biodegradacion de fenoles.pdf

7/29/2019 biodegradacion de fenoles.pdf

1/7

~ Pergamon PIh S0043-1354(97)00014-6War. Res. Vol. 31, No. 8, pp. 2005-2011, 1997 1997 Elsevier Science Ltd. A ll rights reservedPrinted in Great Britain0043-1354/97 $17.00 + 0 .00

BIODEGRADATION OF PHENOL COMPOUNDS INVINASSE USING ASPERGILLUS TERREUS AND

GEOTRICHUM CANDIDUMI . G A R C I A G A R C I A * , J . L . B O N I L L A V E N C E S L A D A , P . R . J I M t ~ N E Z PE I~ IA a n d

E . R A M O S G O M E ZD e p a r t a m e n t o d e I n g e n i e r i a Q u i m i c a , F a c u l t a d d e C i e n c i as , U n i v e r s i d a d d e C 6 r d o b a , A v d a S a n A l b e r t oM a g n o s / n , E - 1 4 0 0 4 C 6 r d o b a , S p a i n

(Received September 1995; accepted in revised fo rm January 1997)A b s t r a c t - - A m o n g s t t h e m e t h o d s f o r t h e p u r i f i c a t i o n o f v i n a s s e , a n a e r o b i c d i g e s t i o n f e a t u r e s s o m eadvantages tha t make i t par t i cu la r ly s u i t ab le fo r p roces s ing th i s t ype o f was te . However , i t s e f f i c i ency i ss o m e w h a t d e c r e a s e d b y t h e p r e s e n ce o f p h e n o l c o m p o u n d s . T h e p r i o r r e m o v a l o f t h es e c o m p o u n d s m a k et h e v i n a s s e m o r e a m e n a b l e t o a n a e r o b i c d i g e s t i o n . A p o s s i b i l i t y o f r e d u c i n g t h e p h e n o l i c c o n t e n t i st h r o u g h a n a e r o b i c b i o l o g i c a l p r e t r e a t m e n t b y s o m e f u n g i . S o , i n t h e f r a m e w o r k o f b r o a d e r r e s e a r c hd i r e c t e d t o w a r d s t h e p u r i f i c a ti o n o f v i n a s s e b y t w o - s t a g e t r e a t m e n t ( a n a e r o b i c f e r m e n t a t i o n f o ll o w e d b yan anaerob ic d iges t ion) , t h i s work s tud ies exc lus ive ly the aerob ic s t age o f Aspergillus terreus a n dGeotrichum candidum. T h e s t u d y i s a i m e d a t f i n d i n g t h e m o s t s u i t a b l e m i c r o o r g a n i s m f o r r e m o v i n gp h e n o l i c c o m p o u n d s . T h e f e r m e n t a t i o n s fo l l o w M o n o d k i n e ti c s. T h e m a t h e m a t i c a l m o d e l i s u s e d in o r d e rt o c o m p a r e t h e m i c r o o r g a n i s m s b e h a v i o u r , a n d i t a l l o w s t h e p r e d i c t i o n o f t h e r e m o v a l o f p h e n o l s . T h ee x t e n t o f r e m o v a l , i n t h e f e r m e n t a t i o n b y A. terreus, o f t o t a l p h e n o l s a n d o - d i p h e n o l s w a s c . 6 6 a n d 9 4 % ,res pec t ive ly ; i n the cas e o f G. candidum, t he r es u l t s were c . 70 and 91%. For A. terreus, t h e m a x i m u mspecif ic growth rate is / lm~x= 0 . 0 6 h - ~, t h e c o n s t a n t Ks= 1 3 , 5 2 5 m g C O D l ~, t h e g r o w t h y i el d i sY~,,s= 0 . 38 mg ce l l s per mg COD and the e f f i c i ency on degrad ing phenol i c compounds i s E = 0 . 033 mgto ta l phe nol s per mg ce l ls . For G. candidum, t hes e cons tan t s a r e /~ , ,x = 0 . 047 h -~ , K} = 4558 m g CO D 1 ~ ,Yi.s = 0 .3 9 m g c el ls p e r n a g C O D a n d E ' = 0 .0 3 6 m g t o t a l p h e n o l s p e r m g c e l ls . T h o u g h b o t hm i c r o o r g a n i s m s b e h a v e i n a s i m i l a r w a y , G. candidum s e e m s t o b e s l ig h t ly b e t t e r f o r r e m o v i n g p h e n o l sf rom v inas s e . 1997 E l s ev ier Sc ience L tdKey words--waste-water, vinas s e , phenol s , Aspergillus terreus, Geotrichum candidum, a n a e r o b i c d i g e s t io n

NOMENCLATUREE = A. terreus ef f i c i ency on degrad ing to t a l phenol i cc o m p o u n d s ( r a g t o t a l p h e n o l s p e r m g c e l l s )

E' = G. candidum ef f i c i ency on degrad ing to t a l phenol i cc o m p o u n d s ( r a g t o t a l p h e n o l s p e r m g c e l l s )F = t o t a l p h e n o l s c o n c e n t r a t i o n ( r a g 1 ~ )Fc,~c = to t a l phe nol s con cen t r a t io n es t imated by th e k ine t i cm o d e l ( m g 1 - ')F0 = in i t i a l t o t a l ph enol s c onc en t r a t ion ( r ag 1 ~)Ks = c o n s t a n t i n e q u a t io n o f M o n o d f o r A. terreus(nag COD 1 ~)K } = c o n s t a n t i n e q u a t i o n o f M o n o d f o r G. candidum( r a g C O D 1 - ' )MS S = mine ra l s us pended s o l ids (mg l ~ )r~ = ce l l g row th r a t e ( r ag 1 j h ')S = u s a b l e C O D ( m g 1 ~)So = in i t i a l us ab le C OD (mg 1 ~)t = t ime (h)T O C = t o t a l o r g a n i c c a r b o n ( m g l ' )TSS = to t a l s us p ended s o l ids (mg 1- ' )V S S = v o l a t il e s u s p e n d e d s o l id s ( m g l - ' )X = c el l c o n c e n t r a t i o n (m g 1 - ' )X0 = in i t i a l ce ll con cen t r a t ion (mg I ~)

* A u t h o r t o w h o m a l l c o r r e s p o n d e n c e s h o u l d b e a d d r e s s e d[Fax : +3 4 57 218 606].

Xca~c = ce l l co nce n t r a t ion es t imated by the k ine t i c mo del(rag 1 t)Yx~s = A. terreus g r o w t h y i e l d o n C O D ( m g c e l l s p e r m gC O D )Y~.,s = G. candidum grow th y ield on C OD ( r ag ce ll s per mgC O D )= specif ic grow th ra te (h -~)~Zmax= maximum s pec i f i c g rowth r a t e fo r A. terreus ( h - ' )~t~ax = max im um specif ic g row th rate for G. candidum

(h ' )

INTRODUCTIONT h e b y - p r o d u c t s o f t h e s u g a r i n d u s t r y i n c l u d em o l a s s e s , t h e r e s i d u e r e m a i n i n g a f t e r s u c r o s e h a sb e e n c r y s t a ll i z e d f r o m t h e p r o c e s s b r o t h . B e c a u s em o l a s s es c o n t a i n s r o u g h l y 5 0 % o f f e rm e n t a b l e s u g a r s( su c r os e , g lu c o s e a n d f r u ct o s e; W a y m a n a n d P a r e k h ,1 9 9 0) , it s m o s t f r e q u e n t a p p l i c a t i o n i s as a s u b s t r a t ef o r f e r m e n t a t i o n p r o c e s s e s . T h u s , i t is u s e d t op r o d u c e a v a r i e ty o f c h e m i c a l p r o d u c t s , f o r e m o s ta m o n g w h i c h i s e t h a n o l .T h e a l c o h o l i n d u s t r y u s e s m o l a s s e s t o p r o d u c ee t h a n o l , n o r m a l l y w i t h t h e a i d o f f e r m e n t a t i o n y e a s ts .

wa 3~,,8 G 2005


2/7

2006 I. Garcia Garcia e t a l .O n c e o b t a i n e d , t h e a l c o h o l m u s t b e r e m o v e d f r o m t h ef e r m e n t a t i o n b r o t h , w h i c h i s u su a l l y d o n e b yd i s t i l l a t io n . T h i s o p e r a t io n l e a v e s a r e s id u e k n o w n a sv i n a s s e , c o n s i s t i n g o f w a t e r a n d n o n - v o l a t i l e c o m -p o n e n t s f r o m t h e f e r m e n t a t i o n b r o t h . T a k i n g i n t oa c c o u n t t h a t a l c o h o l i c f e r m e n t a t i o n i s u s u a l lyf a c i l it a t e d b y a l o w s u g a r c o n c e n t r a t i o n ( 1 2 - 2 0 % ) ,t h e a m o u n t o f v i na s s e p r o d u c e d c a n b e u p t o 1 0 t i m e sl a r g e r t h a n t h a t o f e t h a n o l .

T h e c o m p o s i t io n o f t h i s e f f lu e n t v a r i e s w i th th eo r ig in a n d ty p e o f r a w m a te r i a l u s e d ; a s a ru l e i t i sd a r k - c o l o u r e d a n d h a s a t y p i c a l o d o u r , a s t r o n g l ya c id ic c h a ra c te r (p H -~ 3 . 5) a n d a h ig h o rg a n ic m a t t e rc o n te n t (C O D b e tw e e n 1 0 a n d 8 0 g l - l ) , a l l o f w h ic hp o s e s e r i o u s e n v i r o n m e n t a l p r o b l e m s t h a t e n t a i lp u r i f i c a ti o n o f t h e w a s te p r i o r t o d u m p i n g i n t o p u b l i cw a te r w a y s . In f a c t , v in a s s e c a n in c re a s e th et e m p e ra tu re o f t h e r e c e iv in g w a te r , t h e re b y d e c re a s -i n g t h e a m o u n t o f a v a i l a b l e d i s s o l v ed o x y g en . A l s o ,i t s a c id c h a ra c te r d e c re a s e s th e p H o f th e w a te r ,w h ic h m a y d i s s o lv e s o m e m e ta l s i n f lo w in g .

S e d im e n ta ry s u s p e n d e d s o l id s d e c re a s e b e d p e r -m e a b i l i t y , t h u s h in d e r in g f i l t r a t io n a n d fo s t e r in ga n a e r o b i c f e r m e n t a t i o n a n d t h e r e s u l t i n g u n p l e a s a n to d o u r s . A l s o , t h e t u r b i d i t y o f s u s p e n d e d s o l i d sre s t r i c t s l i g h t p e n e t r a t io n .

T h e p r e s e n c e o f s u r f a c t a n t s i m p e d e s o x y g e nt r a n s f e r a c r o ss t h e a i r - w a t e r i n t e r f ac e a n d d i m i n i s he sw a t e r o x y g e n a t i o n .

In s u m m a ry , t h e e n v i ro n m e n ta l e f f ec t s o f v in a s sec a n c a n c e l t h e n a tu ra l s e l f -p u r i f i c a t io n a b i l i t y o f t h ere c e iv in g w a te r s .

T h e r e a r e a n u m b e r o f a v a i l a b le m e t h o d s f o r t h ee x p l o i t a t i o n a n d p u r i f i c a t i o n o f v i n a ss e , p a r t i c u l a r l yb i o l o g i c a l m e t h o d s ( b o t h a e r o b i c a n d a n a e r o b i c ).N o t w i t h s t a n d i n g i t s s h o r t c o m i n g s , a n a e r o b i c d i g e s -t i o n f e a t u r e s s o m e a d v a n t a g e s t h a t m a k e i t p a r t i c u -l a r ly s u i t a b le fo r p ro c e s s in g th i s t y p e o f w a s te .H o w e v e r , i t s e f fi c i en c y i s s o m e w h a t d e c re a s e d b y th ep r e s en c e o f p h e n o l c o m p o u n d s i n v i n a ss e .

A s n o t e d e a r l i e r , v i n a s s e c o n t a i n s a b u n d a n to r g a n i c m a t t e r t h a t i n c lu d e s a n u m b e r o f p h e n o lc o m p o u n d s ( m o r e t h a n 1 0 a c c o r d i n g to M a e s t r o -D u r / m e t a l . , 1 9 9 3 ) a n d th e i r p o ly m e rs . T h e s ec o m p o u n d s a r e k n o w n t o b e d i ff i cu l t t o d e g r a d eb i o l o g i c a l l y a n d p o s s es s a n t i m i c r o b i a l a n d p h y t o t o x i cp r o p e r t i e s t h a t h i n d e r p u r i f i c a t i o n b y a n a e r o b i cd ig e s t io n (S o r l in i e t a l . , 1 9 8 6 ; S ie r r a -A lv a re z e t a l . ,1 9 88 ; R o d r lg u e z e t a l . , 1988; Ba l ice e t a l . , 1990; Borjae t a l . , 1 99 0 ). I t s e e m s th a t t h e p r io r r e m o v a l o f t h e s ec o m p o u n d s s h o u l d a c c e l er a t e a n a e r o b i c d i g e s ti o n o fth e w a s te , w h ic h s h o u ld r e s u l t i n d e c re a s e d c o s t st h r o u g h a s h o r t e n e d w a t e r r e t e n t i o n t i m e .

A p o s s i b i li t y o f r e m o v i n g t h e s e c o m p o u n d s i st h r o u g h a n a e r o b i c b i o l o g i c a l p r e t r e a t m e n t b ym i c r o o r g a n i s m s . A c c o r d i n g t o l i t e r a t u r e t h i s t r e a t -m e n t i s f a c i l i t a t e d b y s o m e fu n g i . T w o fu n g i w h ic hh a v e t h e a b i l i t y t o b i o d e g r a d e p h e n o l i c c o m p o u n d s i nth i s a n d s im i l a r w a s te -w a te r s a re A s p e r g i l l u s t e r r e u sa n d G e o t r i c h u m c a n d i d u m .

T h e p o s s ib i l i t y o f d ig e s t in g th e f e rm e n te d v in a s s eh a s b e e n t e st e d . A s t u d y o n t h e i m p r o v e m e n t o f t h ek in e t i c s o f a n a e ro b ic d ig e s t io n o f v in a s s e p re v io u s lyt r e a t e d b y G . c a n d i d u m h a s a l r e a d y b e e n p u b l i s h e d( B o r j a e t a l . , 1 9 9 3 a ) . A c o m p a ra t iv e k in e t i c s tu d y o ft h e a n a e r o b i c d i g e s t io n o f n a t u r a l v i n a ss e a n d v i n a ss ep r e v i o u s l y t r e a te d w i t h A . t e r r e u s i s c o n t in u in g .T h e i m p r o v e m e n t i n t h e a n a e r o b i c t r e a t m e n t , b yt h e p r e v i o u s r e m o v a l o f p h e n o l i c c o m p o u n d s u s i n ga e r o b i c b i o l o g i c a l p r o c e d u r e s , h a s b e e n s h o w n b ys e v e r a l a u t h o r s ( B o r j a e t a l . , 1992, 199 3b, c, 19 95;H a n d i , 1 9 9 1 ; H a n d i e t a l . , 1 9 9 1 ; Ma e s t ro -D u r~ ine t a l . , 1991 , 1993; Mart in e t a l . , 1 9 92 ; Ma r t in e z e t a l . ,1 9 92 ) w h o w o rk e d o n e i th e r o l iv e -m i l l w a s te -w a te r ( as im i l a r w a s te in m a n y re s p e c t s ) o r v in a s s e . T h ep r e t r e a t m e n t d e c r ea s e s C O D a n d l e a ve s a r e s i d ue ,a n d t h e h i g h o r g a n i c l o a d , l o w e r t o x i c i t y a n dd i m i n i s h e d p h e n o l c o n t e n t w h i c h m a k e i t p o t e n t i a l l ya m e n a b l e t o s u b s e q u e n t a n a e r o b i c t r e a t m e n t .

I n t h e f r a m e w o r k o f b r o a d e r r e s e a r c h d i r e c t edt o w a r d s t h e p u r i f i c a t i o n o f v i n a s s e b y t w o - s t a g et r e a t m e n t ( a n a e r o b i c f e r m e n t a t i o n f o l l o w e d o f a na n a e r o b ic d ig e s t io n ) , t h i s w o rk s tu d ie s e x c lu s iv e ly th ea e r o b i c s t a g e o f tw o f u n g i. T h e s t u d y i s a i m e d a tf i n d i n g t h e m o s t s u i t a b l e m i c r o o r g a n i s m f o r r e m o v -i n g p h e n o l i c c o m p o u n d s .

A s f a r a s i s k n o w n , a s p e c if i c s tu d y o n th e a e ro b icf e r m e n t a t i o n o f v i n a s s e b y A . t e r r e u s a n d G .c a n d i d u m , w i t h t h e a b o v e - m e n t i o n e d p u r p o s e , h a sn o t y e t b e e n c a r r i e d o u t . A m a t h e m a t i c a l m o d e l i su s e d i n o r d e r t o c o m p a r e t h e m i c r o o r g a n i s m sb e h a v i o u r , a n d i t a l lo w s t h e p r e d i c t i o n o f t h e r e m o v a lo f p h e n o l s.

M A T E R I A L S A N D M E T H O D SVinasse

The vinasse used was obtained from Azucarera delGuadalfeo SA, a sugar factory located in Salobrefia,Gra nada, th at produces ethanol from m olasses of differentorigins. Table 1 shows the composition of the vinasse. Rawvinasse, just s terilized, was used. In order to carry out all theexperiments with the same waste, appr opriate amounts ofvinasse, distributed in 250 ml plastic bottles, w as stored a t- 3 0 C .In o c u l a

A. t e rreus was supplied by the Department ofMicrobiology o f the Fac ulty of Pharmacy of the U niversityof Granada. The fungus is known to degrade phenolcompounds in wastes such as olive-mill waste-waterTable 1. Compositionof the vinasse used

Parameter ValueCOD 75,000TOC 32,000pH 4.4TSS I 1,000MSS 2,000VSS 9,000Total nitrogen 975Total phosphorus 20Total phenols 469Total o-diphenols 34


3/7

Biodegradation of phenolic compounds in vinasse 2007Table 2. Experimental results (concentrations n mg 1 ~) obtained in the fermentation process by Aspergil lus terreus

Time(h ) pH TSS MSS VSS COD TOC Total phenols" Total o-diphenols0244874

96120

4.3 11,700 1700 I0,000 83,000 35,000 469 344.5 12,100 ll00 ll ,000 80,0 00 34,000 420 255.0 16,500 1500 15,000 72,000 30,000 285 165.9 19,400 1400 !8,000 63,000 27,000 224 l06.4 20,600 1600 19,000 60,000 26,000 163 56.6 20,500 1500 19,000 59,000 25,000 160 2

dEpressed as caffeic acid.

(P6rez-Torres, 1988; Mart inez e t a l . , 1992; Borja e t a l . ,1993c). G . c a n d i d u m was supplied by the "Instituto de laGrasa y sus Derivados" (CSIC), Sevilla, and it is depositedin the University of Alberta Mold Herbarium collection,Edmonton, Canada, as UAMH 6257.The inocula were prepared in two steps. Firs t, themicroorganism was grown for 48 h in 125 ml o f a syntheticmedium. The compositi on was glucose 40g l -a, peptonefrom casein 5 g I-~ and peptone from meat 5 g I-L Then, itwas adapted to the waste by supplying the medium with anidentical volume of vinasse and extending the incubation at26C with stirring at 200 rpm for a further 48 h.F e r m e n t a t i o n

Fermentat ion runs, repeated three times, were conductedhatchwise in a thoroughly mixed reactor using sterilizedmedia. In practice, vinasse is a sterile waste -it originates inhigh-temperature industrial processes--which facilitatesculturing populations of a single microorganism.The fermentation tank was a Biostat B 5-1 model fromBraun-Biotech SA (Madrid), that allowed automatedregulation of several experimental parameters.The pH was not controlled, since the fermentation values(~.5-6.5) were suitable for effective growth o f themicroorganisms. However, it was monitored throughout thefermentation.The amount of 02 to be suppl ied was delivered as a sterileair stream at a flow-rate of 2 1 rain- ~. The percent saturationof the medium with oxygen was continuously recorded, andit revealed this experimental condition to result in noconstraints at any time. An exhaust cooler prevented lossesof medium.Stirring was done at a constan t speed of 500 rpm and thefermentation temperature was kept at 26C throughout.Finally, foaming was automatically controlled by meansof an internal probe and, whenever needed, by addingDF-7960 anti-foaming agent from Braun-Biotech SA.Previously experiments, using sterilized media butwithout inoculation, were carried out in order to checkwhether oxidation or air -stripping effects occurred; nochanges were observed as a result of such a procedure. Allthe analyses were carried out in triplicate.Total, mineral and volatile suspended solids (TSS, MSSand VSS, respectively), and chemical oxygen demand(COD), were determined according to the recommendat ionsof the American Public Health Association (APHA-AWWA -WPC F, 1985). VSS was determined by thedifference between TSS and MSS.Total organic carbon (TOC) was quantified by using aDohrmann DC-190 carbon analyser. The instrumentoxidizes all C present to CO2 that is subsequently detectedby IR spectroscopy.

A n a l y s e s o f p h e n o l sThe procedure to obtain the phenolic extract was asfollows. The samples were acidified with HC1 (pH 2.0) andextracted three times with ethyl acetate (v/v) at ambient

temperature. The three organic fractions were combined anddried with anhydrous Na2SO4 for 30-40 rain. The extractwas concentrated to dryness in a rotavapour and redissolved

with a methanol/water mixture (60/40). For the determi-nation o f total phenol content (Box, 1983) the method usedthe Folin-Ciocalteau's phenol reagent (from Merck,Darms tadt ) involving the successive addi tion o f 5 mI sodiumcarbonate (200 gl -~) and 2.5ml Folin-Ciocalteau phenolreagent to a 50.0 ml sample. After 60.0 min at 20C theabsorbance was measured at 725 nm against a distilled waterand reagent blank. In the case of the determination ofo-diphenol content, 2 ml 5% molybdate sodium solution inethanol (50%) was added to 10 ml sample. After 15.0 minat 20C the absorbance was measured at 370 nm comparedagainst a reagent blank prepared at the same time using 2 mlethanol (50%) instead of the molybdate sodium solution.

RESULTS AND DISCUSSIONA s p e r g i l l u s t e r r e u s

Table 2 gives the experimental results obtained. Ascan be seen, the procedure used removed about 66%of total phenols and 94% of o-diphenols.

As expected, COD and T OC varied similarly. Theoverall decrease in COD was about 29%.

As can be inferred from the results, thefermentati on ended after 100-120h. Preliminaryexperiments, not reported, suggested that depletionof nitrogen in the medium was the cause of the endof the fermentation.

Since it is desirable that most of the organic loadshould be removed using an anaerobic purificationprocedure (the more affordable choice), the aerobicpretreat ment should, therefore, remove most phenolswithout eliminating much organic matter. The resultsobtained in this respect are quite interesting, since, as

I2,000 --

10,000 f o l

8000 --~"

"~ 6000 --

4000 --perimental values

2000 ~--- ---Jo - - Calculated valuest F I i I 4 I0 20 40 60 80 100 120 140

Time (h)Fig. 1. Experimental and calculated cell concentra tionsduring fermentation by A s p e r g i l l u s t e r r e u s .


4/7

200810 ,000

8 0 0 0

6 0 0 0

i 4000

2000

I. Garcia Garcia e t a l .

o

o E x p e r i m e n t a l s v a l u e sR e g r e s s i o n , Y X / S = 0 .3 8

I I I I I [5000 10 ,000 15 ,000 20 ,000 25 ,000 30 ,000

Consumed COD, (mg/1)Fig. 2. Variation of the microorganism concentration withthe COD uptake during fermentation by A spe rg i l l u s t e r re us .

50O45 04 0 0 o E x p e r i m e n t a l v a l u e s

, 35 0"~ 300

25 020 0

o o15 0 -1 0 0 I I [ I I I I20 40 60 80 100 120 140

Time (h)Fig. 4. Experimental and calculated values of the phenolconcentration during fermentation by A spe rg i l l u s t e r re us .

noted earlier, the original phenol content was reducedsubstantiallywith a decrease in COD of less than 30%.

From the COD reduction, the total phenols ando-diphenols removed and increase in VSS (a measureof cell growth), it can be shown that the extent ofremoval of organic matter and phenols was related tocell growth.

The results were fitted to a Monod equation inorder to establish correlations between the exper-imental variables and to account for the kineticbehaviour of the ferme ntation process, and it allowedcomparisons between both microorganisms, once itwas shown that the same model was applicable.

There follows the development of the mathematicalmodel used to discuss the results as applied to thissituation.

The material balance for microorganisms in abatch reactor can be expressed mathematically as

dX;'x = -dT (1)10 ,000

~d~' 80 00 - o o

6 0 0 0 -.

4 0 0 0

L9 2000 -- R e g r e s s i o n , E = 0 . 0 3 3

) / I I r I I I0 50 100 150 200 250 300 350

Consumed total phenols, (rag/l)Fig. 3. Variation of the concentration of A spe rg i l l u s t e r re usproduced with phenol removal.

where r x denotes cell growth rate (mg 1 ] h t) andd X / d t cell accumulation.

Usually, in a typical batch process, following a lagphase, cells grow in an exponential manne r with time.Unde r such conditions, the growth rate is given by afirst-order kinetic equati on:

r~ = ~ x X (2)where # is the specific growth r a t e ( h - l ) and X themicroorganism concentration (mg l - l ) .

The Monod model assumes that the relationshipbetween the specific growth rate and the substrateconcentration in a batch culture process limited bythe amo unt of available substrate can be expressed as

~max X S# - K s + S (3)

where #ma~ s the maximum specific growth rate ( h - L ) ,S usable COD (mg 1 ]) and K s a constant (rag 1-~).

Therefore, the material balance can be expressed asdX Sd t - I ~ x X : ~ m a X K s @ S ~g f (4)

Integrating equation (4) requires establishing pre-viously the relationship between S and X.

The relationship between the amount of cellsproduced and substrate uptake (i.e. the yield Y x . s ) hasa decisive influence on both the design and economicfeasibility of a microbiological process. It is ameasure of the growth efficiency in the substrate. Foranalytical and design purposes, the yield is assumedto be constant, which, however, should always beconfirmed experimentally.

After the amou nts of cells produced and substrateuptake over a given period are determined, the yieldcan be calculated from

X - X o = G , s ( S o - S ) (5)


5/7

Biodegradation of phenolic compounds in vinasse 2009Table 3. Experimental esults (concentrations n nag I ~) obtained n the fem entation process by Geotrichum candidum

T i m e(h) p H T S S M S S V S S C O D T O C T o t a l p h e n o ls~ T o t a l o - d i p h e n o ls "0224 267

8912 5

4.4 13,000 3000 10,000 83,000 35,000 469 344.9 14,300 3100 11,200 76,000 31,000 428 285.6 20,000 4500 15,500 65,000 27,000 251 126.5 22,500 5000 17,500 61,000 26,000 224 86.8 23,000 4000 I9,000 60,000 26,000 157 46.6 23,000 4000 19,000 60,000 26,000 143 3

~Expressed as caffeic acid.

X o - XS = So + - - (6)r x / ,w h e r e X 0 i s t h e i n i t i a l m i c r o o r g a n i s m c o n c e n t r a t i o n(m g 1 t ) a n d S o th e in i t i a l u s a b le C O D (m g 1-~) .

T h e m a t e r i a l b a l a n c e c a n t h e r e f o r e b e e x p r e s se d a sd X _ p ~ x X ( + X o - Jd t Ks + SO + X o _ X So (7 )

i n t e g r a t i o n o f w h i c h y i e l d sY~/~SO + Y~/~K~ + X o , /~m~ X t = -- - - -- - - =:. InY x / s s o + Xo o

+ Y x / s K s In Yx/ s s o (8 )Y~:/~So + Xo Yx,,~SO+ Xo - XI f t h e g r o w t h m o d e l c o n s i d e r e d i s c o r r e c t , t h e n t h ee x p e r i m e n t a l m i c r o o r g a n i s m c o n c e n t r a t i o n a n d t i m ev Mu e s s h o u ld f i t e q u a t io n (8) . A s c a n b e s e e n f ro mF ig . 1 , t h e e s t im a te d v a lu e s f i t t e d th e e x p e r im e n ta lp o in t s q u i t e w e l l , w h ic h ju s t i f i e s u s in g th e m o d e l t oa c c o u n t f o r t h e g r o w t h r a t e o f t h e f u n g u s i n th ef e r m e n t a t i o n m e d i u m .

T h e f i t t i n g w a s c a r r i e d o u t b y u s i n g S i g m aP lo t S c ie n t if i c G r a p h in g S y s te m v . 5 .0 a n di t e r a ti v e e q u a t i o n p a r a m e t e r v a l u e a d j u s t m e n t , w h i c hy ie ld s5 0 0

4 5 0

~ 4 0 0 . . . . . G e o t r i c h u m

. ~ 3 0 0o 2 5 0E 2 0 0

c~1 5 0 - ' . . . . . . . . . . . . . . . . . .~00 I F I r I I I

2 0 4 0 6 0 8 0 1 0 0 1 2 0 1 4 0Time (h)

Fig. 5. Predicted values of total phenols concentrationduring fermentations by Aspergillus terreus an d Geotr ichumcandidum.

0.38SO + 5139 + X01 n X0.06 x t = 0 .38 SO+ Xo ~oo5139 0.38SO+ 0.38SO + Xo In 0.38SO + Xo - X (9)

S O , l am a = 0 , 0 6 h - l , Ks ~ 1 3 ,5 25 m g C O D 1 ] a n dYxls = 0.38.

I f a p l o t o f t h e c o n c e n t r a t i o n o f m i c r o o r g a n i s m sp r o d u c e d a g a i n s t t h e C O D u p t a k e i s a s t r a i g h t l i n eo f s l o p e c o i n c i d in g w i t h t h e y i e l d v a l u e o b t a i n e d b yf i t t i n g e q u a t io n (8 ) , t h e n e q u a t io n (9 ) a n d th e m o d e lu s e d w i l l b e v a l id a t e d . A s c a n b e s e e n f ro m F ig . 2 ,b o th r e q u i s i t e s w e re m e t .

F i n a l l y , t h e m o d e l a l l o w s o n e t o p r e d i c t t h ev a r i a t i o n o f t h e p h e n o l c o n c e n t r a t i o n t h r o u g h o u tf e r m e n t a t i o n . T h e r e l a t i o n s h i p b e tw e e n c e l l g r o w t ha n d p h e n o l r e m o v a l i s sh o w n i n F i g . 3 a n d a l l o w s t h ef o l l o w i n g e q u a t i o n t o b e f o r m u l a t e d :

1X - X 0 = ~ ( F 0 - F ) ( 10 )w h e r e X a n d F a r e t h e e x p e r im e n t a l c o n c e n t r a t i o n s o fc e l l s a n d to t a l p h e n o l s (b o th in m g 1 -~ ).T h e s lo p e o f F ig . 3 a l lo w s o n e to d e te rm in e th ee f fi c ie n cy w i t h w h i c h t h e m i c r o o r g a n i s m d e g r a d e sp h e n o l c o m p o u n d s ; t h u s , E = 0 .0 3 3 r e p re s e n t s t h em a s s o f p h e n o l c o m p o u n d s t h a t i s r e m o v e d p e r u n i tm a s s o f m i c r o o r g a n i s m p r o d u c e d . S u c h a c o n s t a n ts h o u ld id e a l ly b e q u i t e h ig h .

A f t e r E h a s b e e n d e t e r m i n e d , t h e c o n c e n t r a t i o n o ft o t a l p h e n o l s a g a i n s t t i m e c a n b e p r e d i c t e d f r o m t h efo l lo w in g e x p re s s io n

1X ~ . , o - X 0 = ~ ( F 0 - F ~ , ~ c ) ( 1 1 )

whe re X~,~ and FoaEc re the ca lc u la t ed ce l l and phe nolc o n c e n t ra t io n s ( r a g 1 - ] ).E q u a t i o n ( 1 l ) c a n b e r e w r i t t e n a s

Foa]~ = Fo - E (Xc,,c - 35o) (12 )F r o m e q u a t i o n ( 9 ) i t f o l l o w s t h a t

X,a,, = f(t)F i g u r e 4 s h o w s t h e e x p e r i m e n t a l v a r i a t i o n o f t h e

p h e n o l c o n c e n t r a t i o n w i t h t i m e a n d a p l o t o fe q u a t io n (1 2 ). A s c a n b e s e e n , th e m o d e l p re d ic t s t h ee x p e r im e n ta l r e s u l t s q u i t e a c c e p ta b ly .G e o t r i c h u m c a n d i d u m

T a b le 3 g iv e s th e e x p e r im e n ta l r e s u l t s o b ta in e d fo rt h i s m i c r o o r g a n i s m . A s c a n b e s e e n , a b o u t 7 0 % o f


6/7

2 0 10 I . G a r c i a G a r c i a et al.t o t a l p h e n o l s a n d 9 1 % o f o - d i p h e n o l s a r e r e m o v e d .T h e o v e r a l l d e c re a s e i n C O D i n t hi s c a se w a s a b o u t2 8 % a n d t h e f e r m e n t a t i o n f i n i sh e d s h o r tl y b e f o r e t h a to f A . t e r r e u s .

F o l l o w i n g a p a r a l l e l d i s c u s s io n , i t c a n b e s h o w nt h a t t h e s a m e k i n e t i c m o d e l i s a p p l i c a b l e , a n d t h ef o l l o w i n g v a l u e s c a n b e f o u n d : # ~ x = 0 . 0 4 7 h -~ ,K 2 = 4 5 5 8 m g C O D I -~ a nd E ' = 0 . 0 3 6 m g t ot alp h e n o l s p e r m g c e l l s .

S i m i l a r b e h a v i o u r t o A . t e r r e u s i s o b s e r v e d . N e v e r -t h e l e s s , t h e k i n e t i c m o d e l p r e d i c t i o n s s e e m t o s u g g e s ts l i g h t ly b e t t e r b e h a v i o u r i n t h e c a s e o f G . e a n d i d u m ,e s p e c i a l ly b e c a u s e o f th e d i f f e r e n c e s b e t w e e n t h e v a l u eo f c o n s t a n t K s . I n F i g . 5 , t h e m o d e l p r e d i c t i o n s o f t h ev a r i a t i o n o f t o t a l p h e n o l c o n c e n t r a t i o n t h r o u g h o u tb o t h f e r m e n t a t i o n s a r e s h o w n . A s c a n b e s e e n , t h e G .e a n d i d u m t o t a l p h e n o l s r e m o v i n g r a t e i s s l i g h t l yh i g h e r th a n t h a t o f A . t e r r e u s .

CONCLUSIONST h e r e s u l t s o b t a i n e d i n t h i s w o r k a l l o w t h e

f o l l o w i n g c o n c l u si o n s t o b e d r a w n .( 1 ) T h e f u n g i A . t e r r e u s a n d G . c a n d i d u m c a n

s i g n if i c an t l y d e g r a d e a e r o b i c a ll y p h e n o l c o m p o u n d si n a l c o h o l in d u s t r i a l w a s t e w i t h o u t t h e n e e d f o r a n yn u t r i e n t s u p p l e m e n t s i n t h e m e d i u m .

( 2) T h e e x t e n t o f r e m o v a l o f o r g a n i c m a t t e r a n dp h e n o l c o m p o u n d s i s r e l a te d t o c e l l g r o w t h . S o , t h ep r e d i c t io n o f th e b i o m a s s c o n t e n t a l l o w s t h ed e t e r m i n a t i o n o f th e r e m o v a l o f p h e n o l s .

( 3) T h e m i c r o o r g a n i s m s f o l l o w M o n o d g r o w t hk i n e ti c s w h e n g r o w n w i t h C O D a s t h e s u b st r a te . T h em o d e l a l l o w s c o m p a r i s o n s b e t w e e n b o t h m i c r o o r g a n -i s m s a s w e l l a s t h e p r e d i c t i o n o f r e m o v a l o f p h e n o l s .T h e k i n e ti c c o n s t a n t s a r e # m ~ x = 0 . 0 6 h -~ a n dK s = 1 3 5 2 5 m g l -~ f o r A . t e r r e u s a n d # n ' ~ x =0 . 0 4 7 h - ~ a n d K ~ = 4 5 5 8 r a g 1 ~ f o r G . e a n d i d u m .

( 4 ) T h e c e l l y i e l d s re l a t i v e t o t h e s u b s t r a t e a r eYx, s = 0 .3 8 m g c e ll s p e r m g C O D f o r A . t e r r e u s a n dY'xls = 0 . 39 m g c e ll s p e r m g C O D f o r G . c a n d i d u m .

( 5 ) T h e e f f i c i e n c i e s w i t h w h i c h A . t e r r e u s a n d G .c a n d i d u m d e g r a d e p h e n o l i c c o m p o u n d s a r e E = 0 .0 3 3a n d E ' = 0 . 0 3 6 r a g t o t al p h e n o l s p e r m g c e ll ,r e s p e c t i v e l y .( 6) T h o u g h b o t h m i c r o o r g a n i s m s b e h a v e i n as i m i l a r w a y , t h e k i n e t i c m o d e l s e e m s t o s u g g e s ts l i g h t ly b e t t e r b e h a v i o u r i n t h e c a s e o f G . c a n d i d u m .A c k n o w l e d g e m e n t s - - T h e authors wish to express theirg ra t i tude to the Comis i6n In te rmin is te r ia l de C ienc ia yTecno log la (p ro jec t AM B--746 /93) as we ll a s the Conse je r iade Educac i6n y C ienc ia de la Jun ta de Anda luc ia fo rfinancial sup port gra nted for the realization of this work.Va luab le he lp f rom the Azucare ra de l Guada l feo (Sa lo -brefia , Spain) is also gratefully acknowledged.

REFERENCESA P H A - A W W A - W P C F ( 1 9 8 5 ) S t a n d a r d M e t h o d s f o r t heE x a m i n a t i o n o f W a t e r a n d W a s t e w a t e r , 16th edn,

Amer ican Pub l ic Hea l th Assoc ia t ion , Wash ing ton ,D C .Balice C. , Carrieri C. and Ce ra O. (1990 ) Caratte rist icheana l i t ic i de l le acqu e d i vege taz ione (Ana ly t ica l fea tu re o fvege ta t ion wa te r o f o l ive ) Riv . I tal . Sostanze Grasse 67 ,9 -16 .Bor ja R . , Mar t in A. , F ies tas J . A. and Maes t ro R . (1990)S tudy o f the inh ib i t ion o f o l ive mi l l was tewate rb iomethana t ion in b io reac to rs wi th mic roorgan ismsimm obi l ized on va r ious types o f suppor t . Grasay Aceites41, 397-403.Bor ja R . , Mar t in A. , Maes t ro R . , Alba J . and F ies tas J . A.(1992) Enhancement o f the anaerob ic d iges t ion o f o l ivemi l l was tewate r by the remova l o f pheno l ic inh ib i to rs .Proc e ss B ioc he m. 27, 231-237.B o r j a R. , M a r t i n A . , M a e s t r o R , L u q u e M . a n d D u r f i n M .M. (1993a) Improvement o f the k ine t ic s o f anaerob icd iges t ion o f v inasse by the remova l o f pheno l icc o m p o u n d s . Biotechnol. Lett. 15, 311-316.B o r j a R ., M a r t i n A . , G 6 m e z L . F . a n d R a m o s - C o r m e n z a n aA. (1993b) Ana erob ic d iges t ion o f o l ive mi l l was tewate rp re t read wi th Azo toba cter chroococcum. Resources ,Conserv . Recycl . 9, 201-211.

Bor ja R . , Gar r ido S . E . , Mar t inez L . , Ramos-ComenzanaA. and Ma r t in A . (1993c) Kine t ic s tudy o f anaerob icd iges t ion o f o l ive mi l l was te -wa te r p rev ious ly fe rmen tedwi th Aspergil lus terreus . Process Bioehem. 28 ,397-404.Bor ja R . , Mar t in A. , Alonso V. , Garc ia I . and Banks C . J .(1995) Influence of different aerobic pretreatments on thekinetics of anaerobic digestion of olive mill wastewater.Wat . Re s . 29, 489-495.Box J . D. (1983) Inves t iga t ion o f the Fo l in -C ioca l teaupheno l reagen t fo r the de te rmina t ion o f po lypheno l icsubstances in natural waters. W a t . R e s . 17, 511-525.Han d i M . (1991) E ffects o f ag i ta t ion and p re t rea tmen t onthe batch anae robic digestion of olive mill waste-water.Bioresource Teehnot. 36, 173-178.Han d i M. and Garc ia J . L . (1991) Com par ison be tweenanaerobic f i l ter and anaerobic contact process forfermented olive waste-waters. Bioresource Technol. 38 ,23-29.Maes t ro -Dur~ in R . , Bor ja -Pad i l la R . , Mar t ln -Mar t in A. ,F ies tas J . A. and Alba J . (1991) B iodegradac i6n de loscompuestos fen61icos presentes en el alpechin (Biodegra-da t ion o f pheno l ic com pounds in o l ive mi l l was te wa te r) .Grasas Aeeites 42, 271-276.Maestro-Dur~in R. , Borja -Padilla R. , Luque-G onzfilez M.and Ma r t in -Ma r t in A . (1993) Compu es tos fen61icos en la saguas residuales de las destilerias vinicas (vinazas)(Phenolic c om pou nds in waste wa ter from wine disti lleries(vinasse). Rev. Espa~ola Ciencia Tecnol . Alimentos 35 ,517-528.Mar t in A. , Bor ja R~ and Ch ica A. (1992) Kine t ic s tudy o fand anaerobic f luidized bed system used for thepur i f ica t ion o f fe rm en ted o l ive mi l l was tewate r . J . Che m.Tech. Biotechnol . 53, 48-92.M a r t l n e z L . , R a m o s - C o m e n z a n a A . , G a r c i a M . P .a n d G a r r i d o S . E . ( 1 9 9 2 ) B i o d e g ra d a c i6 n d ecompuestos fen61icos del aIpechin con Aspergil lus terreus(B iodegrada t ion o f pheno l ic com pounds in o l ive mi l lwas te wa te r by Aspergillus terreus). Grasas Aceites 43 .75-81W ayma n M . and Parekh S . R . (1990) B io techno logy o fbiomass conversion, Biotech. Series 21, Open UniversityPress, Milton Keynes.P6rez -Torres J . (1 98 8) T ransfo rmac idn mic rob ian ade componen tes a rom/ t t icos de l a lpech in (Microb ia lt r a n s f o r m a t i o n o f a r o m a t i c c o m p o u n d s i n o l i v emill waste water) , PhD thesis , De parta me nto deMicrob io log ia , Facu l tad de Farmac ia , U n ivers i ty o fG r a n a d a .Rodr iguez M. , P6rez J . , Ramos-Cormazana A. and


7/7

Biodegrada t ion o f pheno l ic compounds in v inasse 2011Mar t inez J . (1988) E f fec t o f ex t rac ts ob ta ined f rom o l ivemi l l was te -wa te r on Bac i l lum Mega te r ium ATTC 33085 ,J. Appl. Bacteriol. 64, 219-226.S ie r ra -Alvarez R ., F ie ld J . A. and Le t t inga G . (1988)Tox ic idad metanogen ica de compues tos fend l icosy l ip id icos (methanogen ic tox ic i ty o f pheno l ic and l ip idiccompounds), pp. 83-110. In Actas del 4 Seminario deDepuracidn Anaerobia de Aguas Residuates (Proceedingsof the 4th Seminar on Anaerobic Waste-water Treatment)

(Ed i ted by Fdz . -Po lanco F . , Garc ia P . A. and HernandoS.) . University of Valladolid.Sor l in i C . , Andreon i V. , Fe r ra r i A. and Rana l l i G. (1986)The influence of some phenolic acids present in oil millwa te r on mic rob ic g roups fo r the methanogenes is ,pp. 81-88. In International Symposium on Ol ive By-products Volarization, Sev i l la . Food and Agr icu l tu reO r g a n i z a t io n o f t h e U n i t e d N a t i o n s, R o m e .

Documents

biodegradacion de fenoles.pdf