Influence of two inocula levels of Saccharomyces bayanus, BV 818 on fermentation and physico-chemical properties of fermented tomato (Lycopersicon …

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  African Journal of Biotechnology Vol. 11(33), pp. 8241-8249, 24 April, 2012Available online at http://www.academicjournals.org/AJBDOI: 10.5897/AJB11.4300ISSN 1684  – 5315 © 2012 Academic Journals Full Length Research Paper  Influence of two inocula levels of Saccharomyces bayanus, BV 818   on fermentation and physico-chemicalproperties of fermented tomato ( Lycopersicon esculentum Mill  .) juice John Owusu 1,2 , Haile Ma 1,5 *, Ernest Ekow Abano 1,3 and Felix Narku Engmann 1,4 1 School of Food and Biological Engineering, Jiangsu University, 301 Xuefu Road, Zhenjiang 212013, China. 2 Hospitality Department, School of Applied Science and Technology, Koforidua Polytechnic, Koforidua, Ghana. 3 Agricultural Engineering Department, University of Cape Coast, Cape Coast, Ghana. 4 Hotel Catering and Institutional Management Department, Kumasi Polytechnic, Kumasi, Ghana.   5 Key Laboratory for Physical Processing of Agricultural Products of Jiangsu Province, 301 Xuefu Road, Zhenjiang212013 Accepted 2 April, 2012 The influence of two inocula levels of the yeast Saccharomyces bayanus, BV 818, 0.01 (w/v) and 0.02%(w/v) on physico-chemical properties of fermented tomato juice was investigated. The propertiesstudied include alcoholic strength, dry extract, residual sugar, pH, titratable acidity and volatile acidity.During fermentation, the pH, titratable acidity, brix, CO 2 production and changes in phenoliccomposition of the fermented juice were monitored. Both pH and titratable acidity showed an upwardtrend for the fermented juices. Alcoholic strength of the fermented juice produced with 0.01% (w/v)inoculum level was significantly higher (p<0.05) than that produced with 0.02% (w/v). The totalphenolics and pH for wine obtained from 0.02% (w/v) inoculum level were significantly higher (p<0.05)than that from 0.01%. Volatile acidity values of both wines were below the permitted levels. Duringageing, most colour parameters showed higher values in both fermented juices. Inoculum level 0.01%(w/v) gave better physico-chemical qualities and was therefore found to be better than 0.02% (w/v) inproducing fermented tomato juice. The 0.01% (w/v) inoculum-fermented tomato juice scored higher foroverall acceptance than that of 0.02% (w/v) inoculum-fermented tomato juice, but the acceptance levelswere not significant.Key words: Tomato, inoculum level, wine, fermentation. INTRODUCTION Wine is usually made through fermentation of grape juice.However, juices of many fruits are reported to have beenused for making wine. This includes African bush mango(Akubor, 1996), guava (Anderson and Badrie, 2005;Sevda and Rodrigues, 2011), jamun fruit (Chowdhuryand Ray, 2007), litchi fruit (Singh and Kaur, 2009), amlafruit (Soni et al., 2009), kinnow fruit (Panesar et al., *Corresponding author. E-mail: mhl@ujs.edu.cn. Tel: +86-511-88790958. Fax: +86 511 88 78 0201. 2009), apples (Enidiok and Attah, 2010), papaya (Lee etal., 2010), raspberry (Duarte et al., 2010), and tomato2009), apples (Enidiok and Attah, 2010), papaya (Lee etal., 2010), raspberry (Duarte et al., 2010), and tomato(Mathapati et al., 2010). Many fruits and vegetables areknown to be good sources of vitamins, minerals, fibre andphytochemicals. The fermentation of juices of most ofthese fruits is likely to produce wines of varied nutritional,phytochemical and sensory qualities.The production of any alcoholic beverage involvesalcoholic fermentation, which may start spontaneouslyeither by wild yeast or by inoculation of must or juice with  8242 Afr. J. Biotechnol.yeast of known characteristics (Jackson, 2008). Thestrain of the yeast and the level of yeast inoculated areamong the most important factors in determining thequality of the alcoholic beverage produced. The inoculumlevel, which is the quantity of starter culture added tomust or juice to initiate fermentation, is known toinfluence the duration of lag phase, specific growth rate,biomass yield, and the quality of final product in com-mercial industrial fermentation (Sen and Swaminathan,2004). An inoculum level of 7.5% (v/v) is reported to bethe optimum in producing 11% (v/v) ethanol from kinnowfruit (Panesar et al., 2009). The use of inappropriate ino-culum levels may lead to problems such as competitionamong yeast cells leading to premature death and hencelarge quantities of residual sugar, etc. Therefore, it isimportant that the right level of inoculum is utilized infermentation to make maximum use of the availablefermentable sugar.Tomato is one of the most important crops of the world.It is the most prominent source of lycopene (von Elbe andSchwartz, 1996), the carotenoid known to be the mostpowerful antioxidant of all carotenoids (Di Mascio et al.,1989). In addition to lycopene, tomato is endowed withother types of antioxidants, including ascorbic acid,vitamin E, carotenoids and flavonoids (USDA, 2009).Epidemiological studies have suggested that con-sumption of tomato and tomato products is closely linkedwith lower incidence of cardiovascular disease, prostate,gastrointestinal, and epithelial cell cancer (Rao and Rao,2007). Tomato is a perishable commodity and has to beprocessed to extend its shelf life. The well known existingprocessed forms of tomato include juice, paste, puree,soup, ketchup, sauce and canned tomatoes(Motamedzadegan and Tabarestani, 2011). Winemakingcould help to broaden the processing avenues of tomato.Mathapati et al. (2010) have reported the production ofwine from tomato juice using 2% (v/v) Saccharomyces cerevisiae  3282. However, in the present study  Saccharomyces bayanus  was used, because it   is knownto conduct equally effective alcoholic fermentation(Jackson, 2008), produce little volatile acidity, and givemore aromatic alcohols and ethyl esters (Antonelli et al.,1999). The use of different levels of inocula in fermen-tation of tomato juice has not received any attention byany author. Against this background, this study was con-ducted to investigate the effect of two inocula levels (0.01(w/v) and 0.02% (w/v)) of S. bayanus  , BV 818 at 20±1°Con the physico-chemical properties of fermented tomato juice. MATERIALS AND METHODSPreparation of yeast culture The dry yeast used for tomato juice fermentation was S. bayanus  BV 818 (Angel Yeast Company Ltd, Hubei Province, China). Thiswas kept in a refrigerator at 5° C according to the manufacturer’s instructions. The media used to culture the yeast was yeastpeptone dextrose (YPD) (yeast extract  – 0.5% (w/v), peptone -1.0% (w/v), and glucose - 2% (w/v)). The pH of the culture mediawas adjusted to 5.0 using tartaric acid. The culture was put into a-250 Erlenmeyer flask, sterilized in an autoclave at 121°C for 20min, and allowed to cool. Dry yeast of 0.3 g was then suspended inthe 100 ml sterilized medium to obtain yeast concentration of 3 g/L.The suspension was heated to 40°C for 20 min to rehydrate theyeast (Kraus et al., 1981), then cooled to 25°C for adaptation(Jackson, 2008) before it was incubated in an incubator shaker(QYC 211 Incubator Shaker, Shanghai Test Equipment Co. Ltd.) at30°C for 24 h using a speed of 160 rpm. Preparation and inoculation of tomato must The tomato was washed thoroughly with tap water. It was thensterilized with 2% potassium metabisulphite (KSM), and rinsed withdistilled water. The tomatoes were cut into smaller pieces andblended using a Kenwood blender (Philips HR 2006, China).Ammonium phosphate and pectic enzyme concentrations of 0.5 g/Leach was added to the must. Table sugar of a concentration of 200g/L was added (Ribereau-Gayon et al., 2006) to raise the totalsoluble solids (TSS) from 4.9±0.1 to 15.5±0.2 °Brix. The pH of themixture was adjusted from 4.22±0.11 to 3.21±0.01 using equalamounts of tartaric and citric acids, thereby raising the titratableacidity (TA) from 2.8±0.1 to 4.9±0.1 g/L (citric acid). The must waspasteurized at 40°C in a water bath for 1 h. It was then allowed tocool to room temperature and inoculated with the 24-h yeast culture( S. bayanus  BV 818). Inocula levels Must of volume 0.9 L, which weighed 1.06 kg and had dry mattercontent of 5% was inoculated with 30 and 60 ml of the 24-h yeastculture of concentration 3.0 g/L. The must contained 855 ml ofwater and the resulting yeast concentrations were 0.1 (0.01% w/v)and 0.2 g/L (0.02% w/v) for the 30 and 60 ml inocula respectively.The wine produced with 0.01% w/v inoculum level was designatedas fermented tomato juice Lo (FTJ Lo) while the one from 0.02%w/v was designated as fermented tomato juice Hi (FTJ Hi). Fermentation of tomato must The mixture was incubated at 20±1°C for seven days. Duringfermentation, the pH, titratable acidity (TA), total soluble solids(TSS), weight loss of fermentor as a measure of the rate of carbondioxide (CO 2 ) production (El Haloui et al., 1988), total phenols andcolour parameters were monitored starting from the second day upto the seventh day. After fermentation, the wine was cold stabilizedat 5°C for 24 h and centrifuged at 5000 rpm for 10 min. It was thenfiltered and kept at 5°C until needed for analysis. Analysis of fermented tomato juice The method of Sadler and Murphy (2010) was used to determineTA and the results were expressed in g/L citric acid. The pH wasmeasured using a pH meter (PHS-2C Precision pH/mV meter,China) after calibration with solutions of pH 7 and 4, respectivelyaccording to the AOAC (1984). The acid taste index (ATI) wascalculated using the formula: ATI = TA (g/L) - pH (Plane et al.,1980). The TSS was determined with the Abbe Refractometer withtemperature compensation (WAY-2S, Germany) and the valuesexpressed in degree brix (°Brix). Alcoholic strength was measuredusing the spectrophotometric method after distillation of the alcoholas in Caputi et al. (1968). The residual sugar content was mixed     determined by the dinitrosalicyclic (DNS) method (Miller, 1972).Total phenolics were monitored during fermentation by spectralanalysis as described by Somers and Ziemelis (1985). Volatileacidity was determined by AOAC (1960). Dry extract wasdetermined by AOAC (1995) and the results expressed in g/L.Colour parameters were measured following the method of Glories(1984). Absorbance was measured at 420 (A 420 ), 520 (A 520 ) and620 nm (A 620 ) and the values used for calculating colour density,intensity, tint, and % yellow colour, % red colour and % blue colourare as follows:Colour density (CD) = A 420 + A 520 , colour intensity (CI) = A 420 + A 520  + A 620 , colour tint or hue (CT) = A 420  /A 520 , % yellow = 100 x(A 420  /CI), % red = 100 x (A 520  /CI), % blue =100 x ( A620  /CI).The absorbance at 280 and 420 nm is a measure of total phenols(Somers and Ziemelis, 1985) and browning index (Jackson, 2008),respectively. Ageing The wines produced were aged in bottles at 5±2°C for eightmonths. During this period the following parameters weremonitored: pH, TA, TSS, alcoholic strength, residual sugar, volatileacidity, dry extract, acid taste index, colour intensity, tint, density, %yellow, % red, % blue, absorbance at 280 and 420 nm. Sensory evaluation A 10-member semi-trained panel was used to assess the taste,colour, aroma, and the overall acceptance of the two fermentedtomato juices on a 5-point hedonic scale. On the scale: 1, dislikevery much; 2, dislike much; 3, neither like nor dislike; 4, like much;5, like very much. The mean scores of the panelists were analyzed. Statistical analysis The data was analyzed using one-way analysis of variance(ANOVA). The statistical package used for the analysis was SPSSStatistics 17.0. Differences between means were separated usingleast significance difference (LSD). RESULTS AND DISCUSSIONFermentation monitoring The results on fermentation monitoring are shown inFigures 1a, b, 2a, b and 3. In the present study, FTJ Hiachieved maximum CO 2 production (1.6 g/L/h) at about30 h while FTJ Lo had maximum CO 2 production of 1.4g/L/h at about 50 h after commencement of fermentation(Figure 1b). The maximum CO 2 production figuresrecorded in this study were within the range of thosereported for raspberry wine, produced from TSS of 16°Brix (Duarte et al., 2010). The CO 2 production assumeda constant value which was earlier for FTJ Hi than FTJLo. This is an indication that fermentation of FTJ Hi cameto completion earlier than FTJ Lo. A sharp drop in TSSfor both inocula levels was associated with a large CO 2  production (Figures 1a and b). This agrees with what wasreported for papaya juice fermentation in which bothOwusu et al. 8243culture of S. cerevisiae  and Williopsis saturnus  andmonoculture of S. cerevisiae  experienced a sharp fall inbrix with time (Lee et al., 2010).In the course of fermentation, both pH and TA showedupward trend (Figures 2a and b) for the two fermentedtomato juices. Generally, the increase in TA was slightlyhigher for FTJ Lo than FTJ Hi. Increase in TA withfermentation time has been reported for African bushmango wine (Akubor, 1996). The increase in TA withfermentation time may be due to the production of acidsby yeast during fermentation (Ribereau-Gayon et al.,   2006). Titratable acidity increase in the course of fermen-tation is desirable because it can help prevent spoilageby microorganisms. The pH of the tomato must used forproducing the fermented juices was ameliorated withcitric and tartaric acids, so the increase in pH may becaused by crystallization of tartrate (Jackson, 2008). Atthe end of fermentation on the seventh day, the pH ofFTJ Lo was lower than that of FTJ Hi (Figure 2b). Figure2b shows the variation of total phenolics, expressed inabsorbance units (AU) during fermentation. There was aninitial fall in total phenolics up to the second day, followedby gradual increase with sharp increase in pH withfermentation time. The total phenolics of FTJ Hi wereslightly higher than that of FTJ Lo from the first day offermentation to the last day. Increased phenolics contentwith fermentation time may be attributed to increaseddissolution of phenols in increasing ethanol concentrationfrom the tomato pomace (Ribereau-Gayon et al.,   2006).The results show that increasing pH may be acontributory factor in the extraction of phenolics duringfermentation.Colour density of the fermenting must fluctuated overthe fermentation period with increasing phenolic content(Figure 3). This may be due to changes in anthocyaninfrom one form to another with pH changes (Ribereau-Gayon et al.,   2006). Physico-chemical properties of fermented tomatojuice before and after ageing The physico-chemical properties   of the fermented tomato juices produced are shown in Table 1. FTJ Lo had asignificantly lower (p<0.05) pH than FTJ Hi, both beforeand after ageing. Since FTJ Hi was produced by using ahigher inoculum level than FTJ Lo, higher competitionand more yeast deaths, leading to low acid production inthe case of the former than the latter, may account for thepH variation observed (Ribereau-Gayon et al.,   2006). ThepH of the two FTJ Lo and FTJ Hi (Table 1) was around3.5 (Ribereau- Gayon et al.,   2006). The favourable pHrange for white wine is 3.1 to 3.4, and that of red wine is3.3 to 3.6 (Jackson, 2008). Enidiok and Attah (2010)reported pH value of 3.68 and 3.79, respectively for Syzygium malaccensis  and Eugenia owariensis  applewines. Ribereau-Gayon et al.   (2006) have observed thatlow pH values in wines enhance microbiological andphysicochemical stability. Before ageing, the TA of both  8244 Afr. J. Biotechnol. A B Figure 1. Variation in (a) TSS and (b) CO 2 production with fermentation time for the twofermented tomato juices. wines was almost the same, but after ageing, the valueswere the same (Table 1). In addition, the TA values forboth wines after ageing were significantly higher (p<0.05)than those before ageing. Increase in TA afterfermentation has been reported for jamun wine byChowdhury and Ray (2007). Also, an increase in acidityduring a six month ageing period had been reported forwines aged in bottles (Garde-Cerdan et al., 2008). TheTA figures recorded in the present study fall within therange 5.5 to 8.5 g/L in most wines (Jackson, 2008). Theacid taste index values of FTJ Lo was not significantlydifferent (p>0.05) from that of FTJ Hi, both before andafter ageing. However, each fermented tomato juicerecorded a significantly higher acid taste index valuesafter ageing than before ageing (Table 1). Dry red wineshave acid taste index values of two to three and dry whitewines have 2.7 to 3.7, and values too far below theselevels make wine flabby while those too above make thewine sharp and acidic (IIand et al., 2000). In the presentstudy, the acid taste index values before ageing fell withinthe reported figures but were higher after ageing (Table1). This gives an indication that the fermented tomato juices would taste a bit acidic after ageing than before.The TSS of tomato juice was 4.9±0.1 °Brix, but wasameliorated to 15.5±0.1 °Brix. The TSS of FTJ Lo andFTJ Hi were not significantly different (p>0.05) before andafter ageing. Even though the residual sugar valuesrecorded for FTJ Lo was lower than that of FTJ Hi, thedifference was not significant (p>0.05) both before andafter ageing. Both fermented tomato juices gave residualsugar levels lower than 2 g/L and this agrees with whatwas reported by Torija et al. (2003). Residual sugar  Owusu et al. 8245 A B Figure 2. a) Variation in TA, and b) total phenols (TP) and pH with fermentation time. affects the microbial stability of wines (Jackson, 2008).FTJ Lo recorded a significantly higher (p<0.05) alcoholicstrength value than FTJ Hi before ageing, and thisdifference may be attributed to a more efficient sugarconsumption in FTJ Lo than FTJ Hi. Much residual sugarwas left in FTJ Hi than FTJ Lo after fermentation, thoughthis difference was not significant. The alcoholic strengthvalues reported for icewines produced with 0.2 and 0.5 gactive dried wine yeast/L are 7.8 and 12.0%, respectively(Kontkanen et al., 2004). Alcoholic strength values of11.00±0.04 and 10.50±0.03 (%v/v) have been reportedfor S. malaccensis  and E. owariensis  apple wines(Enidiok and Attah, 2010). An inoculum level of 10% (v/v)is reported to produce 9% (v/v) ethanol from Amla (Emblica officinalis Gaertn. ) fruit wine (Soni et al., 2009).Mathapati et al. (2010) have reported a value of 7.88%alcoholic strength for tomato wine by using 2% (v/v) S.cerevisiae  inoculum level. In the fermentation of kinnowsera, cane and kinnow-cane juice involving three inoculalevels, 5, 7 and 9% (v/v), Pratima et al. (2006) reportedmaximum ethanol production for 5% (v/v) inoculum level.The optimum inoculum level for ethanol production inguava wine was reported as 8% (v/v) for S. cerevisiae   NCIM 3095 (Sevda and Rodrigues, 2011). After ageing,even though both fermented juices recorded significantreduction (p<0.05) in alcoholic strength, FTJ Hi had asignificantly higher value than FTJ Lo. Soni et al. (2009)reported a reduction in ethanol content during ageing foramla wine. The reduction in ethanol content duringageing may be due to oxidation of ethanol to acetal-
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