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5 !&'!! " " "! $ " % "! & " " $ "! $ " '()* +,-. */ & 0*1* '%1819: ;!))+.8< =/ )$ ><?@) *' )"'8!!!?8 9768" " " A >-*!!"! v

6 = B"" "&"""%"?" )")C" D" 0" )"'"'" " EF"*!! " " *")""G " H"! " * " C"! "! "" " vi

7 I =,J " K $", M 0,-,-,-,-=C 0 &ML"K < & DN:9;2" ",-,-,- CM@ &".,- ", G 6O " 6O "D 6O "P 6O 9O H,- = " =<,-@ J "@ = < 25 Q " &,- <,- ",- ) " =,J $,-" I L. FG """"H" ".,- vii

8 $ =,- 9O < M GDH << =,- D" C "".,-,- ",- "MG"DPHG" " D C G<HC ",J $ T,JU4R"R:"R6R2S D""P" D " < B,- ;O 9O "&%;"7 < " ;O %6"9GR4SH %6"9TM&%;"7 < " = L T " " L" C = " C$ L",- = =0C ",-"C$,- 9O ;O,J,-= C -"M,-,- C" = viii

9 !( $" $* M * M " " " " DN:9;2" " M " "G 6O " 6O "D 6O "P 6O 9O H! = "! "! = M! 25 Q " $ "?" = $ " G " " " " H" " $ 9O ix

10 'MGDH * M D " M " *! 4RS " G"DPHG""D PH " %;7 M GM! H " TV4R"R:"R6R2S D""P" " B ;O 9O "%&;7 " ;O GR4SH % 69T " % ;7 " T " " "" D!= " $! " ""$ 9O ;O! " ""!= x

11 ))? < W <$. <" < < $< <$ " W. < < W? M $ <<? X <$ <. W < << << <<$W?W & X < W <"KW< "W "<<<<$W " W X DN:9;2" " <$ < M" < &"W "<<<$ G 6O " 6O "D 6O "P 6O 9O H< <"<<<<" < < W< << "!=K25Q" < < << & W< < < W?< << < W <$ < < MW <" < < << X K $<K <" < < <$ < < < < W G " " " <" H". < xi

12 <" < < <$ < $<<<K 9O <<M<<GDH <?<< <<DK <<<"XX W W < " K 4RS! " $ <<<G"DPHG""DPH < W W %;7 << G H X X " <$ X K < $ < <" U 4R"R:" R6 R2S D" " P" " $ <<<B< ;O K 9O "&%K;"7 < " ;O,<<K%6"9GR4SH K%6"9" W&%K;"7< " <$ << <<WX<M"<$ < <<<" <" <? <<<" <" < K Y<?! = <<<" $ < < <.<.<? "<"X $ 9O K ;O?<. < " < << <<<""!=W xii

13 "!$" " " " = " *! " M * = :7 " T W D$ " " " 6" " " $T "! G"! =H 9 xiii

14 23= * $ " = " "$2 5" ;3" DN:9;2 " ""!= "! G;H "! $ G3H * 4T " " " 4G"! =H R G"! H "!:7! xiv

15 !&'!( * )) * * * + ** %( ***!(( *** $ $, - $ $,$,.' $ $,", / $,",$, / $,",$,$, 8 / $,",$,$,$, 0 $,",$,$,", 1 $,",$,", 8 2 $,",$,",$, 2 $,",/, 3 $,",0, 4 $,",1, $ $,",2, $0 $,",3, $1 $,",3,$, 8 = $1 $,",3,", $2 $,",5, $3 $,",4, $5 $,",$! " xv

16 $,/, "$ "/ " ", ('6 7 //!( // ",$, - /0 ",", /3 ",",$, " /3 ",",", /3 ",",/, /5 ",",0, $ /4 ",",1, /4 ",",2,! 0 ",/, 0$ ",/,$, 0$ ",/,", 0" ",/,/, 00 ",/,0, % 01 ",/,1,! 04 ",0, 1 1" / /, ('6 7 13!( 13 /,$, - 15 /,", 2 xvi

17 /,",$, " 2 /,",", 2 /,",/,& 2 /,",0, 2$ /,",0,$, Z 2$ /,",0,", 2" /,",0,/, 2" /,",1, '( 2" /,",2, $ 2/ /,",3, 2/ /,",5, 20 /,",4,) 20 /,",$, * 21 /,/, 21 /,/,$, + 21 /,/,",, 24 /,/,/, $ 3$ /,/,0,) '( "- '(. '( 31 /,0, 32 5" 0 0, ('8 7 53!( 53 0,$, ,", 4 0,",$, " 4 0,",", 4$ 0,",/, & 4$ 0,",0, ) / 4$ xvii

18 0,",1, 4/ 0,",2, 4/ 0,",3,! 40 0,",5, * 40 0,/, 41 0,/,$, 0 / 41 0,/,", 0/ 43 0,/,/,! $1 0,0, $5 $$ 1 1, - ('6 $$1!( $$1 1,$, - $$1 1,", $$3 1,",$, " $$3 1,",", $$3 1,",/,, $$5 1,",0, $$5 1,",1,! $$5 1,",2, * $$4 1,/, $$4 1,0, $"1 $"2 2 2, ('8 7 $"4 xviii

19 !( $"4 2,$, - $/ 2,", $/" 2,",$, " $/" 2,",", $/" 2,",/, & $/" 2,",0, / $/" 2,",1, $/0 2,",2, $/0 2,",3, * $/1 2,/, $/1 2,/,$, / $/1 2,/,", 1 $/3 2,0, $0" 2,1, $05 $04 3 3, & ('8 $1/!( $1/ 3,$, - $1/ 3,", $10 3,",$, " $10 3,",", $11 3,",/,, $11 3,",0, $11 3,",1,! 0 $12 3,/, $12 3,0, $2$ $2" xix

20 5 5, & 9 ( $21!( $21 5,$, - $21 5,", $23 5,",$, ) $23 5,",", $25 5,",/, $25 5,",0, $24 5,",1, % //234 $24 5,/, $3 5,/,$, ) $3$ 5,/,", $ $3" 5,/,/, "% / $30 5,/,/,$, $30 5,/,",",!= $33 5,/,0, "% / $5/ 5,0, $51 $52 4 4, & 9 : $54!( $54 4,$, - $4 4,", $4$ 4,",$, ) $4$ 4,",", $4" 4,",/, $4" xx

21 4,",0, -% /234 5( 6( 4,",1, $40 4,/, $40 4,0, " "$ $4/ $ $, '& "1 $,$, "1 $,", +'& "5!;;<!*!!:$7 DN:9;2!:"7 DN:9;2!:/725Q DN :9;2!* = =:$7!* :$7 &%; xxi

22 xxii

23 + +",$, DN :9;2GHM G8H +",", ;2;M:8GH"DN::R7G8H" DN::R5 GH DN:9;2 GH N G5 1H BG[H:7)) "%;7" 2)6OGHTG\H 69) 6O ":5)D 6O 74)P 6O "& %;7G]H&%;72) 6O E $TG ^;H +",/,* DN:9;2 5 1 )% G:7 )"%;7H"76)BG\H6 O OD 6O TG[H 6O OP 6O T G_H 6O O 6O TG]H 6O O 9O TG`H 6O GHE $TG ^;H + ",0, DN:9;2G\H765"G]H75"G[H:7"G`H67"G_H57GaH :77 1" :7)) "%;7" 2) 6O E $T G^;H +",1,!N G5 1H DN:9;2 :7)) "%;7" 2) 6O BG]HGH G_H25Q"GaH55QG\H47QE $TG^;H +",2, M! DN:9;2b T8"b25"5547Q" +",3,! DN:9;2 E $ $"977G25"5547 QH +",5, DN:9;2" 47 Q A M GH"G8H MGH +",4, S! DN :9;2 :6 1 /1 0" 0/ xxiii

24 661D 6O "577!! :57"65Q97 " " E $T G^2H +/,$, 6O GH"D 6O G8HP 6O GH G]HG\H DN:9;2 :7) ) %;721 T 5 1 $TG V2H +/,",D 6O GHP 6O G8H G]HG\H DN:9;2 :7) ) % ;7 2 1 T :77 1 $TG V2H +/,/, 6O GH"D 6O G8HP 6O GHG]HG\H DN:9;2 :7) ) %;7 2 1 * : $T GV2H +/,0, DN:9;2M! 25Q GHM G8H +/,1, DN:9;2M!25Q )6' GHT B M G8H +/,2, DN:9;2? GH M!25QG8H B +/,3,' DN:9;2 25QGHGHTG8H +/,5, ' DN:9;2 $ GHGHTP 6O G8H"D 6O GH 6O $GH :7) ) %;7 61 * B 666 6O 1"969D 6O 16;6P 6O 1 ;765Q + 0,$, M ) B $ " 3" 3/ xxiv

25 ) O 8GHB8" $ N G:H* G 6 M;H + 0,",! :51 DN :9;2!G HT! G HT! G! H*! G)D(?OHT +0,/,! M:5 1 T8b:6 1 D!G HT! DN:9;2 G HT! G! H! 26!B671G92µΜH + 0,0,! DN:9;2 $8 B T B : G HT =B 6 G HT! B * 26 $ :6 1 * 29*!! :57 " 65 Q 97 " " E $G^;H, + 1,$, S M DN:9;2 )B 6O G HTD 6O G =HT 9O G! H8 :41 "%;7*!!:57"65Q97 " " E $TG^2H +1,", 9O " 6O D 6O G!H G=HG HT G HT G! H* G)D(?OHT 562 $" $0 $3 $"/ $"0 xxv

26 + 2,$ :41!bT=b8) G HT G! HT G H G)D(?OH + 2,", $ DN:9;2 $bt8b 8*B T$B! * " ;65$ :4 1 E $G^;H +3,$, M DN:9;2 )B G HT! G! HT G HT GEH G = H 8 :4 1 "%69G H%;7G H E $T G^2H +3,", 95$:7 M2 1 ;O % * ;O G)D(?OHB0 2 6MG]H" MG\H"%0 2 MG[H" % 60 2GaH +5,$,:5 W" " 557Q +5,", * % &6 M7;E1c1GZ9)Hd"95QGH27QG8H +5,/,!=% 641!GH291=G8H * G)D(?OH B") 6O GHG`H")G0%H 6GHG_H)G0%H 9 M GHG\H )G0%H 2 6M GHG]H $/4 $0" $14 $2 $3" $31 $34 + 5,0,! $5 xxvi

27 $ G H" G H" = $ G = H $G! H"!$ +5,1,=$ %:7G H" G H"! $ G = H $G! H"=$ +5,2,! "! = + 4,$,!* ""!= +4,", 641DGH"761PG8H 941 9O GH ;O GH"%* G)D(?OH DPB") 6O GH G`H")G0%H 6GHG_H")G0%H 9 M GHG\H)G0%H 2 6M GHG]H 9O B 9O GHG`H" G0%H 6O GHG\H"G0%H 9GHG_HG0%H 2 M GHG]H ;O B0 2 6M GHG`H" M GHG_H%0 2 M GHG\H $5" $50 $41 $45 xxvii

28 xxviii

29 %( %(",$,8 04 %(/,$,? 6O "D 6O P 6O 3$ %(/,",! 5 %( 0,$, M = 42 * GM:R42HGM69;1R4H %( 0,", " Z" 44 "!="%;7"%57e %(0,/,8 DN $2 :9;2" * GM:R42H GM 69;1R4H %(1,$, $"$ DN:9;2 %(1,", $"" %(2,$,! $/2 G% & ;7H* GM":R42H GM 69;1R4H %( 2,", 8 DN:9;2" $0$ * GM:R42H GM 69;1R4H %(3,$,8 DN:9;2" $15 %(5,$,'!$ $5 %(5,",'=$ $5$ %( 5,/,! $ %(5,0, =$ $5/ $5/ xxix

30 %(4,$,)"!$ $42 xxx

31 !(( $> GH? S!!!!:+! = (?G H %! % DEAE ;!! %! )! $ '$ +%- - - M A G H B B 1!57S $ B )M$ 6MGDMH ;C D ;%! D = * )$! ', % * %. * :! M E$ C N$$ xxxi

32 xxxii

33 $ - $,$,.'!"T 8$! G"% H" Gf 677;H" " M %G!H " = * B H G"P"" """)"DH GZ!677;HT H G""""D"" ""HG6772HT H G"HG8 6777HT H G"")"PHGE!677:HT H G "HG)! 677RHT H G """P"D"HGf 677:H $ ""M " B 1

34 3) G1=HG$" H* G1=H G $ H Gf 677;H * M GD&E!67:7H $ M * G" "H Gf 677RH " "$ " 6O " 6O " 6O "% 6O " ;O " 9O " 6O " 6O "D 6O " 6O P 6O GM& :RRRT 6776T A 6779T g= g= 6779T E 6779T = 6772T% 677;T) 6773T6774T& 677RT f 67:7H" " & & T $ G :R4:T f :RR;T = 6772HT " 6

35 3) " % D " BGH"GH"GH= GH * $,", :6: ) B M G H " " MG H :6::8 * "! " G H! G8! :RR5H% G 5M97 QHT " 9

36 3)! G8! :RR5HGM H" ""! % " " " * $M 9 :6::: * ":5M65S " $ G47M R7SH cαm G H βmd" G5M:7SHG:M6SHGZ 6776H " " = = M G:RR2H = * βm βm βg:h 9HβG:h;HM! T βg:h 2HM!DM GZ 6776H G 677:H* αm GH" 97M57 S G 677:T? 8 677;HT! $ M "! 2

37 3) :6::6 * " " ""G8 :RR2H*! " $ $ 6O " 6O G8:RR2HfG6776H% G677;H $!!"? G6775H $ $ 9O $" G 6773H"" $""""! 8! $""$ T "" GA:RR7T E!:RR7H G6774H $ G H T " f G H $ G 6773H D 6O ;O Gg= g=6779h 5

38 3) N $ GZ O "D O ") 6O " 6O HfG6773H = " $" = 0=" G8 :RR2T > 6774T 677RH ) T " ZG6779H * $ :6:68 8 " " G8! :RR5H8 GD Z :R33T 8M :R4:H " GDZ:R33T* :RR6H :6:6: " ;

39 3) " " % O M* G8M :R4:T A:RR9H) GD Z :R33T * :RR6T 8! :RR5HT " " "T Gf A:R43H :69 ) %" " % G '$:RR6H %" $ " G8! :RR5H $" %G D 6773H * 6M %0 2M %G6H0 2 %G 3H0" %G%i9H" " % O GD6773H" ;O Gg= g=6779h ) T " :62 3

40 3) * G* *:RR2T 8! :RR5H = *$ G:RRRH $ " " %M A " " T = G H" $ GA 677RH A G6779H G ;O 6O H " " " " "!T! * T G:RR;H "!! " " " GA:RR7T * :RR6T? f!:rr;t!= :RR3T T Aj! 6775H $ GA:RR7H * 8 Gf 677;H 4

41 3) * 8 G:RR2 H 5M27Q :62 ) GH $ G* 6777H* " " %" $ G H" 1 " G% :RR:H % " $ % " % $" GA:RR9T 8! :RR5H'$ T$"$ ;O 9O *G """H $ G%:RR:T* :RR9TA :RR9H $" $ G:R46H 0 G:R46H $ 8" R

42 3) " G:69H ) = $GH G""%H" " & $ " 2 :65 " % "!? " 0 "!8MM*G8*H"&! " $ " " T "! " "? :7

43 3) 8* " " "? " GE!6779 H *?B,9, :;3(,< c:m:d,!g1 H", $!G1 H" G HG1H " *=c:m:d B 3:,93:, (=3:;, <>;3:,< c:m6d *!;3:,< ;3:,<, c:m6d *c:m9db,9% 3: c:m9d ="c:m2d" B,9%(;3:<, c:m2d ::

44 3)?",,!;%<;3:< T,,? )$! =, GH G H? 8*? c:m5db,9,, :=; 0,<=3(;03<,: >>c:m5d G1HT,"",,? *=c:m5d B,:=; 0,<,>93:;, < (=; 03<:, > ;,: < c:m;d *,:=; 0,<,>,:, c:m;d '= = M" :6

45 3) $ :B:" "c:m3db (c:m3d "M $ "+ B +9=>:=>=>c:M4d =>"=>=> M$" G H $ "'="c:mrd c:m:7d" '= B =>:=>93:;+=><(;3:=><=>c:MRd *=>:=>=>" =>+ B =>:=>9+=>0+=>c:M:7d * =>:=> => " +=> :9

46 3) * " +" G H= Z T "! 9 G 92"* 96H G%H! :6; * $ " " $ % GZE:RR5H" " G8A 6775H? T "G :RR4H% " " " G':R33T :RR;T 8! * :RRRH :2

47 3) ="G H* " G*=:RR7H"!1 M & * $ M" G HGf 677;H*" " :63 * " ="T" "" G*=:RR7H :63:8 = * = *" = f G:RR5H " =" "! =!M PG:RR3H"= M! :5

48 3) k = $!G*6777H )" = M G 67:7H = $ = D" =B " " %" G :RR;Tf 677RH :636 GH *" T GE 6779H* " $ "! " $B G" H" G"%"$HGE 6779H " GH" = G H G)! :R46H* :;

49 3) *" " " M G* :RR6T=*$:RRRT) :RRRT) 6777T= 6772HG6776H!M T M " " k "! T :64 8 *! " " G! 67:7H "!"M! "= " " M M G*=:RR7TE!:RR7T* :RR2TA6772T 6774H :3

50 3) " " G! H M G " " " H" ) " G H! GE!:RR7T * :RR2H " B 1 T" " GE! 6779H! M % " "! G! 67:7H 8 " M! "! " G! 67:7H $GE!677:H :6R 8 " " M :4

51 3) G $"! H% " " G:RR;HG 6O " ;O " 6O "D 6O P 6O H = M """DP M "! G:RR3H 8 ;O G$ " $ 7 H ;O GZ 6774H ) 6O " 6O " 6O " 6O " MG' 67:7H 0= G6773H "! "%" =!*% G 6773 H8=M G D6773H :R

52 3) *!! " " " $ $ :6:7! 0 * " " " " G H ' M * G$ H" G 1 H G%" %D0 9 % H" GD0%"D HG7:)H$ 67

53 3) G*H G8 6779T= 6772T6774T 67:7H D" M $,/, % M% " G HG*! 6779T! 6772T*! 6774TZ 67:7T 67:7H* "! =! G6772H *! G6774H '" G67:7H"!= % RR5 4:2S =" "! Z G67:7H "!" = $ "R;;R:5SG%:5HPG%25H 6:

54 3) "$";2S!$ % " $ " " $ G% 677;H D 66

55 3) M&")" M?")"MD"8!"'G:RRRH8 = M?-0 "05":93M:27!="'"A "?N"G:RR3H=M!! 11":M:7 "E* ">)G:RR6H)! $15"9449M 944R " E * " >) G:RR9H) M $14GRH"645:M645; 8"' "* G6779H GEH M =! 53G:H":3M6; 8" " "%" 8" N Z" Z=" "!" Z!" 8 G6772H' M! 40"625M62R 8" D' A G6775H M B?)* 20G6H"R9M:77 8! "Z>""* ">)G:RR5H)! B $! 0/"53RM542 8! " Z > * " >) G:RRRH?)! "/"672M674 8M " A f % G:R4:H M* N!! $ 21G6M9H"44M:63 69

56 3) 8")" 8&" > "' G6777H M 1'1R61:67* 0$D0= 8" " >' G:RR2H 8 / $2";99M;94 8" " > ' G:RR2 H 8 M $! 0$G:H":2RM:52 8"">'G:RR2HM! $2G5H"529M524 8"""">'G:RR2H=$!! 00"6R3M976 " "'" " ")" "? 8 E" G677:H*?! "32":R;3RM:R;46 " >" f G6774H = )Mll$! 34"6R9M6RR ") 8"?"% *" >* G:RR;H = B?)$2"3RM:7: "f">?g6773hp 6O! "G;H"234M246 "f ">?G67:7H'?0$$/2G:H"R5M:76 "%Z"NG6779H= 6O *!!"1G5H"63:M633 &""")"""">E"EG677RH8 GH $0 $//1G:M9H":4:M:44 62

57 3) "?P"f "A>"*G67:7H! k? 55G:H":7RM::5 ">"f"p"p"(">"n"f"n>?"fag6774h ' - $5G2H":774M:7:9 " D" E"' Z!" G6774H 8 M )?! 3G6H":5RM:;R "?)"">"8"l"N"D?""%P"NG677RH ;))< '88@) ) $ "A)" 3:6M3:5 =""*"**$">G6772HGH?$1"::M67 =" *$" > G:RRRH* k GH GH! "$G5H" 29:M293 "*) G:R46H $ " " = $$$0$"39MR2 " '$" > G:RR6H%M M M%$! /3G9H"9RRM 279!"*" "*" " Z" Z&" N")"% " G6772H "PD ++*/G6H"663M696 A"A)G:RR7H * "%?8"?62RM635D N!B)A M% A"A)G:RR9H $- $"0G:H"65M;7 65

58 3) A"A)G677RH8B "?!50G:H" :9M64 A" ) G6772H ' 0 D/E"6:RM696 Aj!" N" m" An," G6775H 8!k! 42":79M :7R A" D" >" 8&" G6779H $* 3G6H"9::M9:R %"'"?"%Z"?"N%"P">%"l"%>">G677;H 8? 2/ $/3G9H":5;RM:53; %")D"'ZG:RR:H) M M GH?B $/3"365M392 %">?""A="G677;H8 " 2 5/G:M2H":7;M::9 Z"E"*G:RR5H M B! 1/G9H" :R5M67; Z" )")" "% " Z 8" G6776H * "2G9H"69RM65; Z""!"*"Z "))"%G67:7H'"P"D $ )?"G6H";6M;; Z" '" 8" D '" A 8" Z G6774H 8 GEH?$/1G9H"676M674 6;

59 3) Z!"DG677;H C '886" 0 6;3M63;! EB )o)?"a8"%g677;h!* 39:3M929?"PN"f ">)"l"'n"ng6775h= 9O $ $0! 2$G2H"3;:M3;5?" N f!" G:RR;H% M =?2/04":;5M:3R )!")" D" > %"% G677RH' k? )? 2G6H":49M:R7 )"""%)'") G6773HGH $M & "$0G:M9H"929M95: )" " "%)"*$" > '") G:RRRH' 6O " 6O 6O B% & $"0":93M:22 )""'")"%)G6777H% 6O " 6O 6O! "/":95M:2: )!"8?""D%">""E?G:R46H D3?! $1"434M44R D&"A)E!"8G67:7H 0 1 2%2 f "?Z"" >"%"N*"DZ623M6R:8'"?B* o' 63

60 3) D" ' Z" G:R33H?B 44"9:3M962 g="g="g6779h GH"DGH" GEH B?2/!$"6:RM66R " E G6774H 8!GH!k B Z"! 44G4H"9:77M9:7R!" " N" N!" >) G67:7H* " "!! $1G:H"4;M:76 " Z D"' G6773H 8 M 0"G9H";65M;94 " Z" D"' D!"' G6773H? M"?! $G:H"R6M:75 " Z" D"' D!"' G6773 H %? )* 22G4H";35M;3R "(("?"NA"P"A)"l"f%"N"P">>G67:7H 8 GH = M? 2/ $33G:M9H";3;M;46 '" G:R33H! $!24"33M4: '"?""")")"""G67:7H ' p!!=p $! 51G9H"3;9M33: ") G6772H +%0D 347M344>f o" 64

61 3) ""G:RR;H $!01"634M645 "E"!"A"""%G6776H 6O B! "0G4H";;9M;;; " Z")" D" Z" " Z ")")"%!"* G67:7H " =! )?"G6H"R9MR3 "">'G:RR;H "! $5" :67RM:6:6 " " >' G:RR3H M 43G9H"623M65: "Af"">">'G:R4:H) B $0$"693M625 ")G:RR2H! M M $G:9H":32:M:356 ">%"N">fZ"G:RR4H 6O $! "G9H"623M65: *!"%%""'"8!">"Z "ZA"'G6779H B: )! $0G;H"269M29; *" )" " " "'" "? A")"%" E? "%?!"' G6777H A " " G' 6777H $ 3"G4H":259M:237 6R

62 3) *" N " A G6777H = $?!31"52:M52; *"N*"fZG:RR2H!= M M M! 1G6H"::9M ::3 * ">)"f"a"a)g:rr2h) M?)$/G6H":6;M:97 *!"%"Z"")"D"Z ")")"%!"*G6774H! " P" D %6 M M 3/GRH":224M :256 *=") G:RR7H "%?8"?965M99RB)A M% E"""E"G6779HZ 8!k! 54G9H"64:M643 E" Z >"!$" A" E"% $" ' G6779H N B! $! 2$":R3M675 E!"8G:RR7H!! E!"8 9M58'"?B' E!"8G677:H$M B $ 2 14"679M6:; E!"8G6779H! E!"8 :3RM:R7)M? "( "B8E $" E!" 8 G6779 H, E!"8:79M:67)M? "( "B8E $" f "%? " G677;H 8 B! $"0G5H"263M25: f " >? G6776H 8 GH! /3G4H"423M457 97

63 3) f " >? " G677RH 8! $"3G6H":R5M66; f ""!"f>ea">g677:h *?"5G9H"92RM952 f " A" A) G:R43H*! M =?B $//"363M393 f "8">'G:RR5H)! $3GRH":773M:7:6 f " 8 " >' G:RR;H' M! $5G5H"59:M59; P") " >' G:RR3H M!! $4G:7H"R59MR55 9:

64

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66 '2 1 ",$, -? * " M * G""$H $T! M GE!:RR7TE!677:H * GE!:RR7TEE!6777TE!677:H $ N $ G H" $ G8! :RR5H*" $ G*=:RR7HM $ " "! " M = "! "! = " G H" 92

67 '2 1 G!H G H GD k:rr2t* 6777T 8L! 677:T A& 6772H 8 =" B" %" ="! =G*=:RR7T :RR;H"! GP ) :RR3T?= 6772H* % G H G:RR6H*" "" M G6:H A B +",$, DN:9;2G!H M G=H 95

68 '2 1 M! " GH" " GH T G)! :R46T:RR6HN GE 6779H '" 6O! G$!D3HM G 6776H! 6O " M M G 6776H A " " $! T " " GE! :RR9TE!)M:RR5T :RR;T8! * :RRRT? 677;H" G* :RR6T=*$:RRRT) :RRRT6777T= 6772H"$!! " "! " $ 9;

69 '2 1 ",", 66:"!* M D N GDNH::R7" DN::R5T DN:9;2 ;2;M:8 G)* 1α" 2D:2D" D3:D3" 3: 3H* DN" Z" $ ;2;M:8 ) * 2 Q N G HB$":7T"67T"67T"67 M 27 N :77! 65 Q! A! 0 277Gf $"ZH":57"62 N 57 1 : " 6!" 2S G1H M M " G6777 " 5 H" 97 ) G*H " " :7)c6MGDM Hd)% "%;7) % ) %! $ " " " = G :RRRT:RRR H " 93

70 '2 1 24G 6777H 25"5547Q" 669 * " G ):RR3H 8" 65"5 1G H") % G:7)"%;7H"2) 6O GH &%;7 2) 6O " )% G:7)"%;7H 76) 6O "D 6O "P 6O " 6O 9O! GD0 9 H 6 "D 6 "P 6 " 6 GD0 9 H 9 G:4 H ""!$ G 67 H 97 ) * G H ;77 G H " 97)*"65* G:4 H! 97 ) * " ;77G H *SGEH B E9388F; : <$388 c6m:d 94

71 '2 1 " " "! * $ 662$ "2577":" G8HG57)"%32H"5$:7 3 1 GH G" " AH 9µ1 "!" 67 $??)8 G?)"f =A %"AH %80M :779G$8257M2R7" 5:7? 5:5H?? 977G?)"% " =HD$:77 &T?)57M "5547Q? 977 " 67! " D $:77 &=977 9R

72 '2 1 *? ) 57M * B A9! ' G:6 c6m6d! " 66;! " 25 Q " $ :6 1 D 6O " 6O 9O!577! :57"!"65Q97 " " $ " T$!G 9SH! % =GMH!277GD!"*" H" 27

73 '2 1 ",/, 69: %G9MRHG:R4RT 6777H*% % BGH &%;7"GH2) 6O GH)% G%;7" :7)H 2) 6O T G 6O H66 G R5S 67 H" 6O " B69) 6O ":5)D 6O 74)P 6O % "DN::R7DN:9;2 G 66M8 H " " 6O "" R5S 5G686HDN:9;2 6O G775)H"DN ::R7 G7: )H G 6777H"! T"DN:9;2 2:

74 ' Settled cells (%) A B Settled cells (%) C D Settling time (min) Settling time (min) +",", ;2;M:8GH"DN::R7G8H"DN ::R5GHDN:9;2GHN G5 1H BG[H:7 )) "%;7" 2) 6O GHTG\H69 ) 6O ":5)D 6O 74)P 6O "&%;7G]H&% ;72) 6O E $TG^;H 696 * " " B 6O "D 6O "P 6O " 6O 9O "76) 69 DN:9;2 G 26

75 '2 1 $ M: H* 695" " 2" 29: 8 ;";9:G$M6H Settled cells (%) Settling time (min) + ",/,* DN:9;2 5 1 )% G:7)" %;7H" 76)BG\H 6O OD 6O TG[H 6O OP 6O TG_H 6O O 6O TG]H 6O O 9O TG`H 6O GHE $TG^;H * P 6O " 6O 6O D 6O G 6776H" 6O T " 6O GA 6772H 29

76 '2 1 *" * $!" $ " " * *" = M 699 =" 6$:7 ; 1G 7: 1H G HT 765 1"" 75 1" G62H * T + / )! G)! :R46T:R46 H " 1 =!" 75 1 M 22

77 ' Settled cells (%) Settling time (min) +",0, DN:9;2 G\H765" G]H 75"G[H:7"G`H 67" G_H 57 GaH:77 1" :7)) "%;7" 2) 6O E $T G^;H 692 % N " " G:RR6H0" G*=:RR7H*"! N!47Q 27S G65HT "2555Q G65H 0 " +/ 57M;7 Q" G):R;2T* 0:R35H*! 25

78 '2 1 47Q!! Settled cells (%) Settling time (min) + "5! N G5 1H DN:9;2 :7)) "% ;7" 2) 6O BG]HGHG_H25Q"GaH55Q G\H47QE $T G^;H N25Q 97)* G H 695" 2" 29:8;";9:G$M9H * G 6776H" 6O " 2;

79 '2 1 *! " 47Q G 6;H" ::5O4µ 9 59O9µ 9!47QG63H A B C D + ",2, M! DN:9;2bT8" b25"5547q" Cell volume (µm 3 ) Live 45ºC 55ºC 80ºC +",3,! DN:9;2E $$" 977G25"5547QH 23

80 ' 2 1 & G M Q G% G M H " 6775H 9" 999" 47 Q! 92H G 25! 55 Q 64H A B C + :9;2" ",5, D N 47 Q A G H" G H G8H 24 M M

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82 '2 1 :R 1 G* 6:H" G6 1H" GM 69;1R4H "D 6O G* 6:H R;SD 6O G 6RH*" 100 Metal removal (%) Number of batch + ",4,S! DN:9;2 : D 6O "577!!:57" 65Q97 " " E $TG^2H ",0, " " D" P" * R5S 5*!TDN:9;2"!" 57

83 '2 1 V75 1 M " 25Q = * " " TR:SD 6O R;SD 6O )" W "! * " " 5:

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85 '2 1 %"'G6775H$ " 2%F $ "38 ")*P ;RRM33;B?="DZ"!"D"Z"*)"ZG6772H 2 30" :2RM:5;?"N%"% ">""f"zg677;h =? $44"394M329 )" " "%)"*$" > '") G:RRRH' 6O " 6O 6O B% & $"0":93M:22 )""'")"%)G6777H% 6O " 6O 6O! "/":95M:2: )!"8?""D%">""E?G:R46H D3?! $1"434M44R )!" 8? " " D% " E? G:R46 H'?! $1"4R7M4RR )" > G:R;2H *?B /1";:M;4 D"k"*G:RR2H =B / $2"9;5M935 " " G:RR;H $! 01"634M645 " E"!" A" " "%) E) G6776H 6O B! "0";;9M;;; 59

86 '2 1 "E""'"%)E)G6777H 6MGDMH %! $""5RM;5 " E )") G:RR3H(?)! $/"R9MR4 " E " > G6777H k %! """:463M:496 "%)E)""?""DE")* G:RRRHDM % $$/40"965M995 "%)E)""8")A'*)G:RRR HDM % $=B $$":9:6M:9:3 ")G:R4RHNB 1"243M2R; ")G:RR6HNB $//":M36 *" D f 0" f? G:R35H?)! 5$"59M53 *" N " A G6777H = $?!31"52:M52; *=") G:RR7H "%?8"?965M99RB)A M% E" Z >"!$" A" E"% $" ' G6779H N B! $! 2$":R3M675 E"'% E!" 8 G6777H 8B q) /":3M62 52

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88

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90 52 1 /,$, - % = " " ""$ * B H $! G"%H" T H T" " )M M "" M "$ "G H 1 " "$*" = GE!6779H 8 " " GE!%:RR5T8!! :RR4TE E! 6777T f 677;H* "= 1 $ GE!677:H! " 58

91 52 1 G= *$:RRRTZ 6775Tf 677;H *? " " GDZ:R33T* :RR6TE! :RR9H0 " B " $ GA:RR7T E!:RR7T A:RR9H! "! "!" " " " GA:RR7T * :RR6T? f!:rr;t!= :RR3T T Aj! 6775H % "! 6O " GHG8 :RR2HT" 6O G * :RR6H 6O!GZ6777H!" G!! H G!25QH!" = *! "!""! " $ " M! 59

92 52 1 /,", 96:"!" D NGDNH:9;2 " M G6"66:H 966 6G666H 25Q 969& G"%rH ;77 *G97)H G H 60

93 :Z Z 577! :77 G2 1 H ) % G:7 ) %;7H )G"DPH 5: cGD0 9 H 6 P 6 d:7771cd 6 d!d 6 G)!H :57 "! G8 ")" AH" 65 Q " G:7 H! 725 µm = $ $ T G :SH!! " "! M" % M! 277 GD!" *" H" *!",Gµ1 H" B,^A$G b H$:7771G$H c9m:d "A$GH" G1H " T GH 61

94 :77! 97G21 H)% G:7)% ;7H G ! =H! :57 65Q: " 725 µm = 9629 *? " : G:65H"! 25QB,9,,:;3(,<c9M6d ",!Gµ1 H", Gµ 1 H $! "?G1µ HT, Gµ1H 965 '( 25Q :$:7 4 1:77µ 65Q! 62

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96 $ 9S G 1H G!H 6 :77 *" G577 " 5 H" 7:) G%;4H GH $ G6SH G!H : :77" 7:) G%;4H" 7:SG 1HM?MGH$ c:7"67"97"57"37:77sg1hd :7 D$" $ 8=k *" )G577H$ >0?>)M95:5!E$: R) G 25 QH" $"!="? G6 1 H " )% G:7 )%;7H6661 " 9691 D6;61 P :57:"65Q"! 65 Q G*'HG8)8M:52H!:77 64

97 52 1 Z89SG 1 H $ 2777;57 M: f M :7* $! " /,/, 99:+ *!G 6O "D 6O P 6O H "%;7" B 5 1 :77 1* G5 1H G:77 1H! %; G%:R46HZ "! 51":5 " 94 22S * $3749µ1 " G 9:MH 97 " B2:57S " 65

98 52 1 $3;R; 6O µ1 " " 97 ;7 $ R: µ1 " 2RS 6O G9:MH$ G H 62 "! "! =!!"$!! :7 G 9:M8 96MH 5 1!"!93;2D 6O µ1 " G 9:M8HT :77 1! " M!6242D 6O µ1 " G 96MH = G5 1H" BM! 94::µ1 "":7G9:MH = G:77 1H" c22 24S" $:26:;7P 6O µ1 " G96M8Hd :7 c27s" $:64P 6O µ M: G96M8Hd 66

99 52 1 Cu 2+ accumulated (µmol/g dry weight biomass) A Ni 2+ accumulated Zn 2+ accumulated (µmol/g dry weight biomass). (µmol/g dry weight biomass) B C Time (min) +/,$, 6O GH"D 6O G8HP 6O GH G]HG\H DN:9;2 :7) ) %;7 21 T 51 $ TGV2H 67

100 52 1 "$ 62!= Ni 2+ accumulated (µmol /g dry weight biomass) A Zn 2+ accumulated (µmol/ g dry weight biomass) B Time (min) + /,", D 6O GHP 6O G8H G]HG\H DN:9;2 :7) ) %;7 2 1 T :77 1 $ TGV2H 68

101 , & G 6O " D 6O P 6O H! M DN:9;2" $ $%G;7H G65 QH! G9:96H";7 )! G,H G,H G H G99H "D 6O P 6O! G 99M8 99MH%M D 6O P 6O G99M899MH 6O " T " G99MH =! "? $ P 6O " G^7452H"? $ G* 9:H*, G $ H D 6O P 6O G :3:2"H G* 9:H 69

102 52 1 q (µmol Cu 2+ /g dry weight biomass) A Ceq (µm Cu 2+ ) q (µmol Ni 2+ /g dry weight biomass) B q (µmol Zn 2+ /g dry weight biomass) Ceq (µm Ni 2+ ) C Ceq (µm Zn 2+ ) +/,/, 6O GH"D 6O G8HP 6O GHG]HG\H DN:9;2 :7) ) %;7 21 * : $TGV2H 70

103 52 1 " B :66 µ 6O 1 :5; µ 6O 1 " G* 9:H "! B 6O VVP 6O D 6O T " %(/,$,? 6O "D 6O P * ( Dµ> '(E D>µE B B B "G :66 :5; 6: :9 7RR6 7R25 ; "G 4 :92 :: :3 7R53 7RRR H "G :6 :; RR7 999$ $! " M ) GH $ T D $ M G%6775H 92"M25 Q *! B 25Q!65QN T " 71

104 52 1 A B +/,0, DN:9;2M!25Q GHM G8H G:RR:H G 677:H " " $ M G95H* A B +/,1, DN:9;2 M! 25 Q )6' GHT B M G8H 72

105 52 1 ) ) G)H D T M G9;H A 1 µm B 1 µm +/,2,Scanning electron microscopy of S. cerevisiae NCYC Live (A) or heatkilled cells at 45 ºC (B). Arrow: bud car. ) M *' * "!B 3R7 ::47 M: T :677b:6R7 T :277:377 M: T 6577b9477 M: M0%MD% T"!6R77 M: G8 677:TA 677:T 6779H* 73

106 52 1! G93H Absorbance A B λ (cm -1 ) +/,3,IR spectra of live and dead cells of S. cerevisiae NCYC Cells heated at 45 ºC (dead cells) (A); live cells (B). )"! $:732 M: G H" βg:h 9H " & G97M25S H "! B!:52: M: GMDDM% H!:;92 M: G^0 DM % H*! $92:; M: "" $ D% G 6779H 74

107 ) '( "- '(. '( * 6O "D 6O P 6O *' 94"! Absorbance A B C D λ (nm) +/,5,' DN:9;2 $ GHGHTP 6O G8H"D 6O GH 6O $GH :7) ) %;76 1 * B666 6O 1"969D 6O 1 6;6P 6O 1 ;765Q 600 "! 6577M9477 M: G94H$!92:; M: " "99;2"96R29645 M: " 6O "P 6O D 6O "* bd%b0% 75

108 52 1!6R64 M: G94H* " " "!":;92:52: M: G H" $:;2R M: G D 6O P 6O H":;5: M: G 6O H:59R M: G P 6O H "! ":732 M: G H" $:747 M: G 6O H * G94H /,0, *! Z N! B G H G H" T G H G H! G8! :RR5H D! = DN:9;2 :7T G ':R;4H" G 9:H $ G51HG) A 76

109 52 1 :R49T 6779H* $"!)!"!! = "D 6O P 6O! M GH G':R;4T:RR;H!"!G25QH G) 6774H "" "! M! G) 6774H!" G25QH! *! $! G( $ H65Q%;7" D 6O P 6O G:3:2"H G* 9:HT 6O "!* 25Q " " " * M * T $ = * βm βm βg:h 9H βg:h;hm!t βg:h 2HM!DM G:5 M;SH 77

110 52 1 *αm GH" 97M57S G 677:T? 8 677;HT G8 :RR2 H G:RR:H" B G95H* )" G9;H"' B! G93HM! " G92H"!=! $ M " 6O = GA:RR7H %%0% 6 G :R4:H" " " * $ $"! G 6779H!"51 6O 57S67 37S;7G$8M:H*" G9:H $! G 94H* 78

111 52 1 $""$ G!"H!* G6779H" D 6O 8 G:RR2H!" $$ 6O * 6O *! DN:9;2 B M" " % * 96 * $! DN:9;2! * 96 " * 96 T! DN:9;2!! = T $ * " " G! H N, 79

112 80 %, ( %(/,",! & %I Dµ> '(E DE?!W :; GE!)M:RR5H? W :;7 2 :; GE!)M:RR5H W : G8!! :RR4H! R7 ee 6 G8)%:RR4H?!W :77 ; 62 GM& :RRRH ; W : G8!! :RR4H?!W :9; ; 62 GM& :RRRH 34R 5 62 Gg=g=6779H!W :R2 ;35 62 G 6779H H W 59 ; : G8!! :RR4H! 654 ee 6 G8)%:RR4H?!W 6R GM& :RRRH I* II D 52 1

113 52 1 " G 25 QH! *! 25Q *!!$ T! $ M Z ;7 " '"! $" " $! 81

114 52 1 M&")" M?")"MD"8!"'G:RRRH8 = M?-0 "05":93M:27!="'"A "?N"G:RR3H=M!! 11":M:7 "E* ">)G:RR6H)! $15"9449M 944R 8!!""8"*>""?)"%"%!"G:RR4H GP" " DH " C /5G;H"6;RM633 8" " "%" 8" N Z" Z=" "!" Z!" 8 G6772H' M! 40"625M62R 8! "Z>""* ">)G:RR5H)! B $! 0/"53RM542 8" " >' G:RR2H 8 / $2";99M;94 8"""">'G:RR2H8 M $1"26RM294 8"""">'G:RR2 H=$!! 00"6R3M976 8">"?=">"A")"nM)"f" X"> '") G677:H = 0""95;RM

115 52 1 8") )%" G:RR4H 8 M! $4"95:M959 " "'" " ")" "? 8 E" G677:H*?! "32":R;3RM:R;46 =" *$" > G:RRRH* k GH GH! "$G5H" 29:M293 "AM'"G:R;4H*P 6O " 6O D 6O!!$$2/"965M997 A"A)G:RR7H * "%?8"?62RM635D N!B)A M% A"A)G:RR9H $- $"0G:H"65M;7 A" "!" A " 8 " )" ) G677:H *' B $!M $43":3RM:4; Aj!" N" m" An," G6775H 8!k! 42":79M :7R %"'G6775H$ " 2%F $ "38 ")*P ;RRM33;B %"G:R46H 0) 0$1) 2R0$ Z"*N"!"Z""N"Z"%8"Z"N"Z"Z"> G6775H +? "" R:MR4?"A8"%G677;H!* 39:3M929 83

116 52 1?" N f!" G:RR;H% M =?2/04":;5M:3R )")"")"A"""%))"E G6774H' k B*! 44G3H" 6:73M6::5 ) ">?A"A)G:R49HP!$?B $"4"926:M9265 D" ' Z" G:R33H?B 44"9:3M962 g="g="g6779h GH"DGH" GEH B?2/!$"6:RM66R " E" E" " G6779H 8!GH 8!k! /5":94RM:9R5 ">'G:RR:H /$40"396M395 "E"%!"Z"%)E)G6779H*$ B? 04"99;M929 "">'G:RR;H "! $5" :67RM:6:6 "Af"">">'G:R4:H) B $0$"693M625 ">%Z"G6777H% 6O 6O! "/"963M96R E"'% E!" 8 G6777H 8B q) /":3M62 84

117 52 1 E!"8G:RR7H!! E!"8 9M58'"?B' E!"8G677:H$M B $ 2 14"679M6:; E!" 8 G6779H! 5M:6 )B8E $" E!"8%"P'G:RR5H8! $$" 695M657 E!" 8 )M" % G:RR5H 8 $! 0""3R3M47; E!"8")"%%"P'G:RR9H!! 0$"46;M46R f "%? " G677;H 8 B! $"0G5H"263M25: 85

118

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120 42 1 / 0,$, - * =M M$ * G" $" H $T " M GE!677:H G 6776H'" "M!25Q" G) 6774H!G) 677RH" M M "M " G! H " " $!" 1G 1 H G! " H" 1 G 6776T) 6774H)" " "! N %" $ G H"" 44

121 42 1 / " % T %" G8 :RR2T = *$:RRRTP 677:T g= g= 6779T E 6779T) * 6772T D 6773H " % G "! H G:R49H G" H 0"% $" G 6772H* $ G " " " H G%:RR:T * :RR9T A :RR9H % $ G 1 HT " G :RR5T 6776 T 6779H '$ T $" $ ;O 9O * " GD Z:R33T=*$:RRRH " =GN 677:T f 677;TA677RH' G6774H G""D" PH T GH %'(" $ * % 6O ""% 4R

122 42 1 / " G% 6775H M GE 6772H $% "! '$ M T!! G 6779T) 677RH!" $ M!! *! $ =" MM 0,", 26:"!" D NGDNH:9;2 M" G6"66:H R7

123 42 1 / 266 " G6777 " 5 H 97)*G)!H M " 25 Q"!" G9H M!" $ 269&! G9"969H 262) / ") 0" " * = " % " $ "G"D"PH"G" ""H ) G% :RR4H *" M MGHG0R;M7R"8"H" Z3725 % D MM GHG"8"H )%6776% R:

124 42 1 / * G B " " $H"%" = " "%97 * = " 725µ= i725µ! 8 6 "Z% G)!H* 725µ= " % * G*H GH *0M5777G=H* G*0H * * M! 277GD!"*"H" :7771G)!H75)%D0 9 = " :77 1 T " ;O G =H * $! R6

125 42 1 / 265 * G" : 6H" "! GD0 9 H 6 "D 6 "P 6 GD0 9 H 9 " )% G:7)H"%;7:" G M 6M "D0 9M 0 2 H ) % $ " " " " = G :RRRT :RRR H 26; )D(?O G 25H G ) 6779H) )D(?O " " G H " G= *$:RRRT) 6772T) 677RH G) 6772H )D(?O*$ G 6O D 6O P 6O H *G9H R9

126 42 1 / 263! * B &%;7 D0%*" 725µ = f" :6 1"!577!":57"65Q97 " " Z G9H 97 $ " T$! G :SH!% 264* $ ""G^ ;H R2

127 42 1 / 0,/, 29: 0 / * " " = " "" G* 2:HT GM 69;1R4H GH " % " *" "! "M GM:R42H * * 2:! M * * $ G694EH%G67H 9O G $:R32H" G* 2:H *"! G" =H" " R5

128 42 1 / %(0,$,M= * GM:R42HGM69;1R4H Wastewater limit discharge criteria Industrial effluent US-EPA Portuguese law ph (units) E h (mv) 238 TC (mg/l) 11.5 TOC (mg/l) 10.7 IC (mg/l) 0.8 Cl - (mg/l) 90.5 F - (mg/l) 12 - NO 3 (mg/l) SO 4 (mg/l) PO 4 (mg/l) 0.08 Cu 2+ (mg/l) (µm) Ni 2+ (mg/l) (µm) Zn 2+ (mg/l) 2.61 NS 0.8 (µm) 39.9 NS 12 Total Cr (mg/l) (µm) Cr 6+ (mg/l) 0.1 <0.06 (µm) 2 <1 *B *0B B R;

129 42 1 / "* 2: G655)94R)"H" "!G7979)H*! """!" GH " ;O 9O Gf?677;H* 2: G H " " G$H! 296 0/ * " " B( ZT( $ Z %* B %" $ " $ G H" *" *( " " G ""H Gf 677RH* " $" "% 9" M R3

130 42 1 /! G" D PH"!" G) 677RH " % ;7 $%$% N %57R7 G8:RR2H" %5M; 6O " 6O " 6O "D 6O "P 6O 9O G> :RR:T 8:RR2T=*$:RRRTg=g=6779T) *6772H%;7" " G $" " " $ M H" G $ H GHG)6772H "( 0"%;% G* 2:H% ; " c 9O d" %" $ G) 6772H"!% " % $ GDPH$ G)6772H * & G H " *" = R4

131 42 1 / "! G* 26H ""$ ") 6O GH " %6T"G0%H 6O ;,< & G0%H 9 * 26$"" 2: %(0,", "Z""!="%;7"%57e log K Ligand Cr(OH) 2+ Cu 2+ Ni 2+ Zn 2+ Reference Biomass 3.56 Ferraz et al. (1999) Machado et al. (2009) Cl a) 0.20 b) 0.41 b) 0.40 b) F a) 1.80 b) 1.40 b) 1.30 b) NO a) 0.50 b) 0.40 b) 0.40 b) HCO 3 - _ -3.56c) Herrero et al. (2005) 1.80 b) 2.10 b) 1.50 b) SO 4 2- H 2 PO d) 2.36 b) 2.30 b) 2.34 b) 6.42 a) _ e " )?B HG0%H 6O O?O% O?O% 60 H)O?)? RR

132 42 1 / H 6O O%0 9 M0 9O% O HG0%H 6O O0 2 6M0%0 2 $ " " 2: H MHPO 4 (aq) H 2 PO HCO3-3 4 MNO 3 (aq) NO - 3 MSO 4 (aq) + 1 B + M n+ MB (s) 5 6 SO Cl - MCl (aq) + F - MF (aq) 2 7 MCO 3 (aq) + H + + 0,$M)B) O 8GHB8"$ N G:H * G 6 M;H ** 26 ""!=" G H * " " $ " )%0 9 ")"))D0 9 "6"9"25" G2:H" $T" 1! 1 = G:H 111" :77

133 42 1 / * 26 = G H 0 "! *! G;H% " * :5 1 " * 2:" G93271H"62S " " S! G 26HT G932 1H"! 95S" ::S G26H % ;O 9O *!! G26H" " $ " ;O 9O 8"! $%T$ 637E%65 ;O 9O G $ :R32H * GH G* 26H" :7:

134 42 1 / " $ G 2 3"H"" G2:H 100 Nickel species (%) x 10-3 M 3.9x 10-2 M (374 mg/l) (3740 mg/l) Sulphate concentration +0,"! :51 DN:9;2! G HT! G HT! G! H*! G)D(?OHT ""!=! BVV PVDGZ * 26H G= *$:RRRT) 677RH* M$ T " " " G8 :RR2H*" " Gc8dH Gs c)dh $ " % " :76

135 42 1 / "!GH=G HG* 26H"!" G"=H"! MM"! "! " = * 2:" " " )D(?O G ) 6779H *" " T " $ :5 :6 1" %;7" G* 2:H* 29M:51! "! G!D0 2 H 0" :61" " 42S! T " "! G!D0 2 H G29M8H:5:61 52$:7 M2 25$:7 M9 ) G) 677RH" :79

136 42 1 / $ "!:4:2R"* $! Concentration (µmol l -1 ) Concentration (µmol l -1 ) A Effluent Batch 1 Batch 2 Batch 3 Batch 4 Batch 5 B Effluent Batch 1 Batch 2 +0,/! M:5 1 T8b:6 1 D!G HT! DN:9;2 G HT! G! H! 26!B671G92µΜH :72

137 42 1 / 299!!"G25QH! T " " " G":6H :"G M "D0 M 9 0 6M 2 H *! ":6 1!" " " G H!! " " * 29 22" *! 22T "!%;7 "!" "!G H)D(?O G)6779H :75

138 :7; %(0,/, 8 DN:9;2" * GM:R42HGM69;1R4H Effluent Limit discharge criteria US-EPA Portuguese law Metal concentration (mg/l) Original effluent After 1 st batch After 2 nd batch After 3 rd batch Industrial 17.8 ± ± ± ± 1 Nickel Synthetic ± ± ± ± 0.3 Synthetic 2 19± ± ± ± 0.3 F )$;96< :6 )% "%;7 :"G M "D0 9 M0 2 6MH 42 1 /

139 42 1 / "! 37";536S G22H" " "! 63" 96 6; 1 " G* 99H "! 6"66:;1 G* 99H" 4R"44R6S " G 22H* "!"! G9R4671H" M GM :R42H GM69;1R4H 100 Nickel removal (%) Number of batch +0,0! DN:9;2 $ 8 B T B:G HT = B 6 G HT! B * 26 $ :6 1 * 29*!!:57"65Q97 " " E $G^;H, 107

140 42 1 / *! *" " "! $! G22H*!! $ G H 1 " $! G 6776 T 6779H * $!! " 0,0, *!$ G " " " " H" " " "!= * " " " "! = 0 " * " " :74

141 42 1 /!* " " GH ;O 9O * " $"!%;7" " Gc8dH" Gsc)dH $f c8d1gs c)dh " "! " G( ZH "! "! "! *!4RS :7R

142 42 1 / References %"fffg:rr4h C C %"f " " E * " >) G:RR9H) M $14GRH"645:M645; 8"">'G:RR2H8M $! 0$G:H":2RM:52 8"""">'G:RR2H8 M $1"26RM294 E" ")!"? A " G6772H 8 M?! 34G:H"2RM5; M 69;1R4 G:RR4H $ le E - LL'I "<M":3;1R49::3M9::4 =" *$" > G:RRRH* k GH GH! "$G5H" 29:M293 "*) G:R49H*M $ 0 $ "G3H":;6M:;; A"A)G:RR9H $- $"0G:H"65M;7 A"A)G677RH8B "?!50G:H" :9M64 %"'"?""'M""E"*E")G6775H ' C * /4G:2H"9:RRM96:7 %")D"'ZG:RR:H) M M GH?B $/3"365M392 ::7

143 42 1 / >"Z "AG:RR:H8! 0G2H" 699M693 )")">"""%"EG677RH' k B?$$2G;H":3R6M:472 )")"")"A"""%))"E G6774H' k B*! 44G3H" 6:73M 6::5 )") *" D G6772H* 20G:H"9RM23 )""')G6772H-)* & 46A H8" -) & B "D* D" ' Z" G:R33H?B 44"9:3M962 g="g="g6779h GH"DGH" GEH B?2/!$"6:RM66R " Z D"' G6773H 8 M 0"G9H";65M;94 $") G:R32H $, $, %" *$" "*")!"? " " > " "? *=") G6779H D!! M! $2G2H"5;3M54: '" " A==" " 8" " 8==")? )=" > G6774H 8! 44G::H"2;42M 2;R9 :::

144 42 1 / " f )" G6779H)D(?OB ) " E 25 f " W)% " )B' "l"*"n"">%"?g6772h"" " ="! B =? ) "31G:H":9:M:2: " "* " >) " G:RR5H M?)$0G9M2H"627M62; "E"!"A"""%)E)G6776H 6O B! "0";;9M;;; "E""'"8"'"%)E)G6776 H E $ k$! 15"49;M42: "E"%!"Z"%)E)G6779H*$ B? 04"99;M929 "%)E)""?""DE")* G:RRRHDM % $$/40"965M995 "%)E)""8")A'*)G:RRR HDM % $=B $$":9:6M:9:3 MG:R42HA "M2271:M4217R:"GH"f "B ::6

145 42 1 / E"""E"G6779HZ 8!k! 54G9H"64:M643 E!"8G677:H$M B $ 2 14"679M6:; f ">"G677;H8 B! $"0"263M25: f">?"ng677;h : $ "2% f"?z" "D%"N245% f " >? " G677RH 8! $"3G6H":R5M66; N" N "!" "!" >) E!" 8 G677:H 8 /1G6:H" 2959M 2954 P " ")" Z?=" D Z G677:H' B /2G9H"92RM9;5 ::9

146

147 1 - ('6 0!( 8 $ " "!" 9O " 6O D 6O "" M * 9O T" 6O D 6O * " 9O $" " 9O *" 9O!*$ 6O D 6O 1,$, - * 2

148 I) * G H " M " "!" B " $" = GA:RR7H) M"%" GA:RR9T D& 67:7H* G H G" H G :R49H " $ ""G"*H G ""HGD&E! 67:7H " " *" " GA :R4;H "=E!G6775H? 9O " 0"&G:RRRH 6O! D *! $ "! " GA :R4;H"* G:R43H 6O 6O! *// * "' G:RR6H! *T ::;

149 I) G H G H "! G6772H *! c 9O " 6O D 6O d "M 1,", 56:" * D NGDNH:9;2! M" G6"66:H 566 " G6777 " 5 H" 97)G*H M 25 Q M!" $ ::3

150 I) G 9"969H 569, * ;O 2" " 26; %;7" " " G=*$:RRRT ) 6772T) 677RH G)6772H*$ G 6O D 6O P 6O H *G9H 565! * 634 1*" $ ;O 9O * ;O " 262 " % &;7 D0%T 725µM = ::4

151 I) f" :4 1"!577!:5765Q97 " " Z G9H 97 $ " T$ G6S"H!% 2"262 56;* $ G^2H 1,/,!" " " *" "" GA 677RH* "" ::R

152 I)!"! M! G8 ;H* 9O D 6O M GM :R42H? GM 69;1R4H G* 5:HT " 6O $G* 5:H 8* ;:"; G 6772H " " * 56" G 9O " 6O D 6O H!" " * D 6O 6O G* 56H* $ 0 "* 56 9O 9O *" 9O " * 5:" 9O "" %;7T!"$ 9O %;7" 9O G* 5:H :67

153 %(1,$, DN:9;2 Metal concentration (mg/l) a Presence of fluoride Limit discharge criteria Effluent b After 1 st After 2 nd After 3 rd batch batch batch US-EPA Portuguese law with 24 ± 3 21 ± ± ± 1 % without 15 ± 3 5 ± ± ± 0.2 :6: with 1.7 ± ± 0.1 ND ND D--E without 1.5 ± ± 0.1 ND ND with 21.1 ± ± ± ± 0.1 ;D--E without 21 ± ± ± ± 0.4 with 1.2 ± 0.1 ND ND ND HD--E 2.61 NS without 1.0 ± 0.1 ND ND ND * G;"8H"M GM:R42HGM69;1R4H DBT DB ) $;94< GEH "G2H %&;7 I)

154 I) B %(1,", A D.E " G "G ; "G C 95; =*$G:RRRH 9RR 992 ) G677RH + : 44; :47 :27 )G6779H " Z" G H!"%;7"%57 " )?B G0%H 6O O?O% O?O% 60 )O?)? " " " G* 5:H" " " c 9O " 6O D 6O d"mgm:r42h GM69;1R4H"" T " 9O 97S G5:H" "D 6O G* 5:H "R:SD 6O " " " G5:H :66

155 I) 100 Metals removal (%) Number of batch +1,$,S M DN:9;2 )B 6O G HTD 6O G =HT 9O G! H8 :41 "%;7*!!:57"65Q97 " " E $T G^2H * $! G56H 9O G* 56HT" "$& 9O G;;S" 56MH* 9O G9:SHG56MH" GR:SH G56M8H" G* 5:5:H 0" 6O D 6O G* 56H*" G H 6O D 6O * :7S 6O D 6O $ :69

156 I) G56MH" G56M8H Metal species (%) A Metal species (%) Cr Cu Ni B 0 Cr Cu Ni +1,", 9O " 6O D 6O G!H G=H G HT G HT G! H* G)D(?OHT 562 $ f" 9O $ " % " " 9O " :62

157 I) G 6O D 6O H*$D 6O *! " "D 6O 1,0,!" " " 9O * $ 9O % " 6O D 6O M :65

158 I) &""A"'$"'ZG:RRRH D /4G:H"2RM56 M 69;1R4 G:RR4H $ le E - LL'I "<M":3;1R49::3M9::4 =" E E!" 8 G6775H 8?" N C * /4G:H"69RM623 =" *$" > G:RRRH* k GH GH! "$G5H" 29:M293 "*) G:R49H*M $ 0 $ "G3H":;6M:;; A"A)G:RR7H * "%?8"?62RM635D N!B)A M% A"A)G:RR9H $- $"0G:H"65M;7 A"A)G677RH8B "?!50G:H" :9M64 A"8"%")")"'"%=")"$")" fg:r4;hghgh$ "G;H";63M;96 )")">"""%"EG677RH' k B?$$2G;H":3R6M:472 )""')G6772H-)* & 46A H8" -) & B "D* D&"A)E!"8G67:7H ""2""$ 2%2 :6;

159 I) f "?Z"">"%"N*"DZ:9M;:8 '"?B*o' D&" A)" E!" 8)" E G67:7H!1 )!!")DNBf ' " A >"!" P"N G:RR6H! M 3"G6R:H"4:MR9 ") G6772H +%0D 347M344>f o" * ">)""AD"'>G:R43H *//!! /G3H"446M44; MG:R42HA "M2271:M4217R:"GH"f "B :63

160

161 2 ('8 7 1!( N = % " $ G""DPH *! " %;7 M GM! H" * "DPGH" D G 8H " % & ;7T 8" ;O 9O %&;7" " TV4R"R:"R6R2S D"" P"!" $ * 5 >*80:7:7761&

162 62 1 2,$, - ) " " & = GE!6779H* " "* G "H" " :77?1 6 GA! :RR;H* $!$" "DP) :67 GE!6779H *M )!B G $HT " G H GE! 677:H* $ " " M " M $ M Gf 677RH " = = " f G677;H" G 130

163 62 1 H G 677RH M " ")""""""D" "" P G :RR5T = *$:RRRT P 677:TD6773Tf 6774T) 6774T) 677RT' 67:7H +/ + / GH" " T G H"!! M G*=677:H * G H "% G "! H G:R49H *! * M GM W H" * M " M 131

164 62 1 2,", ;6:"!" D NGDNH:9;2 M" G6"66:H ;66!" G2"266H ;69& G9" 969H ;62 / * G 8H ) 0" " * = " %"$"G 6O "D 6O "P 6O " ;O H" G" " " H ;O 132

165 62 1 ) G% :RR4H * % %0"26:7 )$ M G)H " " 2"2628" " " M " $ ;O 9O "$G:"6b $ H G*H G!H T ;7Q":7 " 725 µ= * & 7R)G 6777H $G$RM%H" :1" MG$lM:67H M G" 8" H 8 67 % * M" 8 72)" % 6" * D0 9 D"""*"" ;O 2"262 8" =$ 133

166 62 1 ;65 2" 26;" "%;7" G H" G=*$:RRRT)6772T ) 677RH G)6772H* $ G 6O D 6O P 6O H *G9H ;6; * G8H ""DP" &%; D0% 725µM= 8"""DP" $ ;O 9O * ;O " 2"262*" % &; D0%T 725 µm = " :4 1!577!:5765Q97 " " Z G9H

167 62 1 $ T G :S" H!% ;63* $ "" G ^;H ;6" 2,/, ;9: / ' "!"! G PH D GHMDG8H "= G* ;:H 135

168 62 1 %(2,$,! G%&;7H* GM":R42HGM69;1R4H Wastewater limit discharge criteria Industrial effluent US-EPA Portuguese law ph (units) E h (mv) TC (mg/l) TOC (mg/l) IC (mg/l) Cl - (mg/l) F - (mg/l) NO 3 (mg/l) SO 4 2- PO 4 3- (mg/l) (mg/l) A B (mg/l) (µm) (mg/l) (µm) (mg/l) (µm) Total Cr (mg/l) (µm) (mg/l) 0.1 <0.06 < <0.06 (µm) 2 <1 < <1 %7 %.7-7 F GEH "G2H%&;7 ** "%6:""DP M GM:R42H GM69;1R4H' 8"%69""G33SH ;O "D 0"* ;: 136

169 62 1 ;96 1 " M GM W H"%;7" M %"! G& %;H8" ;O 9O " G H %69" B 6%0 2 M ;,<O5% O ;,<O9%0 9 M ;,< 6 9O ;,<O5% 6 0 ;< O90 2 6M;,<c;M:d G$ ^ 667 EH "8 % ;7G* ;:H "% ;7G* ;:H%" G4:SH 8" 27S 9O %G* ;:H%;7= P 6O G5;SP32S 8HP0 2 G26SP65S8H P %;7 " 97S P" 137

170 62 1 " * P G8HG8H *%; BGH$% TGH%;% G* ;:HTGH%" 9O "$G) 6772H" G* ;:H" DP TGH%" M G $" " " $ M H" G $ H GHG)6772H " ")D(?O G) 6779H * $ :4 1" %;7" G* ;:H :41 8 G H %;7 ;: G$ H " " :4 1 " " P" 8" " ;:" " %;7" G* ;:H* & GV 4RSH G H *! 138

171 62 1 " M "" :41 T"c8d1sc)d :7 Metal species (%) Ni Cu Zn A Metal species (%) Ni Metal Cr B Metal + 2,$ :41!b T=b8) G HT G! HT G H G)D(?OH G%&;7H" " " ) " " 139

172 62 1 * ;6 ;6" T G " %;7 H ;6 " GR9S HT" M "DP 535RS"T 8"D ;35RS" " " 8"" " " " M D P " " 62 :: 1" G* ;6H* DP4RR2S"" G;6H8" TD:6 661" G* ;6H* D R6 R7S" " G;6H 140

173 %(2,"8 DN:9;2" * GM:R42HGM69;1R4H 141 DBT DB t)$;96< ttb%&;78bgeh "G2H% &;7 62 1

174 A Metal removal (%) Cu Zn Ni 1 st B 3 rd 2 nd Metal removal (%) Cu Cr Ni 3 rd 1 st2nd +2,"$ DN:9;2 $ b T 8 b 8* B T $ B! * " ;65 $ :4 1 E $G^;H 2,0, " G"$ " $ " 7 " J " B H GE& 677;TZ 6774TE&6774T 677RT & 67:7H% " GA677RH* 142

175 62 1 " G :RR5TP 677:T) 6779T) 6774H' " " MM" GH % B"$ G8! :RR5H)!" $" " $ G! " H!G8! :RR5T) 677RH""DP! BVV PVDG= *$:RRRT) 677RH M $ T " " " G8 :RR2H*" M" Gc8dH Gs c)dh $ " 5" DGZ^992"%;7H" DG"PH D "DM D " 8 143

176 62 1! G& %;H" 8 G* ;:H* $")G0%H $ "$% Bc) $O GHd^Z1c0% M GHd $ TZ $" $ % " 0 9O 6O $G)6772H" G! H 9O 1 6O * ;:"!" M * M " " % % 67"! " %" % 5;G8 :RR2T=*$:RRRTg= g=6779h!" M N " G H" G677RH M! T 6O G 677RH 6O M 144

177 62 1 G 6776H! 6O 9" " 25 Q" " " * =" VR5S 5G) 6774H" 6"M! DP"%;7G9H "! M)"!" & * G H )"! " " *"! $ " " " "! * ;6" " GM:R42T M 69;1R4H * $" %;7" * " " %;7 145

178 62 1 " " *$GR9S H " R3R4S 8"G;6H0 "$G" DPH G;6H % " $ T " " DP ::;S"" 8" D :74S"* B GH " " "" $ G HTGH G= *$:RRRT) 677RH?T " " $ G? 6775T% 677;HTGH$ T " G 9H"$ G 6776 T 6779H**0 i::1 G* ;:H" % " $ "G*HGD*H 146

179 62 1!! G 6772H" * $ G) 6772H " " %M " G* ;6H" D""PV4R"R:"R6R2S" " M! " G :RR;H" = b G :RR3H $ M GP :RR4HT " )" "$" G& 677RHT " S G& 677RH "! )M " * 4R 147

180 62 1 2,0,!" " "%;7" MG" "DPH 9O 1 " GH GH " 9O 1 0"MG""DPH" Gc8dH" GsGc)dH $*:41 6::31DP GH " G H%;7!" " $ " M *"! 148

181 62 1 %"fffg:rr4h C C %"f " 8! "Z>""* ">)G:RR5H)! B $! 0/"53RM542 8"""">'G:RR2H8 M $1"26RM294 &" Z" 8&" " A G67:7H 8 B? 2/ $31G:M9H"::3M:65 "f">?g6774h' 6O " O " O 6O k?2/ $1$G:H";5M37 &""")"""">E"EG677RH8 GH $0 $//1G:M9H":4:M:44 M 69;1R4 G:RR4H $ le E - LL'I "<M":3;1R49::3M9::4 =" *$" > G:RRRH* k GH GH! "$G5H" 29:M293 "*) G:R49H*M $ 0 $ "G3H":;6M:;; A"A)G677RH8B "?!50G:H" :9M64 A!"E"!"AE"E "E"P"?%"?!"E)"?!"D">>G:RR;H C $""G2H"94:M

182 62 1 %"'""8"?""'M"E")G677;H GH 44G:M2H":7;M::; Z" '" 8" D '" A 8" Z G6774H 8 GEH?$/1G9H"676M674?" "'M" " 8" >?" E") %"' G6775H 8 B 8 " " GDH? ) "54G6H"956M954 )")">"""%"EG677RH' k B?$$2G;H":3R6M:472 )")"")"A"""%))"E G6774H' k B*! 44G3H" 6:73M6::5 )""')G6772H-)* & 46A H8" -) & B "D* )" Z " P " "?=" D Z%!" G6779H )? 3G:M 2H"RRM:74 g="g="g6779h GH"DGH" GEH B?2/!$"6:RM66R " Z D"' G6773H 8 M 0"G9H";65M;94 150

183 62 1 " E" P")"'" "*" E" " E" A G677RH! $G::H"6337M633; '"?""")")"""G67:7H ' p!!=p $! 51G9H"3;9M33: " f )" G6779H)D(?OB ) " E 25 f " W)% " )B' ") G6772H +%0D 347M344>f o" " "* " >) " G:RR5H M?)$0G9M2H"627M62; "EG677RH! 2 & "E':79M::6BM "E"!"A"""%)E)G6776H 6O B! "0";;9M;;; "E""'"8"'"%)E)G6776 H E $ k$! 15"49;M42: "E"%!"Z"%)E)G6779H*$ B? 04"99;M929 "">'G:RR;H "! $5" :67RM:6:6 151

184 62 1 " " >' G:RR3H M 43G9H"623M65: *=") G677:H 8 * $!2 14"62:M629 MG:R42HA "M2271:M4217R:"GH"f "B E&" Z G6774H 8! J 0 $ C /2G9H"6RRM975 E&" Z" " Z E") G677;H*!! 0$G2H"459M45R E!"8G677:H$M B $ 2 14"679M6:; E!" 8 G6779H! 5M:6 )B8E $" f ">"G677;H8 B! $"0"263M25: f " >? " G677RH 8! $"3G6H":R5M66; P")">'G:RR4HGEH M!! "G;H";79M;7; P " ")" Z?=" D Z G677:H' B /2G9H"92RM9;5 152

185 3 & ('8 2!( * %M " " "D" ;O 9O ;O GR4SH %69%" ;O %0 2M % O " ;O "% ;7T%$=% * " %;7" " " 6O "D 6O " ;O *! ;O 9O 6 f""kg0:7:7731::637m7:7m7996m:h

186 L2 1"- 3,$, - " " " D P 0 $" D " " " $ GA677RH* G HT"" $!"M! "! " " " " ;O 9O! "!" ;O 9O " " "" 3,", 36:" * D NGDNH:9;2 154

187 L2 1"- M" G6"66:H 366!" G2" 266H 369, ) "" *8" ;* ;O 2" " 26; GH%69" ;O GH%;7" 9O " 6O D 6O %",! %69", ;O g= g= G6779H %;7", 9O =*$G:RRRH*$ 155

188 L2 1"- 6O D 6O * G9H 365! 0 f " :4 1" G 2 H" % 69*!" :57 " 65Q" 577! 97 " "% & ;7 D0%T D $ "97*" " Z " G9"99:H" 97 %G75H %;7$ " T G:SH! 3,/, * " *!!!G=H M! * 3:" "2"%69""G35SH ;O "! M 156

189 L2 1"- GM:R42H:R42H GM 69;1R4HT " f 9O ""G" H" ;O! " G)=:RR9T 6773HT " " 9O GA=677;H $ ;O " G7: 1HG* 3:H ;O 9O!" " T "M B %6" ;O T GH" % ;7 B 9O " 6O D 6O "" $%G69;7H ;O 9O " 6O D 6O "8" :41M! DN:9;2 G;";96H *%69 ;O BH ;O!G%6M65HH %" * % ;7cG 9O " 6O H"D 6O d GH%" 157

190 158 %(3,$8 DN:9;2" Metal concentration (mg/l) a) Limit discharge criteria Industrial After 1 st After 2 nd After 3 rd After 4 th US-EPA Portuguese law effluent b) batch c) batch batch batch Ni(II) ± 2 22± 2 11 ± 1 5 ± 1 3 ± 1 Zn(II) 2.61 NS 1.4 ± 0.2 ND ND ND ND Cu(II) ± ± ± ± ± 0.04 Total Cr ± 3 9± 2 3.7± ± ± 0.4 Cr(VI) NS ± 1 0.3± ± ± 0.1 <0.06 *GM:R42H GM69;1R4H DBT DB H) $;94< H%"2"%69 H %69* %;7 G $ H G H G) 6772H GH $ % L2 1"-

191 L2 1"- * "%69" R3S ;O * 3:" ;O "791" R4ST $ *"%" ;O G3:H" 9O G* 3:H %;7 100 Metals removal (%) Number of batch + 3,$, M DN:9;2 )BG HT! G! HT G HT GEH G =H8 :4 1 "%69 G H%;7 G H E $TG^2H * % % G% i 5H" ;O %0 2 M G36H 159

192 L2 1"- 0 " = $ " "G9H %G%i9H" " % O GD6773H" ;O Gg= g=6779h$ ;O G* 3:H Cr(VI) species in solution (%) ph +3," 95$:7 M2 1GEH% * GEH G)D(?OHB0 2 6MG]H" MG\H"%0 2 MG[H"% 60 2GaH "%;7" "$= " %%67"c 9O " 6O D 6O d G* 3: 3:H0%GH"DGHGH %5M;G8:RR2T=*$:RRRTg=g= 6779H 160

193 L2 1"- ;O %;7" 56SD 6O 34S 6O G3:HT 161

194 L2 1"- " 6O G:71H G* 3:H "D 6O 9:31"G* 3:H" "M G H G HT 43R9SD 6O "G 3:H" " ;O 77;1" G* 3:H0! G& 677RH"$ M =GP:RR4HG 677RH" $ 3,0, *! $ GMH " M ;O M " %69"R4S ;O *" D 6O " ;O " 6O " M G:R42H GM69;1R4H" %;7

195 L2 1"- 8"">'G:RR2H8M $! 0$G:H":2RM:52 &""")"""">E"EG677RH8 GH $0 $//1G:M9H":4:M:44 M 69;1R4 G:RR4H $ le E - LL'I "<M":3;1R49::3M9::4 =" *$" > G:RRRH* k GH GH! "$G5H" 29:M293 A"A)G677RH8B "?!50G:H" :9M64 A=" E ") G677;H $ "1G:7H":77;M:7:5 )""')G6772H-)* & 46A H8" -) & B "D* )=" f G:RR9H %MD M '? - $"/G2H";6;M;99 g="g="g6779h GH"DGH" GEH B?2/!$"6:RM66R " Z D"' G6773H 8 M 0"G9H";65M;94 " "? G6773H B A 1"G:H":M:4

196 L2 1"- " E" P")"'" "*" E" " E" A G677RH! $G::H"6337M633; MG:R42HA "M2271:M4217R:"GH"f "B P")">'G:RR4HGEH M!! "G;H";79M;7; 164

197

198

199 5 & 9 ( 3!( *!! M *! "! = " * B G:H T G6H G%H T G9H T G2H!"! $" =%^:2TG5H=" =$" &%:7* GVRRSH GRRR" R6 RR2S "! =" H*! 5,$, - M M " 7 G>%=)"B%P)*MM:7M 7:R39"H :;5

200 H 1"-. M 8" BGH TGH G H ) *"! " "BG7:)HG%"%D0 9 % HG :R4:T f :RR5T f :RR;H G% 9 00%H G= 6772H" G*H G :R4:T= 6772H G :R4:T f :RR;H% " G B "! 1"! 9O " G :R4:T= 6772H " 0" $"! W G M H" T * " " *?" G*! 6779T*! 6774H "! =! G6772H c *! G6774HdT!" "! = :;;

201 H 1"-. R25S";5RS35RS" "! =" " R5SG*! 6774H % " $"" $G% 677;H *! " " "! =" " " " *" " "! = T! " M" " 5,", 46:) "!= M G ;T H" 25Q Z 557 Q GD "?51;H":5 "!" :;3

202 H 1"-. * * $ %:) G") 9777H"67 25" 477f*"25 %:) G25 477fH 677 G f H " :5GB59f 1H " Q * ""! = " " M" G 2" 262H* "! = 95"64291" 469 8"% % 6 * T 1GZ9)H " :;4

203 H 1"-. M7;Ec1GZ9)d G "A**976H 27 "G577H"95Q27 Q =T = " M "! = " M" *$ 462 2" 26;!=" "$$ G) 6772H 465 % //234 "$GD0%H & % :2! $% 6:2 $! =! $ cdg0%h 6 GHd$=GHcPG0%H 6M 2 GHd"T "% :;R

204 H 1"-. 0% M 0% M *% & D0%=*" G6477 ":7 H! DG0%H 6 D! :7 D0%"%^:2"GH :7D0%%:7T " G6477 ":7 H " % & :7" %93S"="=$"PG0%H 6 GH " G6477 ":7 H = $* G H D0%" %:7" G6477 ":7 H *! = " =""!= M" %D0 9 67SG1H GH! 277 GD!" *" H "!" = G $H 5,/, " 67" "! =" " " ;* "" :37

205 H 1"-. MG;H% " GHGH$ *" "!= " $ " "!" B GH T GH "! 49:) 557Q $ G 677;H G6":7 :5 H *" :5 Z 557 Q 4:" :5 2 " 49:R3;S "*= :3:

206 H 1"-. G H :65G977H" R43S* 977 G ^::SH 2 " 2 Mass decrease (%) Mass decrease (%) Time (h) Time (h) +5,$,:5 W" " 557 Q 496$ $= "! = " T " G?MA@? :36

207 H 1" H* " " 8" " "!= " G8 :R32H" " T" " c4m:d c4m6d" G:RR2HB 6O GHO6 M GH 7 ^792Ec4M:d D0 9 MGHO2% O GHO9 M D0 GH O6% 6 0 GH 7 ^7R;Ec4M6d D$"D0 GH " $ $"D0 6GH "$ )" G*:RR9H % " $B 6GH O6 M 6 M GH 7 ^:9;Ec4M9d 0 6GH O2% O GHO2 M 6% 6 0 GH 7 ^:69Ec4M2d :39

208 H 1"-. %:) = $ G " HG H *277G677677H" %:)$ 466" "! = * 95" "! =" $ " "! = " " 499 "% / 499: G=! :RR5H M72E1c1GZ9)Hd % " G! 6777H M!= M!= M7; E1c1 GZ 9)dT " ""% %6 " 95Q27QG577H :32

209 H 1"-. 46 ;7 " * $ 6O O 7 G! 6777H*" *!27Q 95QG46H A I (A) Time (min) B I (A) Time (min) +5," * % & 6 M7; E1c1GZ9)Hd"95QG!H27QG=H :35

210 H 1"-. f =" " R7 " 27 Q" 97 1 " RR:S95Q"! :97 " 3 1 " RR4S * 9527Q" G MH * " 27 Q" " RRRS" c S G 1 Hd! =" GRR5SH! G6777H "" $""= *! " " G"! =H M7;E c1 GZ 9)Hd G27 QH G577H *! G " " M H G%:RR2H!M= " "! =" :3;

211 H 1"-. "! = 4996!= "! "" *!"" = *! =Gc4M5dc4M;d"" H757EB D 6O GHO6 M D GH 7 ^M765Ec4M5d P 6O GHO6 M P GH 7 ^M73;Ec4M;d *" "! =!="= 7M6E 1 GZ 9)H % 6 D! M7R5 E1c1 GZ 9)Hd = M:6 E1c1 GZ 9)Hd* $! = G765EH "! = "MG*! :33

212 H 1" H % " " $ G% 677;H *!! = " "!" ")D(?O"! = " "% 5:; G49H %3% :9"=$"$PG0%H 2 6M GH" %^:2G49M8H%"! DG0%H 6 GHG49MH*! =! =!! = G 49H" % %^:2% " $ & % G465H :34

213 H 1"-. Fraction (%) Fraction (%) A ph B ph +5,/!=%641!G!H291=G=H * G)D(?OH B " ) 6O GHG`H")G0%H 6GHG_H)G0%H M 9GHG\H)G0%H 6M 2 GHG]H " R3S! G* 4:H"! $"= D!$ 4S G* 4:HT G R7SHG42H :3R

214 H 1"-. %(5,$,'!$ Without wash Washing steps at ph 10* Recovery (%) 97 ± ± 1 96 ± 5 98 ± ± 1 Purity (%) 8 ± 1 40 ± 1 63 ± 7 77 ± 2 92 ± 7 e* D0% %:2T %:7 ) G 6SH = G42H Mass (mg) Without washing Two washing steps at ph 14 plus one at ph 10 Two washing steps at ph 14 plus two at ph ,0! $ G H" G H" = $ G = H $ G! H"!$ " &! f T Two washing steps at ph 14 plus three at ph 10 Two washing steps at ph 14 plus four at ph 10 :47

215 H 1"-. 97M;7 1 D 6 ;% 6 0 G% :RR2H! $ = "!$ $ %^:2 %" = G 49M8H" =" D$" $% :7 * 4: G4R6SH "42 *! $!" R6S" G!M!M HGM%%6775H =G49M8H"%:7" = PG0%H 6 GH"$"% & %:7 = =$G* 46H %(5,",'=$ Washing steps at ph Recovery (%) 100 ± 3 95 ± 6 88 ± ± 6 Purity (%) 55.3 ± ± 2 99 ± ± 0.8 :4:

216 H 1"-. * 559S" G 45HT! G 45H* = $ " :671DG%:RR2H Mass (mg) Number of washing steps at ph ,1 = $ %:7G H" G H"!$G =H$G! H"=$ =$ " " $% :7" G* 4645H "R3SG* 46HT G 45H f " RR RR2S" " G* 46H :46

217 H 1"-. *! = $!! = G* 4942"H " D%^:7" = $" %(5,/!$ Metal concentration (mg/l) Washing waters* 1 st 2 nd 3 rd 4 th 5 th 6 th Cu Ni Zn 1.5 x x x e* D0%%:2T D0%%:7 %(5,0 =$ Metal concentration (mg/l) Washing waters 1 st 2 nd 3 rd Cu Ni Zn * D0%%:7 492 "% / 8 " "! = " " 4;* & B G:H :49

218 H 1"-. TG6HTG9H TG2H! = %:2TG5H= %&:7 Effluent with Cu, Ni and Zn NaOH Effluent with Cu, Ni and Zn at ph 6 Heat-inactivated cells of S. cerevisiae Chapter 6 Treated Effluent Yeast cells loaded with heavy metals Incineration at 550ºC Ashes Acid digestion Acid solution with Cu, Ni and Zn NaOH Solution with Cu, Ni and Zn at ph 2 Electrolysis at controlled potential Cu (s) (Recovery: 99 %) (Purity: 99.9 %) Solution with Ni and Zn at ph 2 NaOH (ph 14) Baths of zinc-nickel alloys Ni(OH) 2 (s) (Recovery: >99 %) (Purity: 92 %) Solution of Zn(OH) 2-4 at ph 14 HCl Zn(OH) 2 (s) (Recovery: >99 %) (Purity: 99.4 %) + 5,2! "! = :42

219 H 1"-. * "!= M!=$"T"! " 5,0,!" " " GH!G!=$H * " 2 " R4S $ "G"!=H :37 * M " "!= GVRRSH GRRR"R6RR2S"!="H* = G MMH :45

220 H 1"-. 8 " A G:R32H) ) 942M94R?B?A? "'G:RR2H 0 '8 D N!B)A M %!"?"D"Z"!""G6777H'" " P $ $02G6M9H"92RM95; =""*"**$">G6772HGH?$1"::M67!"*" "*" " Z" Z&" N")"% " G6772H "PD ++*/G6H"663M696 %"% G:RR2H 0" Z 473M4:7D N!B>f o %">?""A="G677;H8 " 2 5/G:M2H":7;M::9?MA@" >?? ") G6779H f M Mq$ $ ""G6H"R7MR4 )""')G6772H-)* & 46A H8" -) & B "D* ")" ")"!" A " " " ")? " > G677;H B* )? 34G:H"63M2: :4;

221 H 1"-. "Af"">">'G:R4:H) B $0$"693M625 =!"'"?"f%" =")D "ZG:RR5HM 5"G:6H"54M;5 *!"%%""'"8!">"Z "ZA"'G6779H B: )! $0G;H"269M29; *"G:RR9H 1 2:?Bo% *!"%"Z"")"D"Z ")")"%!"*G6774H! " P" D %6 M M 3/GRH":224M :256 M%% G6775H D!!* 0 2 "- B %% f "8">'G:RR5H)! $3GRH":773M:7:6 f " 8 " >' G:RR;H' M! $5G5H"59:M59; :43