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Dziugan et al. Biotechnology for Biofuels 2013, 6:158http://www.biotechnologyforbiofuels.com/content/6/1/158

RESEARCH Open Access

Evaluation of the fermentation of high gravitythick sugar beet juice worts for efficientbioethanol productionPiotr Dziugan1, Maria Balcerek2*, Katarzyna Pielech-Przybylska2 and Piotr Patelski2

Abstract

Background: Sugar beet and intermediates of sugar beet processing are considered to be very attractive feedstockfor ethanol production due to their content of fermentable sugars. In particular, the processing of the intermediatesinto ethanol is considerably facilitated because it does not require pretreatment or enzymatic treatment in contrastto production from starch raw materials. Moreover, the advantage of thick juice is high solid substance andsaccharose content which eliminates problems with the storability of this feedstock.

Results: The objective of this study were to investigate bioethanol production from thick juice worts and theeffects of their concentration, the type of mineral supplement, as well as the dose of yeast inoculum onfermentation dynamics and ethanol yield.The obtained results show that to ensure efficient ethanolic fermentation of high gravity thick juice worts, oneneeds to use a yeast strain with high ethanol tolerance and a large amount of inoculum. The highest ethanol yield(94.9 2.8% of the theoretical yield) and sugars intake of 96.5 2.9% were obtained after the fermentation of wortwith an extract content of 250 g/kg supplemented with diammonium hydrogen phosphate (0.3 g/L of wort) andinoculated with 2 g of Ethanol Red dry yeast per L of wort. An increase in extract content in the fermentationmedium from 250 g/L to 280 g/kg resulted in decreased efficiency of the process. Also the distillates originatingfrom worts with an extract content of 250 g/kg were characterized by lower acetaldehyde concentration thanthose obtained from worts with an extract content of 280 g/kg.

Conclusions: Under the favorable conditions determined in our experiments, 38.9 1.2 L of 100% (v/v) ethylalcohol can be produced from 100 kg of thick juice. The obtained results show that the selection of processconditions and the yeast for the fermentation of worts with a higher sugar content can improve the economicperformance of the alcohol-distilling industry due to more efficient ethanol production, reduced consumption ofcooling water, and energy for ethanol distillation, as well as a decreased volume of fermentation stillage.

Keywords: Bioethanol, Fermentation, Thick juice, Sugar beet, High gravity wort, Yeast

BackgroundBiofuels are defined as solid (biochar), liquid (bioethanol,biobutanol, biodiesel) and gaseous (biogas, biosyngas,biohydrogen) fuels that are mainly derived from biomass.Traditionally, sugar substrates derived from food cropssuch as sugar cane, corn (maize) and sugar beet have beenthe preferred feedstock for the production of biofuels [1].

* Correspondence: [email protected] of Spirit and Yeast Technology, Institute of FermentationTechnology and Microbiology, Lodz University of Technology, 90-924Wolczanska, Lodz 171/173, PolandFull list of author information is available at the end of the article

2013 Dziugan et al.; licensee BioMed CentraCommons Attribution License (http://creativecreproduction in any medium, provided the or

Bioethanol can be produced from all feedstock thatcontain mono-, oligo- and polysaccharides (for example,starch and cellulose) [2]. An advantage of raw materialscontaining simple sugars and disaccharides, such as sac-charose, is the simplified technology of extraction to thewater medium, followed by fermentation to ethanol with-out the need of using additional technological operationsconnected with chemical or enzymatic hydrolysis, whichcould significantly increase the costs of biosynthesis [2].From an economic point of view and in comparison withcereals, sugar beet and beet-processing intermediates con-taining saccharose are very good raw materials for ethanol

l Ltd. This is an open access article distributed under the terms of the Creativeommons.org/licenses/by/2.0), which permits unrestricted use, distribution, andiginal work is properly cited.

mailto:[email protected]://creativecommons.org/licenses/by/2.0

Table 1 Chemical composition of raw material

Physicochemical parameters Thick juice

Solid substance (g/kg) 685.2 11.5

pH 7.4 0.2

Reducing sugars as invert sugar (g/kg) 3.1 0.4

Saccharose (g/kg) 598.4 12.5

Total nitrogen (g/kg) 5.6 0.4

Volatile acids as acetic acid (g/kg) 4.4 0.2

Results expressed as mean values standard error (n = 3).

Dziugan et al. Biotechnology for Biofuels 2013, 6:158 Page 2 of 10http://www.biotechnologyforbiofuels.com/content/6/1/158

production due to their content of fermentable sugars(saccharose) [3,4].The production of ethanol from sugar beet-processing

intermediates (raw, thin, and thick juices) and from bypro-ducts (molasses) constitutes an alternative solution forsugar factories interested in a combined production ofsugar and bioethanol. Furthermore, the use of inter-mediate products of sugar beet processing as raw mate-rials for bioethanol production could be attractive fordistilleries located near the sugar factories, as it wouldminimize high transportation costs. Cooperation betweenthese factories could lead to increased production andutilization of the capacity of both types of facilities.Very high gravity (VHG) processes are extremely at-

tractive and promising for bioethanol production as theyallow significant improvements in overall productivity,thus minimizing production costs thanks to energy sav-ings [5]. On the other hand, the use of VHG technologyimposes greater stress on yeast cells, which has beenassociated with the loss of yeast viability during VHGfermentation, a reduced rate, and incomplete fermenta-tion [6]. Thus, the successful implementation of VHGtechnology in bioethanol production requires the use ofyeast strains that can efficiently ferment high sugarconcentrations (>250 g/L) [7]. Such strains must be resist-ant to the multiple stresses found in the process, includingthe osmotic stress that results from high sugar concentra-tion, the ethanol stress at the end of fermentation, theanaerobic conditions established in large-scale bioreac-tors, and the cell recycling procedures for the utilizationof the yeast biomass for several consecutive fermenta-tion cycles [8,9].Balcerek at al. [10] investigated the effect of various

strains of the yeast Saccharomyces cerevisiae (S. cerevisiae)on the dynamics and efficiency of alcoholic fermentationof thick juice worts. The authors tested strains designatedas M1, M2, M3 (from the Pure Culture Collection of theInstitute of Fermentation Technology and Microbiology,Lodz University of Technology), commonly used for thefermentation of molasses worts, as well as strains desig-nated as Bc-16, D-2, As-4 (purchased from the yeastfactory in Maszewo Lborskie, Poland), used for the fer-mentation of mashes based on starch raw materials. Itwas found that S. cerevisiae strains M1 and M2 dynamic-ally and efficiently (89 to 94% of the theoretical yield)fermented thick juice worts with an extract concentra-tion of 200 g/kg and 250 g/kg, whereas the strain D-2preferred less dense worts (extract concentration of200 g/kg). Gumienna et al. [4] evaluated the efficiencyof alcoholic fermentation of sugar beet and its processingintermediates using commercial yeast strains such as Etha-nol Red and Fermiol (Fermentis Division S.I. Lesaffre,France). Balcerek and Pielech [11] also tested the EthanolRed yeast strain for the fermentation of triticale starch

mashes with a solid substance concentration of approxi-mately 23%. The obtained results showed high ethanolyields (87.54 0.46% to 88.30 0.46% of the theoreticalyield).According to the declaration of the producer (Fermentis

Division S.I.), Ethanol Red is a specially selected strainthat was developed for the ethanol industry. With a highethanol tolerance, this fast acting strain displays higheralcohol yields and maintains higher cell viability, especiallyduring VHG fermentation. Ethanol Red is particularlywell-suited for sugar substrates (sweet juices, molasses)and also saccharified mashes [12].The objective of the presented study was to determine

the effect of thick juice worts concentration, the type ofmineral supplements, and the dose of yeast inoculum onthe dynamics and efficiency of alcoholic fermentation.

Results and discussionChemical characteristics of thick juiceThe chemical composition of thick juice applied in thisstudy was typical of sugar beet processing intermediates(see Table 1). The high content of saccharose (598.4 g/kg)is advantageous from the technological point of viewbecause it promotes a high yield of ethanol from theraw material. Our results are consistent with the datareported by Rankovi et al. [13] with one exception re-lated to the total nitrogen content. The thick juice de-scribed by Rankovi et al. [13] contained four times lesstotal nitrogen (1.4 g/kg) than the raw material used in ourstudy (5.6 g/kg). The differences in the content of nitrogencompounds are probably related to the sugar beet varietiesprocessed in sugar factories in Poland and Serbia [14],and to different sugar beet cultivation conditions andthe technology used for processing it into thick juice(Table 1).The chemical composition of the investigated thick juice

makes this intermediate product of sugar beet processingan attractive feedstock for alcoholic fermentation. Thickjuice is only subjected to initial dilution, pH adjustment,and supplementation with inexpensive mineral sourcesof nitrogen for the yeast (if needed). This makes theoverall process of bioethanol production from thick sugarbeet juice relatively simple in comparison to production


from starch-based raw materials, which require the lib-eration of starch (by pressure cooking or by thoroughgrinding) and its liquefaction and saccharification [11].

The effect of process conditions on fermentationdynamics and the results of fermentation of high gravitythick juice wortsThe effects of initial extract content in thick juice worts,the type of mineral supplement, and the dose of yeastinoculum on the dynamics and efficiency of alcoholicfermentation were determined. The obtained results arepresented in Figures 1, 2, and 3.In all the processing variants, the highest conversion

of saccharose to fermentable sugars (expressed as reducingsugars) as a result of the activity of the -fructofuranosidase(EC 3.2.1.26) present in yeast cells, was observed duringthe first 12 h of fermentation. Due to the prolonged initialphase of fermentation of worts with an extract content of250 g/kg inoculated with a yeast dose of 1.0 g/L and supple-mented with diammonium hydrogen phosphate (0.3 g/L ofwort), ethanol production during the first 6 h of the process

Figure 1 Fermentation dynamics of thick juice worts with an extract(B) Inoculum content of 1.0 g/L; (NH4)2HPO4 + MgSO4 7 H2O. (C) Inoculum(NH4)2HPO4. MgSO4 7 H2O.

was very low (close to zero). In the next 6 h of the process,ethanol concentration increased to 0.8 0.02% (v/v)(Figure 1A). Intensive biosynthesis of ethanol was reportedafter 12 h of fermentation. After 94 h of fermentation,ethanol concentration in the wort reached 12.4 0.3%(v/v). On completion of the process, the real extractconcentration of wort decreased from 250 g/kg to ap-proximately 50 g/kg (by 80%). Also, the concentrationof residual reducing sugars was still sufficiently highand amounted to 35.4 g/L of wort. This indicates thatthe sugar substrates were not fully utilized (Figure 1A).Yeast cells have specific growth requirements leading

to an imbalance or limitations resulting in incompletefermentation. These requirements include specific levelsof nitrogen, carbon, vitamins, water, oxygen, and metalions. Metal ions are required for a number of purposes;they include bulk elements (such as magnesium, calcium,and potassium) and trace elements (such as zinc, copper,and manganese) needed by yeast cells [15]. Magnesium isnecessary for the activation of several glycolytic enzymes,and in practical terms this means that if industrial media

content of 250 g/kg. (A) Inoculum content of 1.0 g/L; (NH4)2HPO4.content of 1.5 g/L; (NH4)2HPO4. (D) Inoculum content of 2.0 g/L;

Figure 2 Fermentation dynamics of thick juice worts with an extract of 280 g/kg. (A) Inoculum content of 1.0 g/L; (NH4)2HPO4. (B) I noculumcontent of 1.0 g/L; (NH4)2HPO4 +MgSO4 7 H2O. (C) I noculum content of 1.5 g/L; (NH4)2HPO4. (D) Inoculum content of 2.0 g/L; (NH4)2HPO4.


are magnesium-limited, the conversion of sugar to alcoholmay be suppressed, leading to slow or incomplete fermen-tation processes [16].Due to the incomplete fermentation of the wort with

an initial extract content of 250 g/kg supplemented withonly diammonium hydrogen phosphate and inoculatedwith 1 g yeast per L of wort, it seemed appropriate toconduct further fermentation experiments using mixednutrients for yeast in the form of diammonium hydrogenphosphate (NH4)2HPO4 (0.3 g/L) and magnesium sulfateheptahydrate (MgSO4 7 H2O) (0.1 g/L) and largeramounts of yeast inoculum.Supplementation of thick juice wort (with an extract

content of 250 g/kg) with MgSO4 7 H2O (in addition todiammonium hydrogen phosphate) did not significantlyimprove the course of the process, so its efficiency wascomparable to the process conducted in the presence ofonly diammonium hydrogen phosphate. Upon comple-tion of this process, ethanol concentration in the wortsupplemented with Mg2+ ions reached 13.0 0.4% (v/v)and was not statistically higher than that in wort with-out the addition of MgSO4 7 H2O (12.4 0.2% v/v,0.05 < P

Figure 3 Fermentation results of thick juice worts. Different letters indicate significant differences (P


ethanol yield under the cited experimental conditions(Figure 3).The obtained results are in accordance with the findings

of Takeshige and Ouchi [17], who reported inhibited yeastgrowth and reduced ethanol yield in the process of molas-ses wort fermentation containing sugar at a concentrationof 300 g/kg. Moreover, Dodi et al. [18], who fermentedthick juice worts, observed that with an increase of fer-mentable sugars content from 5% to 20% (w/w), ethanolyields also increased for both investigated raw materials(molasses and thick juice). However, when an initial sugarscontent of 20% (w/w) was increased to 25% (w/w), theyields dropped significantly, from 67 to 56%. The lowyields obtained by Dodi et al. [18] could have beencaused by the fact that fermentation was carried outusing bakers yeast, which was most likely not adaptedto high-density worts. Furthermore, the worts were notsupplemented with mineral nutrients for yeasts.Hinkov and Bubnk [19], who fermented concentrated

raw sugar beet juice achieved the highest ethanol yield(88.2 to 94.4% of theoretical yield) when the sugar con-centration in the wort amounted to 200 g/kg. The effi-ciency of fermentation and ethanol yield decreased withan increase in wort extract. The distillery yeast strainstested by Hinkov and Bubnk [19] showed an increasedtolerance to osmotic pressure and provided higher yieldsin worts with higher initial concentrations of sugar. Athigh sugar concentrations, it was observed that the yeastexperienced osmotic pressure, which led to plasmolysisand a lower ethanol yield [20]. Based on the obtained fer-mentation coefficients for the studied thick juice worts(Figures 1, 2, 3), the quantity of 100% (v/v) ethanol ob-tained from 100 kg of this raw material was calculated.The results show that 38.9 1.2 L 100% (v/v) ethyl alco-

hol could be produced from 100 kg of thick juice underthe following favorable conditions, established in our ex-periments: extract content of 250 g/kg, yeast dose of 2 g/Lof wort and (NH4)2HPO4 addition of 0.3 g/L of wort.

Analysis of the chemical composition of the obtaineddistillatesThe quality of bioethanol used for fuel purposes is strictlydefined by the Polish national and industrial norms. Highconcentrations of fermentation by-products can cause alower price of the final product. According to some pro-ducers of dehydrated ethanol, higher concentrations ofpollutants in the raw spirit (unpurified ethyl alcohol)can cause fast deterioration of molecular sieves used inthe process of ethanol dehydration [21]. The chemicalcomposition of distillates obtained is shown in Table 2.Methanol concentration in the obtained raw spiritswas low and ranged from 7.7 0.7 to 9.3 0.9 mg/L100% (v/v) ethyl alcohol (no statistically significant dif-ferences, 0.05 < P

Table 2 Chemical composition of distillates obtained from the fermentation of thick juice worts

Compound(mg/L 100% v/vethyl alcohol)

Parameters of fermentation

Extract content of 250 g/kg Extract content of 280 g/kg

Inoculum contentof 1.0 g/L;(NH4)2HPO4

Inoculum contentof 1.0 g/L;(NH4)2HPO4 +MgSO4 7H2O



Inoculum contentof 1.0 g/L; (NH4)2HPO4

Inoculum contentof 1.0 g/L; (NH4)2HPO4 +MgSO4 7H2O



Methanol 8.1 0.8a 7.7 0.7a 9.3 0.9a 7.7 0.8a 9.3 0.9a 8.1 0.8a 9.3 0.9a 9.1 0.9a

Acetaldehyde 763.4 4.2a 1626.0 4.1b 2226.0 6.5c 2214.5 6.3c 3214.5 7.5d 3869.8 8.8e 3971.8 9.2f 4172.9 9.8g

Methyl acetate 8.2 0.8a 9.2 0.8a 8.6 0.8a 9.3 0.8a 7.9 0.8a 9.0 0.8a 8.3 0.8a 9.2 0.8a

Ethyl acetate 266.3 2.5b 248.7 2.5a 284.5 2.8d 270.8 2.6c 248.7 2.5a 246.8 2.5a 243.7 2.5a 285.0 2.8d

Isoamyl acetate 0.0a 0.0a 0.0a 1.9 0.2b 2.6 0.2c 4.2 0.3d 7.1 0.5e 8.7 0.5f

Ethyl butyrate 33.5 0.9c 45.3 1.4d 60.1 1.5f 52.1 1.4e 18.0 0.6a 18.6 0.6a 21.9 0.8b 22.6 0.8b

n-propanol 155.9 2.2bc 177.2 2.8e 187.1 2.8f 158.9 1.6c 152.9 1.6ab 150.3 1.5a 206.1 2.7g 165.9 1.5d

2-methyl-1-propanol 314.3 3.2b 307.2 2.8a 368.9 3.5d 381.1 3.5e 357.8 3.5c 402.2 3.8f 355.7 3.5c 355.3 3.5c

n-butanol 7.4 0.5a 7.8 0.5ab 7.5 0.5a 7.5 0.5a 6.8 0.5a 7.6 0.5a 8.9 0.7b 8.1 0.6b

2-methyl-1-butanol 236.3 2.5a 249.4 2.5b 299.7 2.9e 269.6 2.7c 272.1 2.7c 283.2 2.7d 284.5 2.7d 306.7 3.2f

3-methyl-1-butanol 650.7 3.5a 667.7 3.8b 805.8 4.1d 818.2 4.2e 719.4 3.8c 906.8 4.2g 969.0 4.3h 879.7 4.1f

Results expressed as mean values standard error (n = 3). a-hMean values in lines with different letters are significantly different (P


100% (v/v) ethyl alcohol while the content of 2-methyl-1-propanol was higher and ranged from 307.2 2.8 to402.2 3.8 mg/L 100% (v/v) ethyl alcohol. The amountsof n-butanol in all the tested distillates were relativelysmall (6.8 0.5 to 8.9 0.7 mg/L 100% v/v ethyl alcohol).The most abundant isoamyl alcohol detected in the dis-tillates was 3-methyl-1-butanol (650.7 3.5 to 969.0 4.3 mg/L 100% v/v ethyl alcohol), whereas the contentof 2-methyl-1-butanol ranged from 236.3 2.5 to 306.7 3.2 mg/L 100% (v/v) ethyl alcohol (Table 2). Apart fromthe significantly higher levels of acetaldehyde in the dis-tillates derived from worts with a density of 280 g/kg,there was no correlation between the concentrations ofthe identified byproducts and the fermentation condi-tions (Table 2).The literature provides scant reports on the chemical

composition of raw spirits originating from the inter-mediate products of sugar beet processing. Raw spiritsobtained from the fermentation of thick juice worts werecharacterized by a lower content of higher alcohols thanthose obtained by Balcerek and Pielech-Przybylska [11]following the fermentation of starch mashes (from triti-cale) with the Ethanol Red yeast strain.

ConclusionsThe results of our study prove that the intermediate prod-ucts of sugar beet processing, such as thick juice, may beconsidered an attractive raw material for bioethanol pro-duction. Saccharose is the principal component of its ex-tract, so the only necessary operations before alcoholicfermentation are dilution, pH regulation, and additionof mineral nitrogen sources (if needed). The fermentation ofthick juice worts with an extract content of 250 g/kg using2 g of the dry distillery yeast Ethanol Red (S. cerevisiae)per 1 L of wort supplemented with (NH4)2HPO4 as anutrient for yeast was determined to be favorable, as itenabled a high ethanol yield (38.9 1.2 L 100% v/v ethylalcohol from 100 kg of thick juice).Due to limitations on sugar manufacturing in EU coun-

tries, the capacity of sugar factories is not fully utilizedand they are ready to increase the processing of sugar beetinto intermediates, which could serve as feedstock forbioethanol factories. This would be an alternative to starchprocessing, especially in the years of crop failures. Anothercrucial issue is also the ability of biofuels to reduce green-house gas (GHG) emissions. GHG emissions in the lifecycle of bioethanol depend, among others, on the rawmaterial and technology of production. The productionof ethanol from sugar beet intermediate products is veryfavorable in that it lowers GHG emissions. The resultsof the study presented in this manuscript are aimed toimprove the production process leading to measurableeffects in terms of higher reduction of GHG emissions.

MethodsRaw material and microorganismsThick sugar beet juice was obtained from Dobrzelin SugarFactory (Dobrzelin, Poland). Fermentation was carriedout using a preparation of Ethanol Red dry distilleryyeast (S. cerevisiae), (Fermentis Division S.I.) designedfor the production of alcohol up to 18% (v/v) at hightemperature (35C). The number of living cells at packingwas >2.0 1010 per g, as declared by the manufacturer.

Preparation of fermentation wortsFermentation worts were prepared by diluting thick juicewith distilled water, initially at a ratio of 1:1 w/w, and thenobtaining solutions with an extract content of either 250or 280 g/kg. The worts were acidified with 25% (w/w) sul-furic acid to pH 4.8 and supplemented with (NH4)2HPO4(0.3 g/L) only or with (NH4)2HPO4 (0.3 g/L) and MgSO4 7H2O (0.1 g/L) as nutrients for yeast.

Fermentation variantsThe fermentation variants were as follows:

I. Extract content of 250 g/kg; inoculum content of1.0 g/L; (NH4)2HPO4

II. Extract content of 250 g/kg; inoculum content of1.0 g/L; (NH4)2HPO4 +MgSO4 7 H2O

III. Extract content of 250 g/kg; inoculum content of1.5 g/L; (NH4)2HPO4

IV. Extract content of 250 g/kg; inoculum content of2.0 g/L; (NH4)2HPO4

V. Extract content of 280 g/kg; inoculum content of1.0 g/L; (NH4)2HPO4

VI. Extract content of 280 g/kg; inoculum content of1.0 g/L; (NH4)2HPO4 +MgSO4 7 H2O

VII. Extract content of 280 g/kg; inoculum content of1.5 g/L; (NH4)2HPO4

VIII. Extract content of 280 g/kg, inoculum content of2.0 g/L; (NH4)2HPO4

Fermentation experiments were carried out in 6-L glassflasks, each containing approximately 3 L of wort. Afterinoculation with yeast, which was preliminarily rehydrated,the flasks were closed with stoppers equipped with fermen-tation pipes filled with glycerol and kept in a thermostat-controlled room at 35C. The process was carried out over4 days (96 h). During the fermentation, samples for analysiswere collected and the concentration of ethanol, realextract (after ethanol distillation), reducing sugars, andsaccharose was measured, allowing us to compare the dy-namics and biotechnological factors of the entire process.

DistillationWhen fermentation was complete, all ethanol was distilledfrom worts using a laboratory distillation unit consisting


of a distillation flask, a Liebig cooler, a flask for collectingethanol, and a thermometer. Raw spirits containing 20 to23% (v/v) ethanol were refined to approximately 43% (v/v)in distillation apparatus equipped with a bi-rectifier unit(dephlegmator according to Golodetz), and subjected tochemical analysis.

Analytical methodsThick juice was analyzed by the methods recommendedfor the sugar industry [24]. Solid substance (total extract)was measured by using a hydrometer, which indicates theconcentration of dissolved solids, mostly sugars, calibratedin g of saccharose per kg of water solution. Total nitrogenwas determined by the Kjeldahl method. Volatile acids(expressed as acetic acid) were assayed using steam dis-tillation. Reducing sugars and total sugars (after inversionwith hydrochloric acid) were estimated by the Lane-Eynonmethod. Both were expressed in g of invert sugar per kg ofthick juice. Saccharose concentration was calculated as thedifference between total sugars and reducing sugars (takinginto consideration a conversion coefficient of 0.95). AlsopH was measured (with a digital pH-meter).Worts were analyzed before and after fermentation using

methods recommended for distilleries. Prior to fermenta-tion, the worts were analyzed for pH, total extract, and re-ducing sugars (expressed as invert sugar) and saccharosecontent. On completion of fermentation, the worts wereanalyzed for real extract (after ethanol distillation), etha-nol concentration in wort (using a hydrometer with ascale in % v/v of ethanol) and sugars content.Distillates were analyzed using the Agillent 6890 N gas

chromatograph (USA, Wilmington) equipped with a flame-ionization detector (FID), a split/splitless injector and anHP-Innowax capillary column (60 m 32 mm 0.5 m).The temperature at the injector (split 1:45) and FID waskept at 250C. The temperature program was as follows:40C (6 minutes), an increase to 83C (2C/minutes) andthen to 190C (5C/minutes) (2 minutes). The flowrate of the carrier gas (helium) through the column was2 mL/minute.

Fermentation evaluationThe intake of total sugars (the percentage yield of sugarconsumption during fermentation) was calculated as a ra-tio of sugars used during the fermentation to their contentin the wort prior to this process, and expressed in percent.The yield of ethanol was calculated according to the stoi-chiometric Gay-Lussac equation in relation to total sugarsand expressed as a percentage of the theoretical yield.

Statistical analysisAll samples were prepared and analyzed in triplicate.Statistical analysis was carried out using the MicromalOrigin ver. 6.0 software (Northampton, USA).

Abbreviations(NH4)2HPO4: Diammonium hydrogen phosphate; MgSO4 7 H2O: Magnesiumsulfate heptahydrate; FID: Flame-ionization detector; GHG: Greenhouse gas;VHG: Very high gravity.

Competing interestsThe authors declare that they have no competing interests.

Authors contributionsPD and MB designed the experiments. MB, KP-P and PP performed theexperiments. MB and KP-P wrote the paper. PD and PP were involved inthe evaluation of results and review of the paper. All authors read andapproved the final manuscript.

Author details1Department of Fermentation Technology, Institute of FermentationTechnology and Microbiology, Lodz University of Technology, 90-924Wolczanska, Lodz 171/173, Poland. 2Department of Spirit and YeastTechnology, Institute of Fermentation Technology and Microbiology, LodzUniversity of Technology, 90-924 Wolczanska, Lodz 171/173, Poland.

Received: 3 July 2013 Accepted: 30 October 2013Published: 8 November 2013

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http://www.intechopen.com/books/liquid-gaseous-and-solid-biofuels-conversion-techniques/gas-fermentation-for-commercial-biofuels-productionhttp://www.intechopen.com/books/liquid-gaseous-and-solid-biofuels-conversion-techniques/gas-fermentation-for-commercial-biofuels-productionhttp://www.intechopen.com/books/liquid-gaseous-and-solid-biofuels-conversion-techniques/gas-fermentation-for-commercial-biofuels-productionhttp://www.fermentis.com/wp-content/uploads/2012/06/EthanolRED_EN.pdfhttp://www.fermentis.com/wp-content/uploads/2012/06/EthanolRED_EN.pdf


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doi:10.1186/1754-6834-6-158Cite this article as: Dziugan et al.: Evaluation of the fermentation of highgravity thick sugar beet juice worts for efficient bioethanol production.Biotechnology for Biofuels 2013 6:158.

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AbstractBackgroundResultsConclusions

BackgroundResults and discussionChemical characteristics of thick juiceThe effect of process conditions on fermentation dynamics and the results of fermentation of high gravity thick juice wortsAnalysis of the chemical composition of the obtained distillates

ConclusionsMethodsRaw material and microorganismsPreparation of fermentation wortsFermentation variantsDistillationAnalytical methodsFermentation evaluationStatistical analysisAbbreviations

Competing interestsAuthors contributionsAuthor detailsReferences

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