cid mount of the experimentl smples, were monitored nd determined over the twelve-dy storge t 4 (Figure 1 A, B). The results show tht on the first dy of storge the mount of lctic cid in smples,, nd A ws in the rnge of pproximtely 0.80 g.10 2 g 1, while the ctive cidity vlues were in the rnge etween It is noteworthy tht in the smples contining encpsulted oil (vrint ) nd encpsulted oil nd ntioxidnt (vrint ) the mount of lctic cid tht ws formed ws slightly lower, while the reported vlues for the ph indictor of the sme vrints were higher. This is proly due to the presence of other sustrtes (free croxyl groups of the lginte) which slow down lctose sorption. The oserved trend chnged during storge, when the vlues of lctic cid incresed in ll experimentl smples nd reched vlues of pproximtely 1.0 g.10 2 g 1 in the control, nd pproximtely 0.9 g.10 2 g 1 in vrints,, A, nd. The ctive cidity of these smples ws etween , respectively. The nutritionl nd iologicl vlue ws determined y the mount nd rtio of sturted to unsturted ftty cids g lctic cid.10-2 g -1 product A) ph B) contents, nd the rtio etween essentil nd non-essentil ftty cids. The ftty cid composition of the experimentl smples is given in Tle 2. The results revel tht the content of short-chin sturted ftty cids cproic, cprylic nd cpric is the highest in smples nd, followed y vrints nd, nd the lowest levels re in smples A nd. At the end of storge, the contents of plmitic, luric nd steric ftty cids ccount for the lrgest shre of the totl sturted ftty cids content in ll experimentl smples. The content of plmitic cid is higher in smples, nd in comprison with vrints A, nd, while luric cid is present in greter mount in, nd compred to smples A, nd. In the group of unsturted ftty cids, the oleic cid content is the highest nd is pproximtely twice higher in sm c dy 1 dy 12 dy 1 dy 12 Fig. 1. hnges in cidity (A) nd ph (B) of different yoghurt smples during 12 dys of storge control smple (yoghurt); control smple (yoghurt) with dded ntioxidnt (fennel extrct); yoghurt with directly dded olive oil; A yoghurt with directly dded olive oil nd ntioxidnt (fennel extrct); yoghurt with encpsulted olive oil; yoghurt with encpsulted olive oil nd ntioxidnt (fennel extrct). Different letters in the sme row show sttisticlly significnt differences (p < 0.05) during storge A A

4 322 Mihel Ivnov; Georgi Kostov; Ttyn Blnov; Rdk Vlsev; Glin Uzunov; Mglie Poirieux Tle 2 Ftty cids profile of different yoghurt smples fter 12 dys of storge Ftty cid * A Sturted ftty cids, g.10-2 g -1 extrcted ft proic 6:0 2.12± ± ± ± ± ±0.14 prylic 8:0 3.41± ± ± ±0.23 c 1.85±0.22 c 1.58±0.24 c pric 10:0 0.35± ± ± ± ± ±0.01 Luric 12: ± ± ±0.22 c 6.10±0.36 d 5.72±0.25 d 4.83±0.24 e Tridecylic 13:0 1.92± ± ± ± ± ±0.29 Myristic 14:0 1.45± ± ± ± ± ±0.12 Plmitic 16: ± ± ± ± ± ±1.95 c Mrgric 17:0 0.77± ± ± ± ± ±0.05 Steric 18:0 7.35± ± ± ±0.45 c 4.76±0.35 c 3.60±0.36 d Unsturted ftty cids, g.10-2 g -1 extrcted ft Myristoleic 14:1 0.32± ± ± ± ± ±0.09 Plmitoleic 16:1 3.55± ± ± ± ± ±0.32 Heptdecenoic 17:1 0.34± ± ± ± ± ±0.02 Oleic 18: ± ± ± ±1.34 c 49.27±1.45 d 48.21±1.24 d ssentil ftty cids, g.10-2 g -1 extrcted ft Linoleic 18:2 5.52± ± ± ±1.10 c 10.62±1.43 c 15.30±1.10 d control smple (yoghurt); control smple (yoghurt) with dded ntioxidnt (fennel extrct); yoghurt with directly dded olive oil; A yoghurt with directly dded olive oil nd ntioxidnt (fennel extrct); yoghurt with encpsulted olive oil; yoghurt with encpsulted olive oil nd ntioxidnt (fennel extrct); Different letters in the sme row show sttisticlly significnt differences (p<0.05) etween smples ples nd (out 50 g.10 2 g 1 extrcted ft) compred to vrints nd (pproximtely 25 g.10 2 g 1 extrcted ft). The totl unsturted ftty cids content is 29.78, 28.72, 41.6, 47.98, nd g.10 2 g 1 extrcted ft for vrints,,, A, nd, respectively. The results otined show tht the fermented milk products enriched with olive oil re chrcterized y higher content of unsturted ftty cids compred to the control smple. The highest proportion of polyunsturted ftty cids ws estlished in the smples contining microencpsulted oil in comprison to the control smple nd the smples with emulsified vegetle oil. Polyunsturted long-chin ftty cids omeg-3 nd omeg-6 ftty cids re importnt to humn nutrition. The experimentl smples of fermented milks re chrcterized y significntly high content of essentil ftty cids (p<0.05). The dt in Tle 2 indicte tht the mount of linolenic ftty cid is etween 13 nd 15 g.10 2 g 1 extrcted ft in smples A nd, 8 nd 11 g.10 2 g 1 extrcted ft in smples nd, nd pproximtely 7 g.10 2 g 1 extrcted ft in the control smple with nd without dded ntioxidnt. Determintion of the peroxide vlue indictor llows the determintion of the degree of ft oxidtion in the experimentl smples. Figure 2 shows the results otined for the peroxide vlue indictor for smples, A,, nd determined in over the period of storge t 4±1 for 12 dys. The results presented in Figure 2 show tht there is sttisticlly significnt difference etween the vlues of the peroxide vlue indictor otined on the first nd lst dy of storge. The higher oxidtion stte estlished in smples nd A compred with vrints nd is due to the fct tht encpsultion restricts the contct of Fig. 2. Peroxide vlue chnges in different yoghurt smples during 12 dys of storge yoghurt with directly dded olive oil; A yoghurt with directly dded olive oil nd ntioxidnt (fennel extrct); yoghurt with encpsulted olive oil; yoghurt with encpsulted olive oil nd ntioxidnt (fennel extrct)

5 omprtive Study on the Possiilities of Incorporting Olive Oil nd Nturl Fennel xtrct the oil with oxygen nd the oxidtion process. The dt otined re consistent with the results otined y Tmjidi et l., The results on the chnge in lctic cid cteri count lctocilli nd streptococci in the tested smples re presented in Figure 3. The dt of Figure 3 indicte higher concentrtion of lctic cid cteri in smples nd on the first dy of storge s compred with the other smples. The more intensive microiologicl nd iochemicl processes tht occur during lctose fermenttion in smples,, nd A correlte with Dy 1 the estlished higher levels of the indictors lctic cid nd ph (Figure 1 A, B). The longer exponentil phse estlished in the development of streptococci in smples nd could e explined y the medium rte of milk cidifiction (Figure 1 A, B). A storge temperture elow 5 is unfvourle for the Smple A 0 5 A) log cfu.g -1 Lctocillus delrueckii susp. ulgricus Streptococcus thermophilus Dy 12 Smple A 0 5 log cfu.g -1 B) Lctocillus delrueckii susp. ulgricus Streptococcus thermophilus Fig. 3. hnges in Streptococcus thermophilus nd Lctocillus delrueckii susp. ulgricus numer of different yoghurt smples t 1 st (A) nd 12 th (B) dys of storge Fig. 4. Sensory profile of different yoghurt smples fter 12 dys of storge control smple (yoghurt); control smple (yoghurt) with dded ntioxidnt (fennel extrct); yoghurt with directly dded olive oil; A yoghurt with directly dded olive oil nd ntioxidnt (fennel extrct); yoghurt with encpsulted olive oil; yoghurt with encpsulted olive oil nd ntioxidnt (fennel extrct)

6 324 Mihel Ivnov; Georgi Kostov; Ttyn Blnov; Rdk Vlsev; Glin Uzunov; Mglie Poirieux development of lctic cid cteri. Under low storge temperture conditions, lctocilli nd streptococci counts of fermented milks decrese. The estlished reduction is less pronounced in lctocilli compred with streptococci, which indictes their lower sensitivity to the models developed here. The results of the orgnoleptic evlution of the studied smples, stored for 12 dys t 4 re presented in Figure 4. The results otined show tht smples nd of fermented milks re chrcterized y higher orgnoleptic evlution in comprison with vrints, A, nd. The vrints with encpsulted olive oil nd were wrded higher score y the tsting pnel thn the vrints with emulsified oil nd A. onclusion The enrichment of trditionl diry products with unsturted ftty cids is n innovtive pproch to providing helth ingredients eneficil to humns. The use of microencpsulted vegetle oils nd nturl ntioxidnts ensures incresed stility of the unsturted ftty cids, prolongtion of the product shelf-life nd preservtion of the orgnoleptic properties of the product. Acknowledgments This study ws crried out with the finncil support of the Scientific Investigtion Section Project 7/15-H, fund Science, University of Food Technologies, Plovdiv, Bulgri. References Belitz, H., W. Grosh nd P. Schieerle, Food hemistry. Springer, Berlin. Bermúdez-Aguirre, D. nd G. Bros-ánovs, Qulity of selected cheeses fortified with vegetle nd niml sources of omeg-3. Food Science nd Technology, 44: Bligh,. nd W. Dyer, A rpid method of totl lipid extrction nd purifiction. ndin Journl of Biochemistry nd Physiology, 37: Bulgrin ntionl stndrd (BNS) , Milk nd milk products. Determintion of cidity (Bg). Bulgrin ntionl stndrd (BNS) , Milk nd milk products. Оrgnoleptic nlysis (Bg). Dwczynski,., L. Mrtin, A. Wgner nd G. Jhreis, n-3 L-PUFA-enriched diry products re le to reduce crdiovsculr risk fctors: doule-lind, cross-over study. linicl Nutrition, 29 (5): Dwczynski,., R. Schuert, G. Hein, A. Mueller, T. idner, H. Vogelsng, S. Bsu nd G. Jhreis, Long-term moderte intervention with n-3 long-chin PUFA-supplemented diry products: effects on pthophysiologicl iomrkers in ptients with rheumtoid rthritis. British Journl of Nutrition, 101 (10): Gunstone, F., The hemistry of Oils nd Fts. Wiley-Blckwell, pp Hsenhuettl, G. nd R. Hrtel, Food mulsifiers nd their Applictions. Springer, Berlin, pp Hunter, J., Scientists discuss dietry ftty cids. Journl of Americn hemicl Society, 66: IDF Stndrt 117B:1997, Milk nd Milk Products-ogurt numertion of chrcteristic microorgnisms olony-count technique t 37 degrees (Lctocillus delrueckii susp. ulgricus nd Streptococcus thermophilus) re present nd vile. Interntionl Diry Federtion, Brussels, Belgium. IDF Stndrt 122:1996, Milk nd Milk Products Preprtions of Smples nd Dilutions for Microiologicl xmintion, IDF 122:1996. Interntionl Diry Federtion, Brussels, Belgium. ISO 1735 IDF 5:2004, Determintion of ft content in crem cheese Grvimetry (Schmid-Bondzynski-Rtzlff). ISO 5508:2004, Animl nd vegetle fts nd oils Anlysis y gs chromtogrphy of methyl esters of ftty cids. ISO 5509:2000, Animl nd vegetle fts nd oils Preprtion of methyl esters of ftty cids. ISO N 3960:2008, Animl nd vegetle fts nd oils. Determintion of peroxide vlue. Iodometric (visul) end point determintion. Joshi, S., Nutrition & Dietetics. Tt McGrw-Hill duction, New Delhi, pp Kiritskis, P., A. Kiritskis,. Lenrt, W. Willet nd R. Hernndez, Olive Oil, From the Tree to the Tle, Second dition. Wiley-Blckwell. Poirieux, M., G. Kostov, R. Denkov, V. Shopsk, M. Ivnov, T. B. R. Vlsev, Otining cpsules with high oil phse content for ppliction in diry industry, rohydrtes Polymer (in press). Lopez-Huerts,., Helth effects of oleic cid nd long chin omeg-3 ftty cids (PA nd DHA), enriched milks, A review of intervention studies. Phrmcologicl Reserch, 61: Min, D. nd H. Lee, hemistry of lipid oxidtion. In: R. Ternishi, F. Wick nd I. Hornstein (ds.) Flvor hemistry: Thirty ers of Progress. Kluwer Plenum Pulishers, New ork, pp Nenov, N., T. Atnsov, I. Stoilov, V. Gochev, T. Girov nd A. Stoynov, Low temperture extrction of essentil oil ering plnts y liquificte gses. 11. Dill (Anethum grveolens L.). Scientific works Food science, engineering nd technologies 2013, Octoer 2013, Plovdiv, LX: (Bg). Tmime, A. nd R. Roinson, oghurt Science nd Technology, 3 rd dition. R Press. Tmjidi, F., A. Nsirpour nd M. Shhedi, Physicochemicl nd sensory properties of yogurt enriched with microencpsulted fish oil. Interntionl Journl of Food Science & Technology. 18: Vlsev, R., M. Ivnov, N. Petkov, A. Stoynov nd M. Schreiner, Otining stle food emulsions etween milk nd corn oil. Bulgrin Journl of Agriculturl Science, 20 (5): Received Novemer, 25, 2016; ccepted for printing Jnury, 9, 2017