effect of yeast fermentation of green coffee beans on...

Research ArticleEffect of Yeast Fermentation of Green Coffee Beans onAntioxidant Activity and Consumer Acceptability

Han Sub Kwak 1 Yoonhwa Jeong 2 and Misook Kim 2

1Division of Functional Food Research Korea Food Research Institute 245 Nongsaengmyeong-ro Iseo-myeon Wanju-gunJeollabuk-do 55365 Republic of Korea2Department of Food Science and Nutrition Dankook University 19 Dandae-ro 119 Dongnam-gu Cheonan-siChungnam 31116 Republic of Korea

Correspondence should be addressed to Misook Kim mkim5dankookackr

Received 16 November 2017 Accepted 23 January 2018 Published 6 March 2018

Academic Editor Gabriel Henrique Horta de Oliveira

Copyright copy 2018 Han Sub Kwak et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

This study assessed the functionality and consumer acceptance of yeast fermented coffee beans Green coffee beans were fermentedfor 24 hwith three different yeast strains to increase functionalityThe yeast fermentationwas effective in fortifying the functionalityof coffee by significantly increasing antioxidant activity according to the results of ORAC and SOD-like assay (119875 lt 005) The TPCand TFC contents in the fermented coffee beans were significantly higher than those in the controls (119875 lt 005) The consumeracceptance for the fermented coffee beans was slightly lower than that of the controls Fermentation seemed to influence thearoma and flavor of coffee However agglomerative hierarchical clustering analysis revealed that approximately 39 of consumerssignificantly liked one of the fermented coffees (F3) more than the controls (119875 lt 005) These consumers indicated that the yeastfermentation of green coffee beans did not generate a negative aroma or flavor and can be attractive with high antioxidant activity

1 Introduction

Coffee is one of the most traded commodities worldwide Itis harvested in tropical regions andmostly exported to devel-oped countries such as the USA Europe Russia Japan andKorea [1 2]Many coffee drinkers consume coffee on a regularbasis similar to tea and coffee is regarded as a hedonic foodHowever coffee contains significant amounts of phenoliccompounds such as chlorogenic and hydroxycinnamic acidsand antioxidants including caffeine melanoidins and otherMaillard reaction products and volatile compounds [3ndash5]Levels of chlorogenic and hydroxycinnamic acids are alsodetermined on the final aroma and taste of the roasted coffee[6 7] Richelle et al [8] demonstrated that a cup of Robustaand Arabica coffee had two times more antioxidant activitythan a cup of green and black tea A cup of coffee per daywas found to reduce the relative risk of diabetes by 7 inmeta-analysis [9] andmoderate coffee drinking (below 4 cupsper day) showed the strongest inverse relation to heart failure[10] In some European countries coffee is one of the majorsources of antioxidants in the human diet [11ndash13]

The functionality of food products can be increased byfermentation Ginseng is one of the best known functionalfoods Ginsenoside contents and SOD-like activity afterfermentation were significantly increased by solid-microbialfermentation of white ginseng [14 15] Increased antioxi-dant activities and phenolic compounds are also frequentlyobserved in fermented tea products Jayabalan et al [16]showed an increase in epicatechin isomers over 18 daysof kombucha tea fermentation DPPH radical scavengingactivity and total polyphenol content in Pu-Erh tea weresignificantly increased by fermentation [17]

The fermentation of coffee is known as coffee cherryfermentation and effectively removes themucilage layer priorto the drying process for obtaining green coffee beans [18]Therefore the primary objective of coffee cherry fermen-tation is to improve the ease of obtaining green coffeebeans rather than to increase the functionality of the coffeebeans Green coffee beans can gain higher functionalitywith additional processing steps such as soaking in fruitextracts and fermentation Lim et al [19] reported higherantioxidant activity total polyphenol contents and consumer

HindawiJournal of Food QualityVolume 2018 Article ID 5967130 8 pageshttpsdoiorg10115520185967130

2 Journal of Food Quality

acceptability after soaking green coffee beans in mulberryextract However the fermentation of green coffee beanshas not been widely studied as a second processing stepfor increasing antioxidant activity and phenolic compoundsAs the fermentation of tea products increases their antiox-idant activity and the number of phenolic compounds theantioxidant activity and phenolic compounds in green coffeebeans could also be increased by fermentationThe fermentedcoffee beans can become fortified functional coffee beansTherefore the objective of this study was to ferment greencoffee beans using commercial yeasts and investigate theresulting physicochemical properties antioxidant activitytotal polyphenol and flavonoid contents and consumeracceptability with the intention ofmaking fortified functionalcoffee beans

2 Materials and Methods

21 Chemicals Sodium carbonate (Na2CO3) sodium nitrite(NaNO2) monosodium phosphate (NaH2PO4) disodiumphosphate (Na2HPO4) potassium acetate (CH3COOK) andaluminumchloride (AlCl3sdot6H2O)were purchased fromDuk-san Pure Chemicals (Ansan-si Korea) HCl NaCl ethanoland methanol were supplied by Samchun Pure Chemicals(Pyeongtaek-si Korea) D-Glucosewas bought fromDuchefaBiochemie BV (Haarlem Netherlands) YPD was purchasedfrom Becton Dickinson (Sparks MD USA) Folin-Ciocal-teursquos phenol reagent fluorescein sodium 221015840-azobis(2-am-idinopropane)dihydrochloride (AAPH) 25-dinitrosalicylicacid (DNS) trolox ethylenediaminetetraacetic acid (EDTA)pyrogallol quercetin and gallic acid were supplied by Sigma-Aldrich LLC (St Louis MO USA)

22 Coffee Fermentation Roasting andBrewing Green coffeebeans (300 g Brazil Ipanema Euro Natural Coiners Inter-national Ltd Bucheon-si Korea) were put into a water(450mL) and yeast (375mL 10 times 107 CFUmL) mixtureThree different commercial yeasts (Saccharomyces species)were used for fermentation F1 (Lalvin 71B Lallemand IncMontreal Canada) F2 (Lalvin Cy3079 Lallemand Inc) andF3 (BDX Lallemand Inc) Fermentation was conducted for24 h at 30∘C After fermentation green coffee beans werewashed three times using sterile water The fermented coffeebeans were dried in a dry oven at 45∘C until their moisturecontent reached 10 Two controls were used in this studyone control (C1) was the original green coffee beans and theother was a control (C2) treated with the same procedure asthe yeast fermented coffee beans but without yeasts

Coffee roasting was conducted with an automatic coffeeroaster to ensure a similar roasting condition for all greencoffee beans (Behmor 1600 Behmor Inc Incline Village NVUSA) The green coffee beans were put into the roaster androasted at maximum heating power (1600W) for 16minThe roasted coffee beans were cooled for 15min and kept atroom temperature for three days The roasted coffee beanswere ground using the medium option on a coffee grinder(KG79 DersquoLonghi Milano Italy) The ground coffee (36 g)was brewedwith 600mLof filteredwater using a coffeemaker(HD7564 Philips Netherlands)

23 Fermentation Characteristics The pH of the fermenta-tion solution was measured using an electronic pH meter(Orion 3 star Thermo Fisher Scientific Inc Waltham MAUSA) at 0 and 24 h

The reducing sugar content in the green coffee beanswas determined by the DNS method [20] with some mod-ifications A sample (200 uL) was mixed with 600 uL DNSsolution and heated at 100∘C for 5min Water (3mL) wasadded to the reaction solution and its absorbance wasmeasured at 550 nm D-Glucose solutions (01ndash25mgmL)were used for a standard curve (119877 = 0997)

Yeast-containing solutions for green coffee bean fermen-tation were serially diluted with 085 NaCl solution andspread onto the surface of YPD agar plates The spreadplates were incubated for 24 h at 30∘C to determine the totalmicrobial counts

24 Physicochemical Characteristics of Roasted Coffee Mois-ture content determination was performed according to theAssociation of Official Agricultural Chemists [21] guidelinesApproximately 1 g of ground coffee beans was dried at 105∘Cafter roasting until the weight became consistent

Crude ash content was also measured with the Associ-ation of Official Agricultural Chemists [21] guidelines Theroasted coffee beans (1 g) were incinerated at 550∘C until theweight became consistent Crude ash content was determinedgravimetrically

The color of the ground coffee after roasting was mea-sured using a Hunter colorimeter system (JC-801S ColorTechno system Co Tokyo Japan) Ground coffee (2 g) wasput into a small Petri dish for measurement Lightness (119871)redness (119886) and yellowness (119887) were measured 119871 119886 and 119887values for the standard white color plate were 119871 = 9838 119886 =029 and 119887 = minus041 respectively

25 Antioxidant Activity Oxygen radical absorbance capac-ity (ORAC) wasmeasured using themethod described by Ouet al [22] with some modifications for in vitro antioxidantactivity Phosphate (NaH2PO4-Na2HPO4) buffer (10mM pH70) was used to dissolve fluorescein powder Coffee extract(50 120583L) and 25mM fluorescein solution (150 120583L) were mixedand then incubated in a dark room for 10min A volumeof 25 120583L of 120mM AAPH solution was added to the coffeeextract and fluorescein mixture The control was 10mMphosphate buffer instead of coffee extract Fluorescence wasmeasured by the microplate reader (Spectra max M2 Molec-ular Devices LLC Sunnyvale CA USA) Measurementswere taken every minute for 90min (excitation wavelength485 nm emission wavelength 535 nm) The ORAC valueswere calculated by the following formula and presented as120583Mtrolox equivalentmL of coffee (120583MTEmL)

ORAC (120583M TEg)

= 119862Trolox times (AUCSample minus AUCBlank) times 119896(AUCTrolox minus AUCBlank)

(1)

where 119862Trolox 119896 and AUC were the concentration of trolox(5 120583M) the sample dilution factor and the area under

Journal of Food Quality 3

Table 1 Change of pH reducing sugar and viable cells before and after fermentation

Sample pH Reducing sugar (mgmL) Viable cell count (log CFUmL)0 h 24 h 0 h 24 h 0 h 24 h

C2 598 513 068 plusmn 001c(1) 257 plusmn 013a lt116 plusmn 053 778 plusmn 070aF1 579 492 112 plusmn 002b 251 plusmn 012a 650 plusmn 583 931 plusmn 085aF2 593 452 128 plusmn 003a 247 plusmn 009a 641 plusmn 562 925 plusmn 086aF3 578 504 112 plusmn 001b 267 plusmn 009a 630 plusmn 530 903 plusmn 081b(1)Different superscripts within a column meant significant difference at 119875 lt 005 by Fisherrsquos least significant difference test

the curve respectively AUC was calculated based on thefollowing formula

AUC = 1 +90

sum119899=1

1198911198991198910 (2)

where 119891119899 is the fluorescence at time 119899 (min)Superoxide dismutase-like (SOD-like) activity was deter-

mined using the method described by S Marklund andG Marklund [23] with some modifications Coffee extract(400 120583L) Tris-HCl buffer (600 120583L 50mM tris(hydroxym-ethyl)aminomethane and 10mM ethylenediaminetetraaceticacid (EDTA) pH 80) and 72mM pyrogallol (40 120583L) weremixed together and kept at 25∘C for 10minThe reaction wasstopped by adding 01 N HCl (20 120583L) The absorbance wasmeasured at 420 nm using a UVvisible spectrophotometer(Ultrospec 2100 Pro Biochrom Ltd Cambridge UK) SOD-like activity was calculated based on the following equation

SOD-like activity () = (1 minus 119860119861) times 100 (3)

where 119860 is absorbance of the sample and 119861 is absorbance ofthe control

26 Total Polyphenol and Flavonoid Contents The total pol-yphenol content (TPC) of brewed coffee was measured usingSingletonrsquos method [24] with some modifications Coffeeextract (20120583L) was diluted with 1580120583L of distilled waterDiluted coffee (160 120583L) was mixed with Folin-Ciocalteursquosphenol reagent (10 120583L) and allowed to sit for 8min A volumeof 30 120583L of 20 Na2CO3 solution was added and the mixturewas incubated in a dark room for 2 h Distilled water insteadof coffee extract was used as a control The absorbance wasmeasured at 765 nm by amicroplate reader (Spectra maxM2Molecular Devices LLC) Gallic acid solutions (0-1mgmL)were used to generate a standard curve (1198772 = 0997) Theresults were presented as mg gallic acid equivalentmL ofcoffee extract (mg GAEmL)

The total flavonoid content (TFC) for each coffee extractwas evaluated according to themethoddescribed byDewantostudy with some modifications [25] Coffee extract (250 120583L)distilled water (1mL) and 75 120583L of 5 NaNO2 were mixedtogether After 5min 10 AlCl3sdot6H2O solution (150 120583L) wasadded and incubated for 6min 1 N NaOH (500120583L) wasinjected and the mixture was incubated for 11min A blanksample was substituted for the diluted coffee extract withdistilled water The absorbance of the sample was measured

at 510 nm against the blank sample using a microplate reader(Spectra maxM2Molecular Devices LLC)The difference inabsorbance between the sample and the blank was comparedto the absorbance of the quercetin solution used as thepositive control (1198772 = 0999)The amount of flavonoids in thecoffee was presented as mg quercetin equivalentmL of coffeeextract

27 Consumer Acceptance Test The consumer acceptancetest for the yeast fermented coffee was performed with 74subjects (24 males and 50 females ages 19ndash30) who werestudents and staff at Dankook University (Yongin-si Korea)Subjects were coffee drinkers that consumed Americanocoffee (espresso coffee extract + hot water) at least once aweek Consumer testingwas conducted in an open area underincandescent lighting Subjects were assigned to tables andwere prohibited from talking and using cellphones during thetest Coffee (20mL) was served in 60mL paper cups using acompleted balanced design [26] in order to minimize carry-over effects The temperature of the coffee was approximately60∘C Acceptance of overall quality coffee aroma bitter tasteastringent taste and smooth mouthfeel was rated using anine-point hedonic scale on paper ballots Each categoryon the scale was labeled with numbers and descriptors inorder to give a clear understanding of the scale Starting fromthe left side the scale was labeled as 1 = dislike extremely2 = dislike very much 3 = dislike moderately 4 = dislikeslightly 5 = neither like nor dislike 6 = like slightly 7 = likemoderately 8 = like very much and 9 = extremely like A cupof filtered water was served as a palate cleanser Subjects weremonetarily compensated after the test (approximately $85)

28 Statistical Analyses Results were analyzed using Excelintegrated statistical software (XLSTAT version 2012 Addin-soft Paris France) in this study Analysis of variance(ANOVA) was performed to identify significant differencesacross samples Post hoc analysis was carried out usingFisherrsquos least significant test at 119875 lt 005when the significancewas observed by ANOVA Agglomerative hierarchical clus-tering (AHC) analysis was conducted to generate consumersegment groups based on their overall quality ratings

3 Results and Discussion

31 Fermentation Characteristics Measurements of pH val-ues reducing sugar contents and viable cell counts wereconducted to investigate the effect of yeast fermentation ongreen coffee beans (Table 1) Reducing sugar contents for


Table 2 Moisture content crude ash and color of ground roasted coffee beans after 24 h of yeast fermentation

Sample Moisture content () Crude ash () Color119871 119886 119887 Δ119864

C1 150 plusmn 009a(1) 473 plusmn 005a 3152 plusmn 101a 791 plusmn 044a 1678 plusmn 192a 6960 plusmn 127bC2 130 plusmn 005b 408 plusmn 006cd 2524 plusmn 062c 490 plusmn 025b 870 plusmn 089c 7400 plusmn 062aF1 127 plusmn 017b 405 plusmn 006d 3062 plusmn 081ab 711 plusmn 094a 1474 plusmn 080ab 6991 plusmn 052bF2 123 plusmn 006b 415 plusmn 005c 2517 plusmn 068c 443 plusmn 026b 763 plusmn 053c 7390 plusmn 061aF3 131 plusmn 010b 440 plusmn 005b 2978 plusmn 097b 697 plusmn 054a 1375 plusmn 108b 7051 plusmn 070b(1)Different superscripts within a column meant significant difference at P lt 005 by Fisherrsquos least significant difference test

2025b

1411b

3495a 3744a 3409a

C1 C2 F1 F2 F30

10

20

30

40

50

ORA

C (T

E

Mm

L of

coffe

e)

(a)

3367c 2963c

8276a 7979ab

6958b

C1 C2 F1 F2 F30

20

40

60

80

100

SOD

-like

activ

ity (

)

(b)

Figure 1 Antioxidant activities of yeast fermented coffee extracts by oxygen radical absorbance capacity (a) and superoxide dismutase-like(SOD-like) activity (b) assays Different letters meant significant difference at P lt 005 by Fisherrsquos least significant difference test

C2 F1 F2 and F3 were 2567 2507 2469 and 2673mgpH decreased after 24 h of fermentation The pH for C2also decreased This decrease would be due to the solubleorganic acids released from green coffee beans [27] andproduced by the fermentation of naturally occurring lacticacid bacteria in coffee beans After 24 h of fermentation therewas no significant difference in reducing sugar content in thefermented solution (119875 gt 005) Viable cell counts for C2F1 F2 and F3 showed that fermentation was conducted byyeasts (Table 1) The initial number of viable microbial cellswas less than 116 log CFUmL The number of microbialcells naturally present in green coffee beans increased to778 CFUmL after 24 h fermentation Solutions from F1 F2and F3 fermented by starter culture showed approximately25 higher log CFUmL of solution than that from C2This provided supporting evidence for the proliferation ofyeasts during fermentation and the role of yeasts in thefermentationThroughout the 24 h of fermentation there wasno drastic change in pH and reducing sugar content henceyeast fermentation occurred across all samples

32 Quality Characteristics of Fermented Coffee Beans Themoisture content crude ash and color of fermented coffeebeans after roasting are presented in Table 2 No signifi-cant difference across fermented coffee beans and C2 wasobserved This indicates that the drying and roasting of thefermented green coffee beans were performed identicallyThemoisture content of C1 was 150 andwas significantly higher

than those of C2 and the fermented coffee beans (119875 lt 005)This was due to slight overdrying prior to roasting The ashcontent was also significantly higher in C1 at 473 whilethe others ranged from 405 to 440 Moisture content andcrude ash showed a strong negative correlation (119877 = 0894119875 = 0041) The color of the ground roasted coffee beansshowed differences across the samples C1 F1 and F3 hadhigher 119871 119886 and 119887 values and lower Δ119864 values than C2 andF2 The color of the ground coffee beans showed a differentpattern in comparison with the moisture content and crudeashTherefore the different colors of the roasted coffee beanslikely partially originated from the fermentation as well as thedifference in moisture contents

33 Antioxidant Activity The antioxidant activities of thecoffee extracts in ORAC and SOD-like assays are shownin Figure 1 ORAC for the yeast fermented coffee extractswere 3495 3744 and 3409120583MTEmL for F1 F2 and F3respectively (Figure 1(a)) ORAC for C1 and C2 were 2025and 1411 120583MTEmL respectively There was no significantdifference in ORAC values after soaking the green coffeebeans for 24 h (119875 gt 005) however a slight decrease inthe ORAC value was observed in C2 It seemed that thesoluble antioxidants in green coffee beans might be elutedinto water The ORAC values of the yeast fermented coffeeextracts (F1 F2 and F3) were significantly higher than thosefrom the controls (C1 and C2) (119875 lt 005) There wasno significant difference across the fermented coffee beans


086c

072c

111b 118ab130a

C1 C2 F1 F2 F300

05

10

15TP

C (G

AE

mg

mL

of co

ffee)

(a)

089cd077d

101bc 104b

121a

C1 C2 F1 F2 F300

05

10

15

TFC

(mM

QE

mL

of co

ffee)

(b)

Figure 2 Total polyphenol (a) and flavonoid (b) contents of yeast fermented coffee extracts Different letters meant significant difference atP lt 005 by Fisherrsquos least significant difference test

(119875 lt 005) In SOD-like activity F1 F2 and F3 showed8276 7979 and 6958 respectively (Figure 1(b)) On theother hand C1 and C2 showed 3367 and 2963 SOD-likeactivity respectively and were not significantly different (119875 gt005) An increase of more than two times the SOD-likeactivity was observed after 24 h of yeast fermentation Amongthe fermented samples F1 showed the highest activity andthe activity was significantly higher than F3 (119875 lt 005)Soaking green coffee beans in water (C2) did not significantlyinfluence the antioxidant activity of the coffee extracts (119875 gt005) although the ORAC value of C2 was slightly lowerthan that of C1 Fermentation for 24 h was effective inincreasing the antioxidant activity significantly The increaseof antioxidant activity by fermentation was similar to otherfermented materials such as ginseng garlic tea and soy[15 16 28 29]

34 Total Polyphenol and Flavonoid Contents The TPC andTFC of the yeast fermented coffee extracts are presented inFigure 2 The yeast fermented coffee extracts (F1 F2 andF3) had 111 118 and 130GAEmgmL of coffee extractrespectively (Figure 2(a)) F3 had significantly higher TPCthan F1 (119875 lt 005) Control samples (C1 and C2) hadsignificantly lower amounts of TPC than the yeast fermentedcoffee extracts (119875 lt 005) C2 had the lowest TPC at072GAEmgmL This was due to the elution of solublephenolic compounds in the green coffee beans during thesoaking period (24 h) A similar finding was observed aftersoaking green coffee beans in mulberry extract [19] In Limrsquosstudy the TPC of the coffee extracts decreased after 6 h ofsoaking Despite the decrease in TPC due to the soakingprocess in this study fermentation positively influenced TPCin the coffee extracts During the fermentation processstrongly bound phenolic compounds in the cell wall maybecome weakened [30] making them easier to extract afterroasting Phenolic compounds in coffee are mostly in theforms of chlorogenic acidswith 5-O-caffeoyl-quinic acid suchas caffeic ferulic p-coumaric and caffeoylquinic acid [3]Feruloylquinic acid di-caffeoyl-quinic acid and proantho-cyanidins are also detected in coffee [3]

In TFC the yeast fermented coffee extracts (F1 F2and F3) had 101 104 and 121 QEmgmL of coffee extractrespectively and F3 had significantly higher (119875 lt 005)TFC than F1 and F2 (Figure 2(b)) C1 and C2 had loweramounts of TFC than the yeast fermented coffee extractsYeast fermentation was effective in increasing the number offlavonoids in the coffee extracts The increase in TFC mightbe due to the conversion of insoluble phenolic compoundsinto soluble flavonoids during fermentation [31] The ratio ofTFC to TPC in C1 and C2 was 103 and 107 while F1 F2and F3 had ratios of 091 088 and 093 respectively Theratio was lower in the yeast fermented coffee extracts Fer-mentation was more effective in producing soluble phenoliccompounds than flavonoidsThis result contrastedwith thosefor fermented Columbian coffee [32] However Columbiancoffee fermentation was conducted with broken green beanswhich might elute more soluble phenolic compounds easilyover seven days of fermentation This would also influencethe roasting process

35 Consumer Acceptance Test Consumer acceptance for thefermented and control coffee extracts is shown in Table 3Mean overall quality rating was the highest in C1 followedby F3 F1 and F2 had significantly lower overall qualityratings than C1 at 435 and 443 respectively (119875 lt 005)C2 also received slightly lower overall quality ratings thanC1 Therefore the soaking process does appear to slightlyinfluence the coffee quality Color acceptance was not sig-nificantly different (119875 lt 005) which meant that therewas no difference in the appearance of the coffee extractAroma acceptance ratings for the fermented coffee samples(F1 F2 and F3) were significantly lower than those forthe controls (C1 and C2) (119875 lt 005) The consumeracceptability was within the range of the previous acceptancestudy using various roasting conditions [33] Fermentationmight negatively influence the aroma acceptance of coffeeAcross the fermented coffee samples (F1 F2 and F3) thedifferences in consumer acceptability might be related to thepresence of different volatile compounds in coffee beans afterfermentation Different yeasts showed different flavor profiles


Table 3 Overall quality and acceptances of color aroma sourness bitterness astringency and mouthfeel of yeast fermented coffee extractby 74 consumers

Sample Overall quality Color Aroma Sourness Bitterness Astringency MouthfeelC1 509 plusmn 172a(1) 559 plusmn 176a 562 plusmn 163a 512 plusmn 186a 451 plusmn 190a 481 plusmn 180a 580 plusmn 175aC2 469 plusmn 165ab 530 plusmn 175a 512 plusmn 170a 507 plusmn 149a 412 plusmn 194ab 457 plusmn 182ab 535 plusmn 168abF1 435 plusmn 170b 518 plusmn 148a 408 plusmn 185b 478 plusmn 150a 384 plusmn 176b 420 plusmn 176b 499 plusmn 172bF2 443 plusmn 146b 523 plusmn 144a 451 plusmn 151b 419 plusmn 158b 401 plusmn 185ab 403 plusmn 179b 485 plusmn 166bF3 484 plusmn 167ab 554 plusmn 160a 445 plusmn 195b 484 plusmn 147a 441 plusmn 189ab 436 plusmn 158ab 512 plusmn 157b(1)Different superscripts within a column meant significant difference at P lt 005 by Fisherrsquos least significant difference test

Table 4 Agglomerative hierarchical clustering (AHC) analysis of overall quality

Class Subject(1) C1 C2 F1 F2 F31 28 379 plusmn 120b(2)B(3) 382 plusmn 161bB 357 plusmn 148bB 489 plusmn 166aA 532 plusmn 161aA2 43 595 plusmn 153aA 523 plusmn 149bA 481 plusmn 172bcA 409 plusmn 129dB 451 plusmn 171cdB(1)Three subjects were removed from AHC analysis because they marked same ratings for entire samples (2)Different superscripts within a row meantsignificant difference at 119875 lt 005 by Fisherrsquos least significant difference test (3)Different subscripts within a column meant significant difference at 119875 lt 005

in rice distilled liquor [34] and cachaca [35] Acceptanceratings for sourness bitterness astringency and mouthfeelwere lower in the yeast fermented coffee samples than in C1and C2

In order to segment consumers into a small number ofgroups AHCanalysis was performedusing the overall qualityratings for all except three consumers that selected the sameacceptance ratings for the entire sample (Table 4) Consumerswere divided into two clusters generated by AHC analysisCluster 1 was composed of 394 of the total consumersThese were the consumers who preferred fermented coffee(F2 and F3) while disliking the controls (C1 and C2) (119875 lt005) On the other hand cluster 2 was composed of 606of consumers These consumers had higher overall qualityratings for the controls (C1 and C2) They also rated F1 as anaverage of 481 and had the least acceptance for F2 at 409Approximately 40 of consumers preferred the fermentedcoffee extracts (F2 and F3) although fermented coffee hadlower mean overall quality ratings for the entire group ofconsumers Therefore the fermented coffee samples (F2 andF3) did not have lower acceptance ratings for all consumersthey were acceptable and showed higher acceptability for394of consumersThis result provides supporting evidencethat fermentation does not have a negative influence onthe consumer acceptance of coffee Consumers are thereforesegmented and another coffee product can be created forapproximately 40 of coffee consumers In addition the highantioxidant activity and phenolic compounds in fermentedcoffee can be attractive to those consumers with moderateacceptance for fermented coffee The consumer acceptancewould also increase with awareness of the high antioxidantactivity and phenolic compounds content as shown in aprevious blind and informed consumer acceptance test forblueberry functional beverages [36]

4 Conclusion

Yeast fermentation of green coffee beans for 24 h was effec-tive in fortifying the functionality of coffee by inducing

a significant increase in antioxidant activity TPC and TFCYeast fermentation of green coffee beans causes bound phe-nolic compounds to be released after roastingThe consumeracceptance for the fermented coffee beans was slightly lowerthan for the controls Fermentation might negatively influ-ence the aroma and flavor of coffee extracts However theconsumer segmentation revealed that approximately 394ofconsumers preferred one of the fermented coffees (F3) morethan the controls Therefore it can be concluded that yeastfermentation did not always generate a negative aroma andflavor for consumers If fermentation was carried out withproperly selected yeasts fermented coffee can be attractiveto coffee consumers and coffee manufacturers can diversifytheir products with higher functionality

Conflicts of Interest

The authors declare that they have no conflicts of interest

References

[1] H D Vieira ldquoCoffee The plant and its cultivationrdquo in Plant-Parasitic Nematodes of Coffee M Souza Ed pp 3ndash18 SpringerDordrecht Netherlands 2008

[2] ldquoInternational Coffee Organization (ICO)rdquo 2015 httpwwwicoorgtrade statisticsasp

[3] P Parras MMartınez-Tome A M Jimenez andM AMurcialdquoAntioxidant capacity of coffees of several origins brewedfollowing three different proceduresrdquo Food Chemistry vol 102no 3 pp 582ndash592 2007

[4] M Perez-Martınez B Caemmerer M P de Pena C Con-cepcion and L W Kroh ldquoInfluence of brewing method andacidity regulators on the antioxidant capacity of coffee brewsrdquoJournal of Agricultural and Food Chemistry vol 58 no 5 pp2958ndash2965 2010

[5] I Sanchez-Gonzalez A Jimenez-Escrig and F Saura-CalixtoldquoIn vitro antioxidant activity of coffees brewed using differentprocedures (Italian espresso and filter)rdquo Food Chemistry vol90 no 1-2 pp 133ndash139 2005


[6] J R Oliveira-Neto S G Rezende C De Fatima Reis S RBenjamin M L Rocha and E De Souza Gil ldquoElectrochemicalbehavior and determination of major phenolic antioxidants inselected coffee samplesrdquo Food Chemistry vol 190 no 1 ArticleID 17651 pp 506ndash512 2016

[7] J A Vignoli M C Viegas D G Bassoli and M D T BenassildquoRoasting process affects differently the bioactive compoundsand the antioxidant activity of arabica and robusta coffeesrdquo FoodResearch International vol 61 no 1 pp 279ndash285 2014

[8] M Richelle I Tavazzi and E Offord ldquoComparison of theantioxidant activity of commonly consumed polyphenolic bev-erages (coffee cocoa and tea) prepared per cup servingrdquoJournal of Agricultural and Food Chemistry vol 49 no 7 pp3438ndash3442 2001

[9] R Huxley C M Y Lee F Barzi et al ldquoCoffee decaffeinatedcoffee and tea consumption in relation to incident type 2diabetes mellitus a systematic review with meta-analysisrdquorchives of Internal Medicine vol 169 no 22 pp 2053ndash20632009

[10] E Mostofsky M S Rice E B Levitan and M A MittlemanldquoHabitual coffee consumption and risk of heart failure a dose-response meta-analysisrdquo Circulation Heart Failure vol 5 no 4pp 401ndash405 2012

[11] R PulidoMHernnndez-Garcıa and F Saura-Calixto ldquoContri-bution of beverages to the intake of lipophilic and hydrophilicantioxidants in the Spanish dietrdquo European Journal of ClinicalNutrition vol 57 no 10 pp 1275ndash1282 2003

[12] S A Qureshi A C Lund M B Veieroslashd et al ldquoFood itemscontributing most to variation in antioxidant intake a cross-sectional study amongNorwegian womenrdquo BMCPublic Healthvol 14 no 1 article no 45 2014

[13] A Svilaas A K Sakhi L F Andersen et al ldquoIntakes ofantioxidants in coffee wine and vegetables are correlated withplasma carotenoids in humansrdquo Journal of Nutrition vol 134no 3 pp 562ndash567 2004

[14] B-G Kim S-Y Choi M-R Kim H J Suh and H JPark ldquoChanges of ginsenosides in Korean red ginseng (Panaxginseng) fermented by Lactobacillus plantarum M1rdquo ProcessBiochemistry vol 45 no 8 pp 1319ndash1324 2010

[15] S-I Lim C-W Cho U-K Choi and Y-C Kim ldquoAntioxidantactivity and ginsenoside pattern of fermented white ginsengrdquoJournal of Ginseng Research vol 34 no 3 pp 168ndash174 2010

[16] R Jayabalan S Marimuthu and K Swaminathan ldquoChanges incontent of organic acids and tea polyphenols during kombuchatea fermentationrdquo Food Chemistry vol 102 no 1 pp 392ndash3982007

[17] K-C Jeng C-S Chen Y-P Fang R C-WHou andY-S ChenldquoEffect of microbial fermentation on content of statin GABAand polyphenols in Pu-erh teardquo Journal of Agricultural and FoodChemistry vol 55 no 21 pp 8787ndash8792 2007

[18] C F Silva L R Batista LMAbreu E SDias andR F SchwanldquoSuccession of bacterial and fungal communities during naturalcoffee (Coffea arabica) fermentationrdquo Food Microbiology vol25 no 8 pp 951ndash957 2008

[19] H H Lim S Ji H S Kwak et al ldquoQuality characteristics ofcoffee brewed from green beans soaked in mulberry (Morusbombycis) extractrdquo Journal of the Korean Society of Food Scienceand Nutrition vol 44 no 4 pp 579ndash585 2015

[20] T K Ghose ldquoMeasurement of cellulase activitiesrdquo Pure andApplied Chemistry vol 59 no 2 pp 257ndash268 1987

[21] AOAC Official Methods of Analysis Association of OfficialAnalytical Chemists Gaithersburg MD USA 18th editionedition 2005

[22] BOuMHampsch-Woodill andR L Prior ldquoDevelopment andvalidation of an improved oxygen radical absorbance capacityassay using fluorescein as the fluorescent proberdquo Journal ofAgricultural and Food Chemistry vol 49 no 10 pp 4619ndash46262001

[23] S Marklund and G Marklund ldquoInvolvement of the superoxideanion radical in the autoxidation of pyrogallol and a convenientassay for superoxide dismutaserdquo European Journal of Biochem-istry vol 47 no 3 pp 469ndash474 1974

[24] V L Singleton R Orthofer and R M Lamuela-RaventosldquoAnalysis of total phenols and other oxidation substrates andantioxidants by means of folin-ciocalteu reagentrdquo Methods inEnzymology vol 299 pp 152ndash178 1999

[25] V Dewanto X Wu and R H Liu ldquoProcessed sweet corn hashigher antioxidant activityrdquo Journal of Agricultural and FoodChemistry vol 50 no 17 pp 4959ndash4964 2002

[26] H J MacFie N Bratchell K Greenhoff and L V VallisldquoDesigns to balance the effect of order of presentation and first-order carry-over effects in hall testsrdquo Journal of Sensory Studiesvol 4 no 2 pp 129ndash148 1989

[27] C I Rodrigues L Marta R Maia M Miranda M Ribeir-inho and C Maguas ldquoApplication of solid-phase extractionto brewed coffee caffeine and organic acid determination byUVHPLCrdquo Journal of Food Composition and Analysis vol 20no 5 pp 440ndash448 2007

[28] Y-H Pyo T-C Lee and Y-C Lee ldquoEffect of lactic acidfermentation on enrichment of antioxidant properties andbioactive isoflavones in soybeanrdquo Journal of Food Science vol70 no 3 pp S215ndashS220 2005

[29] E Sato M Kohno H Hamano and Y Niwano ldquoIncreasedanti-oxidative potency of garlic by spontaneous short-termfermentationrdquo Plant Foods for Human Nutrition vol 61 no 4pp 157ndash160 2006

[30] M Shi Y Yang Q Wang Y Zhang Y Wang and Z ZhangldquoProduction of total polyphenol from fermented soybean curdresidue by Lentinus edodesrdquo International Journal of FoodScience amp Technology vol 47 no 6 pp 1215ndash1221 2012

[31] Y-K Lee S-I Lee J-S Kim et al ldquoAntioxidant activity ofgreen tea fermented with Monascus pilosusrdquo Journal of AppliedBiological Chemistry vol 55 no 1 pp 19ndash25 2012

[32] J-Y Shin H Kim D-G Kim G-H Baek H-S Jeong andK-W Yu ldquoPharmacological activities of coffee roasted fromfermented green coffee beans with fungal mycelia in solid-state culturerdquo Journal of the Korean Society of Food Science andNutrition vol 42 no 3 pp 487ndash496 2013

[33] L C Mendes H C De Menezes M Aparecida and A PDa Silva ldquoOptimization of the roasting of robusta coffee (Ccanephora conillon) using acceptability tests and RSMrdquo FoodQuality and Preference vol 12 no 2 pp 153ndash162 2001

[34] H S Kwak J S Seo Y Hur et al ldquoInfluence of yeaststrains on the physicochemical characteristics methanol andacetaldehyde profiles and volatile compounds for Korean ricedistilled spiritrdquo Journal of the Institute of Brewing vol 121 no 4pp 574ndash580 2015

[35] E S Oliveira H M A B Cardello E M Jeronimo E L RochaSouza and G E Serra ldquoThe influence of different yeasts onthe fermentation composition and sensory quality of cachacardquoWorld Journal of Microbiology and Biotechnology vol 21 no 5pp 707ndash715 2005


[36] M K Kim andH S Kwak ldquoInfluence of functional informationon consumer liking and consumer perception related to healthclaims for blueberry functional beveragesrdquo International Jour-nal of Food Science amp Technology vol 50 no 1 pp 70ndash76 2015

Hindawiwwwhindawicom

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MicrobiologyHindawiwwwhindawicom

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Submit your manuscripts atwwwhindawicom


acceptability after soaking green coffee beans in mulberryextract However the fermentation of green coffee beanshas not been widely studied as a second processing stepfor increasing antioxidant activity and phenolic compoundsAs the fermentation of tea products increases their antiox-idant activity and the number of phenolic compounds theantioxidant activity and phenolic compounds in green coffeebeans could also be increased by fermentationThe fermentedcoffee beans can become fortified functional coffee beansTherefore the objective of this study was to ferment greencoffee beans using commercial yeasts and investigate theresulting physicochemical properties antioxidant activitytotal polyphenol and flavonoid contents and consumeracceptability with the intention ofmaking fortified functionalcoffee beans

2 Materials and Methods

21 Chemicals Sodium carbonate (Na2CO3) sodium nitrite(NaNO2) monosodium phosphate (NaH2PO4) disodiumphosphate (Na2HPO4) potassium acetate (CH3COOK) andaluminumchloride (AlCl3sdot6H2O)were purchased fromDuk-san Pure Chemicals (Ansan-si Korea) HCl NaCl ethanoland methanol were supplied by Samchun Pure Chemicals(Pyeongtaek-si Korea) D-Glucosewas bought fromDuchefaBiochemie BV (Haarlem Netherlands) YPD was purchasedfrom Becton Dickinson (Sparks MD USA) Folin-Ciocal-teursquos phenol reagent fluorescein sodium 221015840-azobis(2-am-idinopropane)dihydrochloride (AAPH) 25-dinitrosalicylicacid (DNS) trolox ethylenediaminetetraacetic acid (EDTA)pyrogallol quercetin and gallic acid were supplied by Sigma-Aldrich LLC (St Louis MO USA)

22 Coffee Fermentation Roasting andBrewing Green coffeebeans (300 g Brazil Ipanema Euro Natural Coiners Inter-national Ltd Bucheon-si Korea) were put into a water(450mL) and yeast (375mL 10 times 107 CFUmL) mixtureThree different commercial yeasts (Saccharomyces species)were used for fermentation F1 (Lalvin 71B Lallemand IncMontreal Canada) F2 (Lalvin Cy3079 Lallemand Inc) andF3 (BDX Lallemand Inc) Fermentation was conducted for24 h at 30∘C After fermentation green coffee beans werewashed three times using sterile water The fermented coffeebeans were dried in a dry oven at 45∘C until their moisturecontent reached 10 Two controls were used in this studyone control (C1) was the original green coffee beans and theother was a control (C2) treated with the same procedure asthe yeast fermented coffee beans but without yeasts

Coffee roasting was conducted with an automatic coffeeroaster to ensure a similar roasting condition for all greencoffee beans (Behmor 1600 Behmor Inc Incline Village NVUSA) The green coffee beans were put into the roaster androasted at maximum heating power (1600W) for 16minThe roasted coffee beans were cooled for 15min and kept atroom temperature for three days The roasted coffee beanswere ground using the medium option on a coffee grinder(KG79 DersquoLonghi Milano Italy) The ground coffee (36 g)was brewedwith 600mLof filteredwater using a coffeemaker(HD7564 Philips Netherlands)

23 Fermentation Characteristics The pH of the fermenta-tion solution was measured using an electronic pH meter(Orion 3 star Thermo Fisher Scientific Inc Waltham MAUSA) at 0 and 24 h

The reducing sugar content in the green coffee beanswas determined by the DNS method [20] with some mod-ifications A sample (200 uL) was mixed with 600 uL DNSsolution and heated at 100∘C for 5min Water (3mL) wasadded to the reaction solution and its absorbance wasmeasured at 550 nm D-Glucose solutions (01ndash25mgmL)were used for a standard curve (119877 = 0997)

Yeast-containing solutions for green coffee bean fermen-tation were serially diluted with 085 NaCl solution andspread onto the surface of YPD agar plates The spreadplates were incubated for 24 h at 30∘C to determine the totalmicrobial counts

24 Physicochemical Characteristics of Roasted Coffee Mois-ture content determination was performed according to theAssociation of Official Agricultural Chemists [21] guidelinesApproximately 1 g of ground coffee beans was dried at 105∘Cafter roasting until the weight became consistent

Crude ash content was also measured with the Associ-ation of Official Agricultural Chemists [21] guidelines Theroasted coffee beans (1 g) were incinerated at 550∘C until theweight became consistent Crude ash content was determinedgravimetrically

The color of the ground coffee after roasting was mea-sured using a Hunter colorimeter system (JC-801S ColorTechno system Co Tokyo Japan) Ground coffee (2 g) wasput into a small Petri dish for measurement Lightness (119871)redness (119886) and yellowness (119887) were measured 119871 119886 and 119887values for the standard white color plate were 119871 = 9838 119886 =029 and 119887 = minus041 respectively

25 Antioxidant Activity Oxygen radical absorbance capac-ity (ORAC) wasmeasured using themethod described by Ouet al [22] with some modifications for in vitro antioxidantactivity Phosphate (NaH2PO4-Na2HPO4) buffer (10mM pH70) was used to dissolve fluorescein powder Coffee extract(50 120583L) and 25mM fluorescein solution (150 120583L) were mixedand then incubated in a dark room for 10min A volumeof 25 120583L of 120mM AAPH solution was added to the coffeeextract and fluorescein mixture The control was 10mMphosphate buffer instead of coffee extract Fluorescence wasmeasured by the microplate reader (Spectra max M2 Molec-ular Devices LLC Sunnyvale CA USA) Measurementswere taken every minute for 90min (excitation wavelength485 nm emission wavelength 535 nm) The ORAC valueswere calculated by the following formula and presented as120583Mtrolox equivalentmL of coffee (120583MTEmL)

ORAC (120583M TEg)

= 119862Trolox times (AUCSample minus AUCBlank) times 119896(AUCTrolox minus AUCBlank)

(1)

where 119862Trolox 119896 and AUC were the concentration of trolox(5 120583M) the sample dilution factor and the area under






AUC = 1 +90

sum119899=1

1198911198991198910 (2)
















2025b

1411b

3495a 3744a 3409a

C1 C2 F1 F2 F30

10

20

30

40

50

ORA

C (T

E

Mm

L of

coffe

e)

(a)

3367c 2963c

8276a 7979ab

6958b

C1 C2 F1 F2 F30

20

40

60

80

100

SOD

-like

activ

ity (

)

(b)







086c

072c

111b 118ab130a

C1 C2 F1 F2 F300

05

10

15TP

C (G

AE

mg

mL

of co

ffee)

(a)

089cd077d

101bc 104b

121a

C1 C2 F1 F2 F300

05

10

15

TFC

(mM

QE

mL

of co

ffee)

(b)













4 Conclusion





References









































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018







AUC = 1 +90

sum119899=1

1198911198991198910 (2)
















2025b

1411b

3495a 3744a 3409a

C1 C2 F1 F2 F30

10

20

30

40

50

ORA

C (T

E

Mm

L of

coffe

e)

(a)

3367c 2963c

8276a 7979ab

6958b

C1 C2 F1 F2 F30

20

40

60

80

100

SOD

-like

activ

ity (

)

(b)







086c

072c

111b 118ab130a

C1 C2 F1 F2 F300

05

10

15TP

C (G

AE

mg

mL

of co

ffee)

(a)

089cd077d

101bc 104b

121a

C1 C2 F1 F2 F300

05

10

15

TFC

(mM

QE

mL

of co

ffee)

(b)













4 Conclusion





References









































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018






2025b

1411b

3495a 3744a 3409a

C1 C2 F1 F2 F30

10

20

30

40

50

ORA

C (T

E

Mm

L of

coffe

e)

(a)

3367c 2963c

8276a 7979ab

6958b

C1 C2 F1 F2 F30

20

40

60

80

100

SOD

-like

activ

ity (

)

(b)







086c

072c

111b 118ab130a

C1 C2 F1 F2 F300

05

10

15TP

C (G

AE

mg

mL

of co

ffee)

(a)

089cd077d

101bc 104b

121a

C1 C2 F1 F2 F300

05

10

15

TFC

(mM

QE

mL

of co

ffee)

(b)













4 Conclusion





References









































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018



086c

072c

111b 118ab130a

C1 C2 F1 F2 F300

05

10

15TP

C (G

AE

mg

mL

of co

ffee)

(a)

089cd077d

101bc 104b

121a

C1 C2 F1 F2 F300

05

10

15

TFC

(mM

QE

mL

of co

ffee)

(b)













4 Conclusion





References









































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018









4 Conclusion





References









































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018





































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018


effect of yeast fermentation of green coffee beans on...

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