torres et al. (1997) - amino acid chronology of the lower pleistocene deposits of venta micena...

8/7/2019 TORRES et al. (1997) - Amino acid chronology of the Lower Pleistocene deposits of Venta Micena (Orce, Granada, A

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PergamonPII: S0146-6380(96)00131-3

Org. Geochern.Vol. 26, No. 1/2, pp. 85 -97, 1997 1997 ElsevierScience Ltd

All rights reserved.Printed in Great Britain0146-6380/97 $17.00 + 0.00

Am ino acid chronology of the Lower Pleistocene deposits of Venta

Micena (Orce, Granada, Andalusia, Spain)T. T O R R E S , J . F. L L A M A S , L . C A N O I R A , P. G A R C I A - A L O N S O , A . G A R C I A -

C O RT E S a n d H . M A N S I L L AEscuela T6cnica Superior de Ingenieros de Mina s de M adrid, R ios Rosa s 21, 28003 Madrid, Spain

Ab strac t--Th is pap er deals with a mod el adjustment fo r the estimation of age by means of amino acidracemization analysis. Two model families were obtained on the basis of the different genera of mol-luscs analysed, and were applied to a palaeontological site located in the Cfillar-Baza basin: "VentaM icena" (Orce, Gr anad a). The ana lytical results obtained from the stud y of fossil gastropods have pro -vide d a ve ry coherent average dati ng o f 983 ___ 58 Ky , this co inciding to a large extent w ith the mo stwidespread palaeontological, geological and stratigraph ical datings for the site. There is, however, noagreement with certain recent theories that situate Venta Micena chronologically at the Pliocene-Pleistocene bound ary (ca. 1.6 My ) or before. The v alidity of the da ting fit was tested on the basis of

anoth er classical pala eontolog ical site, C611ar-Baza, where a coherent dating of 441 + 27 Ky wasobtained. 1997 Elsevier Science Ltd

K e y w o r d s - - a m i n oacid racemization, a minoc hronostra tigraphy, aminochronology, Lower Pleistocene,M iddle Pleistocene

INTRODUCTION

O n e o f t h e m o s t i m p o r t a n t p r o b l e m s i n v o l v e d i nP l e i st o c e n e p a l a e o e n v i r o n m e n t a l a n d p a l a e o c l i m a t o -l og i ca l s t ud i e s i n Spa in , and o the r p a r t s o f t hew or ld , i s t ha t o f p l ac ing t he obse rva t i o ns on ach r onos t r a t i g r aph i ca l s ca l e a l l owing t he ve loc i t y o ri soch ron y o f t he p roce s se s t o be e s t ab l i shed .Tra d i t i o na l l y, r e l a t i ve da t i ng ha s been accomp l i s hedt h r o u g h g e o m o r p h o l o g i c a l , p a l a e o n to l o g i c a l a n da rc ha eo lo g i c a l s t ud ie s i n wh ich fo rms , f o s s il s a nda r t i f a c t s w e re p l aced on t he s t r a t i g r ap h i ca l s ca l eacc o r d i ng t o (more o r l e s s ) sub j ec t i ve ana ly s i s , i . e .t he r e l a t i ve t opog raph i c a l l eve l o f f l uv i a l t e r r ace s o rg l ac i a l mo r a ine s , t he evo lu t i ona ry s t age o f f o s s i lr em a ins , a c co rd ing t o an a s sumed ve loc i t y o f evo l -u t i on o r t he morpho log i ca l cha r ac t e r i s t i c s o f s t onea r t i f a c t s . Obv ious ly t he se me thods do n o t wo rkaccu ra t e ly b e c ause i n mos t c a se s t he r e i s i n f a c t af ee dbac k p r o c e s s : f o rms and depos i t s a r e u sed t op l ace f o s s il and a r chaeo log i ca l r ema in s on t h ech ron os t r a t i g r aph i ca l s ca l e and l a t e r p r ov i de t hec lue f o r d a t i ng o f t he p roce s se s .

R a d i o m e t r i c s . L m e t h o d s ( 1 4 C , U - s e ri e s, K / A r,ESR, TL . . . ) a r e pa r t i cu l a r l y u se fu l t o o l s f o r da t i nga nd a r e w ide ly u sed i n sp i t e o f t he r ange m e thodl i m i t a t i ons ; howeve r, t he mos t f r equen t p r ob l e m i sr e l a t ed t o t h e pos s ib i l i t y o f f i nd ing s amp le s f o r da t -i ng pu rp os e s (we igh t o r na tu r e ) . The amin o ac i dr a c e m i z a t i o n d a t i n g m e t h o d p r o v i d e s c e rt a i n a d v a n -t ages : o n l y mino r s amp l ing p roce s se s a r e r equ i r ed( 80 m g m a x i m u m ) a n d a w i d e g r o u p o f m a t e r i a l s

a r e a v a i l ab l e f o r da t i ng , e . g. she l ls f r om m o l l u sca

(Goodfr iend , 1987; Wehmil le r, 1993) , foramini fera( Wehm i l l e r, 199 3 ) , o s t r a co da (To r r e se t a l . , 1995)and e ggs ( Mey e r, 1992 ) ; bon es and t e e th ( Ba dae ta l . , 1973a; Marzin, 1990; Elstere t a l . , 1991), wood( Ru t t e r and V l aho s , 1 988) , spe l e o th e m e s (Lau r i t z ene t a l . , 1994) , o r even rocks and sof t sed iments

( H e a r t ye t a l . , 1992).Th e a m i no a c i d an a l y s i s o f f o s s i l s was i n i t i a t ed

m a ny ye a r s a go (Ab e l s on , 1954 ) a nd ha s s i nc e b eencon t i nued b y many r e se a r ch e r s ( H a re , 1 96 9 , 19 7 l ;Bad a an d P r o t ch s , 1 973 ; B a da a nd S c h r oed e r, 1972 ;M i l l e r a nd H a re , 197 5) . To da y, a m i no a c i d r acemi -za t i o n r a t i o ana ly s i s makes i t po s s ib l e t o d e t e rminet h e p a l a e o c l i m a t o l o g i c a l d a t a o f a r e g io n ( B a d ae ta l . , 1973b; Wehmil le r, 1993) . Amino ac id racemiza-t i on ana ly s i s a l so a l l o ws amino s t r a t i g r a phy ( Mi l l e rand Ha re , 19 80 ) a nd amin och ro no l ogy t o be e s t ab -l i she d . Amin os t r a t i g r aphy cons i s ts o f a r r a ng ing geo -logica l s ites in to a sequen ce on th e bas is ofob s e rv ed c lu s t e r s o f r a c em i z a t i on r a t i o s , e s t ab l i shedus in g r ep re sen t a t i ve s o f on e zoo l og i c a l g e nus. T h i sm e t hod i s an ex c e l l en t t o o l f o r t he co r r e l a t i on o fs e a l eve l o sc i l l a t i on - l i n ked dep os i t s - -m ar i n e an df luv i a l t e r r ace s i nd i ca t i v e o f wa rm and s t ab l e c l i -m a t e p e r i o ds (Mi l l e r a nd M a nge ru d , 1985;K a u f ma n , 1992 ; H e a r tye t a l . , 1992 ) - - an d i s e s -pec i a l l y u se fu l f o r t he de t e r m in a t i on o f neo t ec to n i c sa f f ec t i ng P l e i s t o c ene d e po s i t s (D umase t a l . , 1988).I n so me ca se s , an d ma in ly w he r e a lmos t con t i nuoussedimenta t ion has taken p lace , as in the C611ar-Baza Ba s i n , i t i s ne c e s s a ry t o p roduce num er i ca l -

age r e su l t s , t h e mos t com m on be ing age -ca l i b r a t ed85


2/13

86 T. Torres e t a l .

( 14 C o r U - s e r i e s ) r e s u l t s ( G o o d f r i e n d , 1 9 87 ; D u m a se t a l . ,1988; Hear tye t a l . ,1992; Wehmi l le r, 1993) .

S o m e y e a r s a g o , Ve n t a M i c e n a , a s m a l l - h o l d i n gl o c a t e d c lo s e to O r c e i n G r a n a d a , w a s w i d e l y m e n -t i o n e d i n t h e m a s s m e d i a a s a r e s u l t o f a c o n t r o v e r -s ia l f i n d in g o f h u m a n / n o n - h u m a n r e m a i n s ( G i b e r te t a l . ,1989, 1992b) . I t i s no t o ur ob jec t ive he re tog e t i n v o l v e d i n p a l a e o a n t h r o p o l o g i c a l q u e s t i o n s,b u t r a t h e r t o p l a c e t h is p a l a e o n t o l o g i c a l s i te o n aP l e i s t o c e n e t i m e - s c a l e u s i n g t h e a m i n o a c i d r a c e m i -z a t i o n d a t i n g m e t h o d . T h i s s i t e , a l o n g w i t h m a n yo t h e r s i n t h e C f i l l a r - B a z a B a s in , w a s d a t e d u s i n gt h i s m e t h o d d u r i n g a l e n g t h y d a t i n g c a m p a i g n c a r -r i e d o u t f o r d r a w i n g u p o f t h e " I n s t i t u t oTe c n o l 6 g i c o G e o m i n e r o d e E s p a f i a " 1 : 50 0 00 g e o -l o g i c a l m a p ( To r r e s , 1 9 95 ).

E X P E R I M E N TA L

G e o g r a p h i c a l s i t u a t io n a n d p a l a e o n t o l o g i c a l - g e o l o g i -c a l s e t t i n g s

Ve n t a M i c e n a i s a w e l l - k n o w n p a l e o n t o l o g i c a ls i t e o f U p p e r Vi l l a f r a n c h i a n a g e ( F i g . 1 ) l o c a t e d i nt h e G u a d i x - B a z a B a s i n . T h i s b a s i n c o n s i s t s o f al o n g , n a r r o w t e c t o n i c a l l y c o n t r o l l e d d e p r e s s i o n , t h em a j o r a x i s o f w h i c h i s o r i e n t e d in a N E - S W d i r ec -t i o n . T h e G u a d i x - B a z a B a s i n c o n s t i t u t e s a n e x c e p -t i o n t o t h e b e h a v i o u r o f I b e r i a n P e n i n s u l a

P l e i s t o c e n e b a s i n s : s i n c e a t t h e e n d o f P l i o c e n et i m e s t h e y w e r e a l l w o r k i n g u n d e r a n i n c i s i o n - d o m i -n a t e d r e g i m e , a n d i n s p i t e o f s h o r t - t e r m a l l u v i a lp l a t f o r m b u i l d i n g , t h e e r o s i o n c o n t r o l l e d b y f a l l i n gs e a l e v el w a s t h e o r i g i n o f i n c o m p l e t e s t r a t i g r a p h i c a ls e c t i o n s w i t h a l l u v i a l o r l a c u s t r i n e t e r r a c e d d e p o s i t s ,r e p r e s e n t i n g o n l y w a r m ( s t a b l e ) i n t e r v a l s . D u r i n gt h e P l i o c e n e a n d m o s t o f t h e P l e i s t o c e n e , t h eG u a d i x - B a z a B a s in o p e r a t e d a s a c l a s si c e n d o r h e i cb a s i n , a n d t h e r e w a s s t a c k i n g o f a th i c k , a l m o s tc o n t i n u o u s a l l u v i a l - l a c u s t r i n e - p a l u s t r i n e s e qu e n c e.T h e c o n t i n u i t y o f t h e G u a d i x - B a z a B a s i n ' sP l i o c e n e - P l e i s t o c e n e g e o l o g i c a l r e c o r d , a n d t h el a r g e n u m b e r o f p a l a e o n t o l o g i c a l l o ca l it i es p r e -s e r v e d c o n s t i t u t e a r e m a r k a b l e e x c e p t i o n . S o m ea u t h o r s ( A n a d 6 ne t a l . ,1987; Agus t i , 1984 ; Alberd ie t a l . , 1 98 9 ) h a v e e s t a b l i s h e d n u m e r o u s c h r o n o -s t r a t ig r a p h i c a l s e q ue n c e s b a s e d o n p a l a e o n t o l o g i c a ld a t a b u t w i t h o u t n u m e r i c a l d a ti n g .

P l i o c e n e - P l e is t o c e n e s e d i m e n t s a p p e a r a t t h e S Wl i m i t o f t h e b a s i n a n d a t i t s n o r th e r n b o u n d a r y( I T G E , 1 98 9 ). L o w e r P l e i s t o c e n e d e p o s i t s a r e w e l lr e p r e s e n t e d i n t h e c e n t r a l p a r t , w h e r e a s m i d -P l e i s t o c e n e m a t e r i a l s a r e t o b e f o u n d a t i t s s o u t h e r nl i m i t . T h e O r c e s i t e h a s b e e n w r o n g l y p l a c e d i n aM i d d l e P l e is t o c en e d a t e d o u t c r o p .

T h e P l e i s t o c e n e s e d i m e n t a t i o n o f t h e O r c e ( Ve n t aM i c e n a ) a r e a ( F i g . 2 ) c o n s i s t s o f f i n e - g r a in e d d e t r i -

P I / ,

~ ~ _ t . _ . . ~ . . " '~ "i f . " / i I . . ' 1 -

. . . . . . . ( t, ~ / / / _ / / / /

, / / / / / / /

x ' .g .g X "g x" x" ~ A ln ler lo

~ ~ ~ , ~ , < ,~ 1 2 0 K i n I

Plio-Pleistocene Lower-Middle Middle Other Pleistocene Mountain RangePleistocene Pleistocene Basins (Sierra)

i .

Fig. 1. Geo grap hical and geological situation of Orce (Venta M icena) area (ITG E, 1989, modified).


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Am ino acid chronology of Venta Micena deposits 87

N

U - - - l s4

m 3

- 7

1 K m 1

Fig. 2. Geolog y of the Venta M icena (VM ) site (Soriaet al . , 1987, modified). (1) M esozoic rockthreshold; (2) fluvial-alluvia l fan Pleistocene deposits; (3) palustrine lacustrine car bon ate deposits withVenta Micena (VM) palaeontological site; (4) Middle and Upper Pleistocene alluvial deposits; (5)

Holocene depo sits.

t a l d e p o s i t s a t t h e l o w e r p a r t ( S o r i aet al., 1987),f o l lo w e d b y c a r b o n a t e d e p o s i t s o f l a c u s t ri n e o r i g in .T h e l o w e r- m i d d l e P l e i s t o c e n e s e d i m e n t s w e r eu n c o n f o r m a b l y c a p p e d w i t h r e c e n t g r a ve l s o f a l lu -v i a l o r i g i n . M e s o z o i c l i m e s t o n e o u t c r o p s a c t e d a sl o c a l t h r e s h o l d s d u r i n g P l e i s t o c e n e d e p o s i t i o n . T h eb o t t o m s o f t h e v a l l e y s a r e f i ll e d w i t h r e c e n t( H o l o c e n e ) d e p o s i t s .

T h e Ve n t a M i c e n a s e c t i o n ( A n a d 6 net al . , 1987)( F i g . 3 ) m a y b e d e s c r i b e d a s f o l l o w s :

( a ) d o l o m i t i c m a r l s , d o l o s t o n e s a n d c h a n n e l l e dg r a v e l a n d s a n d ;

( b ) lu t i te s a n d m a r l s w i th a b u n d a n t o r g a n i cd e b r i s c o n t a i n i n g f o r a m i n i f e r a , g a s t r o p o d a a n do s t r a c o d a , t h a t h a v e b e e n s a m p l e d f o r a m i n oa c i d r a c e m i z a t i o n d a t i n g ;

( c ) s l i g h t l y s a n d y l u t i t i c l i m e s t o n e s - - t h e Ve n t aM i c e n a p a l a e o n t o l o g i c a l si te a p p e a r s a t t h e t o po f t h i s s t r a t u m ;

( d ) p a r t i a l l y c o v e r e d , l u t i t i c l im e s t o n e s a n dm a r l s , d o l o s t o n e s a n d d o l o m i t i c m a r l s - - c h a n -n e l le d g r a v el a n d s a n d a r e c o m m o n ;

OG 26/1 2 D

( e ) n o d u l o u s l u t i t i c l i m e s t o n e s a n d l i m e s t o n e sw i t h g a s t r o p o d a c a s t s.

T h e g e n e si s o f t h e Ve n t a M i c e n a p a l a e o n t o l o g i c a ls i t e ( c ) m a y b e i n t e r p r e t e d a s a m a rg i n a l d e p o s i t o ft h e C f i l l a r - B a z a B a s i n w h i c h o c c u r r e d d u r i n g al a c u s t r i n e e x p a n s i o n e v e n t o v e r l a p p i n g a p r e v i o u sm u d f l a t [ ( a ) a n d ( b ) ] ; t h i s l a c u s t r i n e e n v i r o n m e n tw a s t e m p o r a r i l y i n t e r r u p t e d b y c h a n n e l l e d d e p o s i t s(d) .

T h e Ve n t a M i c e n a v e r t e b r a t e a s s o c i a t io n i s m a d eu p o f t h e f o l l o w i n g : U r s i d a e(Ursus e t ruscus) ,C a n i d a e (Canis e t ruscus mosbachens i s , Cuon pr i scus ,Vulpes praeglacial is , Xenocyon s p . ) , F e l i d a e(Homother ium la t idens , Megante reon cu l t r idens adro-ver i , Lynx sp. , Panthera cf. gombaszogens i s ) ,H i e n i d a e ( Pac hycro cuta brevirostr is ruizi) ,P e r i s s o d a c t y l a(Equus s tenonis , Dicerorh inus e t rus -cus , Hippopotamus incogni tus ) , P r o b o s c i d e a( Arch id i skodon mer id iona li s )a n d A r t i o d a c t y l a( Me gaceros so lh iacus , Cervus (9.) elaphoides,Preaeovivossp. , Capra alba, Soergel ia minor, Bisons p . ) , a s w e l l a s t h e c o n t r o v e r s i a lH o m o sp . r emains .T h e r e a r e a l s o r o d e n t a n d l a g o m o r p h r e m a i n s


4/13

88 T. Torre set aL

fm

@

E LimestoneLutiticlimestone

F ~ Dolostone

Lutitic dolostone

[ '~ Dolomitic marl

Lutite

Sand

Gravel

d

eM

C

b

a

~ R D

Fig. 3. Stratigraphical section of Venta M icena area(Anad6net al., 1987, modified). VM fossil mamm als b ear-ing s trata . AARD sampled bed for amino acid racemiza-

t ion dat ing.

( A g n s t i et al. (1987)) : Allophaiomys pliocaenicus,Castillomys crusafonti, Apodemusaft . mystacinus,Eliomis intermedius, Hystrix majora n d Prolagus cf.calpensis; a n d m a n y m o l l u s c a g e n e r a :Melanoides,Melanopsis , Pomatias, H yd ro bia , Me rcur ia ,Hauffenia, Bithynia, P lano rbis, Anisus, G yraulus,Armiger, Ancylus, Acroloxus, Lymnaea, Cochlicopa,Vertigo, Pupilla, Helicellaa n d Pisidium, cf.R o b l e set al. ( 19 8 7) . F o r a m i n i f e r a a n d c r u s t a c e a a r e a l s of r e q u e n t .

A c c o r d i n g t o p a l a e o n t o l o g i c a l as s e m b l a g e s,a n dt o t h e g e o m o r p h o l o g i c a l c h a r a c te r i s ti c s a n d p a l a e o -m a g n e t i s m o f th e a r e a , Ve n t a M i c e n a ( A l b e r d ietaL , 1 98 9 ) w a s d a t e d n e a r 1 .3 M y . L a t e r ( G i b e r tetal., 1 9 92 a) a n a g e o f 1 .6 M y w a s e s t a b l i s h e d .

Sampling

A 5 - k g s a m p l e w a s t a k e n f r o m a l u t i t i c s t r a t u ml o c a t e d i m m e d i a t e l y b e l o w t h e V e n t a M i c e n a f o ss il i-f e r o u s h o r i z o n ( F i g . 3 ) . G a s t r o p o d a e x t r a c t i o n w a sa c c o m p l i s h e d i n a c c o r d a n c e w i t h t h e e s t a b l i s h e dp r o c e s s ( To r r e set al., 1 9 9 4 ) . Tw e n t y - t h r e e i n d i v i d -

u a l s a m p l e s w e r e p i c k e d o u t f r o m t h e s i e v e d s a m p l ea n d a n a l y s e d : t h e y p r o v e d t o b e s i xGastropodainde t , ope rcu la ( B i t h y n i a s p . ) , t h r e e Radix(Lymnaea) sp . , one Planorbis sp . , one Bithynia sp .a n d 1 2 Fragmenta indet.( p r o b a b l yBithynia sp . )

Sample preparation and analysis

T h e g l a s s w a r e u s e d f o r a n a l y s i s ( e x c e p t t h eP a s t e u r p i p e t t e s) w a s c l e a n e d b y b a k i n g i n a n o v e na t 5 0 0 C f o r a b o u t 2 h . E p p e n d o r f p l a s t ic m i c r ot e s t t u b e s , p l a s t i c m i c r o p i p e t t e t i p s a n d P a s t e u r p i p -e t t e s w e r e n e w f r o m t h e f a c t o r y. Te f l o n l i n e r s a n ds e p t a w e r e t h o r o u g h l y w a s h e d w i t h p e t r o l e u m e t h e r

a n d a c e t o n e a n d r i n s e d t h r e e t i m e s w i t h u l t r a c l e a nw a t e r. A l l t h e w a t e r u s e d i n t h e a n a l y s i s w a s M i l l i -Q q u a l i t y f ro m M i l l i p o r e. C o n c e n t r a t e d h y d r o f l u o -r i c a n d h y d r o c h l o r i c a c i d s a n d t r i f l u o r o a c e t i c a c i da n h y d r i d e w e r e M e r c k a n a l y t i c a l g r a d e . T h i o n y lc h l o r i d e w a s p u r c h a s e d f ro m F l u k a A G . I s o p r o p y ia l c o h o l a n d n - h e x a n e w e re M e r c k H P L C g r a d e ,a n d d i c h l o r o m e t h a n e w a s M e r c k s p e c t r o s c o p yg r a d e .

A l l t h e s h e l l s u s e d f o r a m i n o a c i d r a c e m i z a t i o na n a l y s i s w e r e t h o r o u g h l y c l e a n e d b y a c o m b i n a t i o no f u l t r a s o n i c a t i o n a n d m e c h a n i c a l c l e a n i n g w i t h ad e n t a l d r i l l , a n d r i n s e d t h o r o u g h l y w i t h d e i o n i z e d

w a t e r , 2 N h y d r o c h l o r i c a c i d a n d a g a i n w i t h d e i o n -i z e d w a t e r. H y d r o l y s i s o f c a. 8 0 m g o f s h e l l w a s c a r -r i e d o u t i n a m i x t u r e o f 1 2 N h y d r o c h l o r i c a c i d(2 .9 /~1 mg -1 ) a nd 6 N hy dro ch lo r ic ac id (100 /~1), int e s t t u b e s w i t h Te f l o n - l i n e d s c r e w c a p s c l o s e d i n a na t m o s p h e r e o f n i t r o g e n , i n a h e a t i n g b l o c k a t 1 00 Cf o r 2 0 h . D e s a l t i n g w a s a c c o m p l i s h e d i n c o n i c a l1 .5 m l E p p e n d o r f p l a s t i c m i c r o t e s t t u b e s w i t h c a p s ,w i t h c o n c e n t r a t e d h y d r o f l u o r i c a c i d a d d e d( 1.2 5 /~ 1 m g - l o f s a m p l e ) , t h i s b e i n g m i x e d w i t h am e c h a n i c a l Vo r t e x s h a k e r, a n d c e n t r i f u g e d f o r4 m i n i n a n E p p e n d o r f c e n tr i fu g e . T h e s u p e r n a t a n tw a s t r a n s f e r r e d t o n e w 1 .5 m l E p p e n d o r f m i c r o t e s tt u b e s , f r o z e n i n l i q u i d n i t r o g e n , a n d v a c u u m d r i e di n a p l a s t i c d e s i c c a t o r. S a m p l e s w e r e r e d i s s o l v e dw i t h 8 0 # 1 d i s t i ll e d w a t e r, m i x e d i n t h e V o r t e x s h a -k e r, c e n t r i f u g e d f o r a f e w s e c o n d s t o e n s u r e t h e d e -p o s i t i o n o f a l l th e d r o p l e t s , a n d t h e n t r a n s f e r r e d t o2 m l g l a s s v i a l s w i t h s c r e w c a p s a n d Te f l o n - l i n e ds e p t a . Wa t e r w a s e v a p o r a t e d a t v a c u u m f r o m t h ev i a l s i n t h e p l a s t i c d e s i c c a t o r, t h e c a p s n o t b e i n gt i g h t l y c l o s e d .

T h e f i r s t a m i n o a c i d d e r i v a t i z a t i o n s t e p c o n s i s t e do f e s s t e r i f i c a t io n w i t h 2 5 0 /~ 1 o f 3 M t h i o n y l c h l o r i d ei n i s o p r o p a n o l . T h e v i a l s w e r e t i g h t l y c l o s e d i n an i t r o g e n a t m o s p h e r e a n d l e f t t o r e a c t o n t h e h e a t i n gb l o c k a t 1 0 0 C f o r j u s t 1 h . T h e t o p s o f t h e v i a l s


5/13

Amino acid chronology of Venta Micena deposits 89

were then unscrewed (but not removed) in a hoodand vacuum dried in a plastic desiccator just to thepoint of dryness. The second derivatization stepconsisted o f N-trifluoroacetylation with 150 #1 oftrifluoroacetic acid anhydride (25% in dichloro-methane). The vials were tightly closed in a nitroge natmosphere a nd heated at 100C for just 5 min onthe heating block. They were subsequently allowedto cool and opened in a hood, where the dichloro-methane solvent and the unreacted trifluoroaceticacid anhydride were evaporated under a gentle flowof nitrogen. Later the samples were dissolved in125/~1 of n-hexane, shaken in the vortex, and mostof the n-hexane evaporated in a stream of nitrogento a final volume of 15-25/A, then being transferredto 150 #1 inject ion vials.

One pl o f sample was injected into a Hewlett-Packard 5890 gas chromatograph. The injection

port was kept at 215C and set for splitless modefor the first 75 s, at the b eginni ng of which thesample was injected, and later set to split mode. Weused helium as the carrier gas at a head columnpressure of 5.8 psi, an d a Chirasil-V al fused silicacol umn (25 m x 0.39 mm x 0.25 mm) from Chro m-pack. The programme we used was as follows: 50C(1 min), he at at 40C min -l to 115C, rema inin g at115C for 12min, heating at 3Cmin -l to 190C,remainin g at 190C for 1 0min, cooling down to50C and r emaining at this temperature betweenruns (80C if the time between runs is longer, typi-cally overnight). The detector was a FID set at300C. Integration of the peak areas was carriedout using the PEAK96 integration program fromHewlett-Packard, which runs on a PC computer.44 shows a typical gas chromatogram of a sample.The sensitivity limits of the meth od may be fixed

, . . 4

"7

o, .

i , ~ ! !~ ! ] ! I !

a _

i ! f ' i "

i ! i i ' *Fig. 4. Typical chromatogram from a Bulimus sp. sample from Venta Micena.


6/13

90 T. Tor tes et al.

Table 1. Comparision of D/L ratios between the interlaboratory results exercise and the results obtained by theamino ac id racemization laboratory of E.T.S.I., Minas

D/L [sample (powder)]

Amino acid ILC-A ILC-B ILC-C

A/ I ILR 0.212 0.072 0.54 +_ 0.0162 1.215 0.030LAB 0.180 0.650 1.245

PRO ILR 0.278 0.068 0.595 + 0.210 0.81 0.26

LAB 0.195 0.507 0.786LE U ILR 0.196 0.042 0.497 + 0.098 0.833 __+0,086LAB 0,182 0.444 0.849

ASP ILR 0.378 0.056 0,705 0,056 0.894 0,158LAB 0.373 0.728 0.914

PHE ILR 0.239 0.040 0,583 0.059 0.873 0.178LAB 0.220 0.608 0.885

GLU ILR 0.203 0.022 0.432 0.034 0.849 0.070LAB 0.185 0.426 0.832

ILR = Interlaboratory results (confidence interval was calculated as 2tr).LAB = laboratory E.T.S.I., Minas.

e i t h e r i n a c c o r d a n c e w i t h t h e r a c e m i z a t i o n i n d u c e d

b y th e m e t h o d ( 1 - 5 % , Ta b l e 2 , " To d a y " r o w ) o rw i t h th e m i n i m u m c o n c e n t r a t io n o f a m i n o a c i dd e t e c t e d , w h i c h i s n o t a l i m i t i n g f a c t o r ( i t d e p e n d so n t h e a g e o f t h e s a m p l e s , s i n c e it a ff e c t s t h e m o r er e c e n t s a m p l e s i n w h i c h t h e D - a m i n o a c i d m i g h t n o tb e d e t e c t e d ) . I n a l l t h e s a m p l e s a n a l y s e d t o d a t et h i s p r o b l e m h a s b e e n s o l v e d b y w e i g h i n g a l a r g e ra m o u n t o f t he s a m p le .

Dating method cal ibrat ion

A f t e r s e t t i n g u p t h e a n a l y t i c a l m e t h o d i n o u r l a b -o r a t o r y, w e w o r k e d t o o b t a i n a n e x t e r n a l v a l id a t i o no f t h e m e t h o d i n o r d e r t o c o n t r o l a n y k i n d o f sy s -

t e m a t i c e r r o r t h a t m i g h t i n t r o d u c e a b i a s i n t h er e s u l t s . T h r e e s a m p l e s f r o m a n i n t e r l a b o r a t o r ya m i n o a c i d r a c e m i z a t i o n e x e r ci s e ( We h m i l l er , 1 98 4)w e r e a ls o a n a l y s ed : I L C - A ( S a x i d o m u s sp. ca .5 0 K y ) , I L C - B (Mercenar ia sp . be tween 100 and2 5 0 K y ) a n d I L C - C (Mercenar ia sp . ca 1 .000 Ky) .T h e s e s a m p l e s w e r e a n a l y s e d b y 11 la b o r a t o r i e s i na n e x e r c is e o f i n t e r l a b o r a t o r y c o n t r o l . T h e l a c k o fb i a s a n d t h e v a l i d i t y o f t h e p r o c e s s f i t p r o v i d e d b yo u r l a b o r a t o r y w e r e c o n f i r m e d ( Ta b l e 1 ) .

S a m p l e s o f m o l l u sc s f r o m U / T h m e t h o d d a t e dP l e i s t o c e n e f l u v i a l t r a v e r t i n e t e r r a c e s i n t h e P r i e g oa r e a ( To r r e s et al., 1 9 9 4 ) w e r e c o l l e c t e d , p r e p a r e d

a n d a n a l y s e d . T h e a g e o f t h e s a m p l e d t e r r a c e s r a n -g e d b e t w e e n 6 a n d 113 K y. I n o r d e r t o c a l c u l a t e t h er a c e m i z a t i o n i n d u c e d d u r i n g s a m p l e p r e p a r a t i o n ,s e v e n s a m p l e s o f g a s t r o p o d s c o l l e c te d l i v e w e r e a l s o

p r e p a r e d a n d a n a l y s e d . Wi t h a v i e w t o c o v e r i n g

o l d e r g e o l o g i c a l r e c o r d s,D/L

a m i n o a c i d r a t i o s f r o m1 00 to 2 5 0 K y a n d c a . 1 0 0 0 K y o l d l oc a l it i es( We h m i l l e r , 1 98 4) w e r e a l s o u s e d .

S p u r i o u s r e s u l t s w e r e o b s e r v e d d u r i n g i n t e r p re t -a t i o n o f c h r o m a t o g r a p h i c a n a l ys i s o f t h e s am p l e sf r o m P r i e g o : a n o m a l o u sO/L r a t i o s i n s a m p l e s f r o mt h e s a m e s t r a t i g r a p h i c l e v e l ( t r a v e r t i n e t e r r a c e ) . A l lt h e o b s e r v e d d e v i a t i o n s a p p e a r e d a s a n e x c e ss o f D-a m i n o a c i d s a n d s e e m e d t o b e r e la t e d t o t h eo b s e r v e d a l g a e - f u n g i m a t c o v e r i n g s o m e o f t h es a m p l e d g a s t r o p o d s h el ls . S o m e b a c t e r i a h a v e D -a m i n o a c i d s i n t h e i r c e l l u l a r w a l l s ( L e i v e a n d D a v i s ,1 9 80 ) a n d a l g a e a n d f u n g i m a t s a r e u s u a l b a c t e r i a

h o u s e r s . B y t h e t i m e Ve n t a M i c e n a w a s s a m p l e d ,w e h a d a l r e a d y m o d i f i e d o u r r e c o v e r y p r o c e s s i no r d e r t o r e d u c e t h e d a n g e r o f c o n t a m i n a t i o n : t h es a m p l e s w e r e o b t a i n e d f r o m d e e p e r p a r ts o f th es t r a t a w h e r e t h e r e is n o i n f l u en c e b y t h e a t m o s p h e r ea n d s u n l i g h t . I n o r d e r t o a v o i d t h e i n f lu e n c e o f t h es p u r i o u s v a l u e s , a p r e v i o u s s i n g l e r e g r e s s i o n a n a l y -s i s o f t h e D - a n d L - a m i n o a c i d p a i r s i n t h e s a m p l e sw a s p e r f o r m e d . Ta b l e 2 s h o w s t h e a v e r a g e r at i o s o fr a c e m i z a t i o n o f P r ie g o s a m p l e s .

F i n a l ly, p r e d i c t i o n m o d e l s w e r e c a l c u l a t e d u s i n ga s e t o f 3 0 U / T h - d a t e d s a m p l e s f r o m P r i e g o : 2 1f r e s h w a t e r g a s t r o p o d s , s i x t e r r e s t r i a l g a s t r o p o d s

a n d t h r e e f r e s h w a t e r p e l e c i p o d a , a s w e l l a s t h r e es a m p l e s o f a n i n t e r l a b o r a t o r y c o m p a r i s o n e x e rc i s e( We h m i l l e r , 1 9 84 ) . T h e r a c e m i z a t i o n m o d e l s a r eb a s e d o n a f i r s t - o r d e r r e v e r s i b l e r e a c t i o n ( B a d a a n d

Table 2. AverageD/L ratios of m ollusc samples from U /Th-dated travertine terraces in P riego (Cuenca, Central Spain)

AGE average (U/Th) A/I Leu D/L Asp D/L Pile D/L Glu D/L

To day 0.000 + 0.000 0.008 + 0.0002 0.051 0.008 0.019 0.011 0.016 0.0126.000 0.041 0.008 0.215 + 0.051 0.029 0.044 0.028 0.02212,500 0.149 0.078 0.071 0.018 0.257 + 0.019 0.079 0.009 0.070 0.0062 0 , 0 0 0 0 . 1 3 3 0 . 0 0 9 0 . 3 4 6 + 0 . 0 4 0 O . 1 2 1 0 . 0 3 1 0 . 1 0 5 0 . 0 1 3

1 0 5 , 0 0 0 0 . 2 3 7 + 0 . 0 1 6 0 . 4 2 3 0 . 0 2 6 0 . 5 9 4 0 , 0 2 6 0 . 4 7 9 0 . 0 4 0 0 , 2 5 2 0 . 0 3 1


7/13


P r o t c h s , 1 97 3 ; S c h r o e d e r a n d B a d a , 1 97 6 ; B a d a ,1 9 8 5 ) , t h e a l g o r i t h m s b e i n g b a s e d o n a t i m e s q u a r e -r o o t ( x / t ) a d j u s t m e n t ( G o o d f r i e n d , 1 9 8 7 ) . T h i sw o u l d a p p e a r t o b e j u s t i f i e d s i n c e i t s u s e s t a b i l i z e st h e v a r i a n c e o f t h e e r r o r t h a t , i n t h e c a s e o f t im e ( t )a d j u s t m e n t , b e c o m e s p r o g r e s s i v e l y g r e a t e r w i t hg r o w i n g a g e s a m p l e s .

T h e m o d e l s o b t a i n e d a r e a s f o l lo w s :

L EU ~/t - - - (1 .17 4- 0.62) + (11.38 4- 0.88)

x I n { [1 + ( D / L ) ] / [ 1 - ( D / L ) ] } ,

cor re la t ion coeff i c ien t = 0 .9774 (n = 33)

AS P x / t - - - ( -2 .17 4 - 1.02) + (10 .02 4 - 0 .85)

x In{J1 + (D/L ) ] / [1 - (D/L) ]} ,

cor r e la t io n coeff i c ien t = 0 .9740 (n = 32)

PH E ~ / t = (0 .99 4 - 0 .78) + (10 .26 4 - 0 .74)

ln{[ l + (D /L) ] / [1 - (D /L) ]} ,


G L U ~/ t = (2 .16 + 0 .65) + (12 .44 4 - 0 .82)

x ln{[ l + (D /L) ] / [1 - (D/L) ]} ,


T h e a l g o r i t h m s e l e c t e d f o r t h e i s o l e u c i n e e p i m e r i -z a t i o n m o d e l , f o l lo w i n g t h e a d j u s t m e n t o f d i f fe r e n tm o d e l s , w a s t h e o n e c o m m o n l y p r o p o s e d b y n u m e r -o u s a u t h o r s ( M i t t e r e r, 19 75 ; G o o d f r i e n d a n dM i t t e r e r, 1 9 8 8; G o o d f r i e n d a n d M e y e r, 1 9 9 1) . I nt h i s c a s e t h e b e s t f i t w a s o b t a i n e d t h r o u g h t i m e ( t )a d j u s t m e n t ( c . c . 0 . 9 8 7 2 ) r a t h e r t h a n t i m e s q u a r e -roo t (c . c . 0 .9533) . The mode l i s a s fo l lows :

A / ! t = ( -3 4 .9 9 4 - 25 .8 ) + (267 .14 4 - 20 .32)

x 1n{0.565/[0.565 - ( A /I) / (1 + A/I )]},


A l t h o u g h t h e f i n a l e q u i l i b r i u m s t a t e i s a f f e c t e do n l y b y t e m p e r a t u r e , f o r t h e I l e / A l o e p i m e r i z a t i o nr e a c t i o n i t d e m o n s t r a t e d t h e e x i s t e n c e o f a r e l a t i o n -s h i p i n F o r a m i n i f e r a b e t w e e n r a c e m i z a t i o n p e r c e n -t a g e s, a g e a n d t h e c u r r e n t m e a n a n n u a l t e m p e r a t u r e( C M AT ) ( We h m i l l e r, 1 9 8 4 ) . I n o u r s t u d y, w e h a v ed e t e r m i n e d t h e C M A T f o r th e a r e a s o f P r i e g o( To r r e s e t a l . , 1 9 9 4 ) a n d R e d u e f i a ( L l a m a se t a l . ,1 9 95 ), w h i c h a r e c l i m a t o l o g i c a l l y s i m i l a r t o t h e z o n eu n d e r s t u d y, a C M AT o f 11 - 1 4 C h a v i n g b e e nf o u n d . T h u s , i t h a s b e e n c o n s i d e r e d t h a t t h e s a m em o d e l s w o u l d b e a p p l i c a b l e . A s r e g a r d s t h e t y p e o ff o s s il s u n d e r s t u d y, t h e i n i t i a l re s u l t s p o i n t e d t o t h ee x i s t e n c e o f d i f f e r e n c e s b e t w e e n a m i n o a c i d k i n e t i c sd e p e n d i n g o n t h e g e n e r a o f m o l l u s c a a n a l y se d .T h e s e d i f f e re n c e s w e r e m o r e m a r k e d i n t h e o l d e s ts a m p l e s a n a l y s e d . I n 1 0 s a m p l e s o f a n c i e n t f r e s h -w a t e r g a s t r o p o d a ( P l a n o r b i s s p . a n d R a d i x sp.)f r o m P r i e g o ( To r r e se t a l . , 1994) th i s e ffec t was v ery

m a r k e d f o r g l u t a m i c a c i d b u t n e g l i g i b le f o r l e u c in e .A p o o r c o r r e l a t i o n b e t w e e n t h e r a c e m i z a t i o n r a t i o so f d i f f e r e n t a m i n o a c i d s m i g h t b e e x p l a i n e d i nt e r m s o f e a r l y d i a g e n e s i s ( G o o d f r i e n d , 1 9 91 ), e . g .t h e l o s s o f t h e m o s t e a s il y h y d r o l y z a b l e a m i n o a c i df r o m t h e t e r m i n a l p r o t e i n c h a i n s , a s p a r t i c a c i d ,m i g h t b e r e f l e c t e d i n l o w e r D / L r a t i o s , t h i s a p p a r -e n t l y b e i n g u n c o r r e l a t e d w i t h t h e h i g h e r D / L r a t i o so f t h e l e s s e a s i l y h y d r o l y z a b l e a m i n o a c i d s , s u c h a si so leuc ine .

Te n i n d i v i d u a l s a m p l e s f ro m t h e P r i e g o a r e a w e r ed a t e d a c c o r d i n g t o t h eD/L l e u c i n e r a t i o s , a n d a na v e r a g e v a l u e o f 7 3 3 + 14 0 k y w a s o b t a i n e d . F o rt h e P r i e g o a r e a a c u r r e n t m e a n a n n u a l t e m p e r a t u r e( C M AT ) o f b e t w e e n 11 a n d 1 4 C w a s o b t a i n e d .T h e s e v a l u e s w e r e c o h e r e n t w i t h t h o s e c o r r e s p o n d -i n g t o t h i s g e o g r a p h i c a l l o c a t i o n , a n d t h e C M ATw a s u s e d a c c o r d i n g t o t h e W e h m i l l e r ( 1 99 3 ) c r i t e r ia .F r o m o u r p o i n t o f v ie w, te m p e r a t u r e m i g h t a f f ec to n l y t h e f i n a l A l o / I l e e q u i l i b r i u m s t a g e . A c c o r d i n gt o t h e s e r e s u l ts , o t h e r a l g o r i t h m f a m i l i e s w e r ea d j u s t e d u s i n g o n l yP l a n o r b y s sp . and R a d i x sp .s a m p l e r e s u l t s , t h e a d j u s t e d m o d e l s b e i n g a s f o l -l o w s :

L E U x/ t = (0.94 + 0.63) + (11.81 4- 0.70)

x l n { [1 + ( D / L ) ] / [ 1 - ( D / L ) ] } ,

cor re la t i on coeff i c ien t = 0 .9870 (n = 32)

AS P ~ / t = ( -3 .3 4 4 - 2 .31) + (12.38 4 - 2 .24)

x In{J1 + (D/L ) ] / [1 - (D/L) ]} ,


PH E ~ / t - - (0 .48 4- 2 .88) + (15 .64 4- 3 .71)

x In{[1 + (D/L ) ] / [1 - (D/L ) ]} ,


G L U ~/ t = (0 .33 4- 1 .44) + (21 .93 2 .48)

l n { [1 + ( D / L ) ] / [ I - ( D / L ) ] } ,


I n t h i s c a se t h e b e s t m o d e l f o u n d f o r A l o / I l e e p i-m e r i z a t i o n w a s t h e s q u a r e - r o o t o f t i m e( x / t )a d j u s t e d ( c . c . 0 . 9 4 9 5 ) . Ti m e ( t ) a d j u s t m e n t h a d a

low er cor r e la t ion coff i c ien t va lue (c . c. 0 .9359) .

A / I ~ / t = ( - 0 . 02 4 - 1 .42) + (21.85 4 - 2 .31)

x ln{0 .565 / [0 .565 - (A / I ) / (1 + A/ l ) ]} ,

cor re la t i on coeff i c ien t = 0 .9495 (n = 21)

RESULTS AND DISCUSSION

Ve n t a M i c e n a

A t V e n t a M i c e n a , o n l y 2 1 a n a l y t i c a l r e s u l t s w e r eo b t a i n e d f r o m t h e 2 3 o r i g i n a l s a m p l e s , tw o b e i n gl o s t d u r i n g t h e s a m p l e p r e p a r a t i o n p r o c e s s . T h e


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92 T. Torre set al.

S i m i l a r i t y

- 3 0 1 . 2 7 - -

- 1 6 7 . 5 1 - -

Clt~tcr ol"t~hservatiom(15 ~ - 5 variables)I.h~,gc n~tlxxf WardI)istM:astxe:Euelide,~

- 3 3 . 7 6

1 0 0 . 0 0

0 0 0 0 0 0 ?f G Gf

- "C~ror~."o _ _ = ~ , " ObmrvaUons

G G f G G f G f G f

Fig. 5. Similarity cluster o f amin o ac id racemization ratios (Ile/A lo isoleucine, leucine, aspa rtic acid,glutamic acid) of Venta M icena samples.

r e m a i n i n g s a m p l e s c o u l d b e c l a s s e d i n t o t w og r o u p s : Opercula a n d Gast ropoda . T h e f o r m e r w e r e2 - 3 m m s i z ed , s u b e i r c u l a r - e l l i p s o i d a l s h a p e d d i s h e so f a r a g o n i t e , p r o b a b l y o fB i t h y n i as p . T h r e e d i f f e r -e n t g e n e r a w e r e g r o u p e d i n t oGastropoda.

F i r s t , a c l u s t e r a n a l y s i s w a s p e r f o r m e d w i t h G Ca n a l y s i s r e s u l t s , t h r e e d i f f e r e n t g r o u p s b e i n g i d e n t -i f i e d . O n e o f t h e s e g r o u p s , w h i c h d i f f e r e d w i d e l yf r o m t h e o t h e r t w o , i n c l u d ed f o u r s a m p l e s w i t hv e r y l o w r a c e m i z a t i o n r a t e s , t h i s d i s a g r e e i n g w i t h

3imilarity

-273.34

- 1 4 8 . 8 ~ - -

Clu~,,~rol'ol:servatiom(1 6ohservatiam- 4 variables)LU/~a~ n~thodWardDia.IVleas~e:Euclidean

- 2 4 . 4 5

100.00

G G f G G G f G f C rf C , f C O O

o = " G a s ~ o p ~ " Observations( ) = " O i x ' r c ~ "f = ~ a ig a = r = '

0 0 0 0 ?f

Fig. 6. Similarity cluster o f amin o acid racemization ratios (Ile/A lo isoleucine, leucine, aspa rtic acid,phenilalanine, glutamic acid) of Venta Micena samples.


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AMINOCHRONOLOGYX ~ a I q i ~n a

1800

I I I I I I I I I I

l ie LBJ ~ ~ ~ i le leu ~ p Id~ 91u

A AUppercase: '~ :~ ta LA " {detemmedor Tra~amm:)lowercase : GA.STi~~:X 3A " ~,det~( ,, i, -. x l o r T - r ~ : l

Fig. 7. Box and whisker plot of calculated ages from different amino acids and materials from VentaMicena.

the other two. These results were interpreted asbeing due to the influence of contamination, andwere discarded. A second cluster analysis of the17 remaining samples revealed the existence oftwo groups: Opercula,along with an unidentified

fragmenta, an d Gastropoda (two Radix sp., oneBulimus sp. and six unidentified gastropod frag-menta). In the second group a fragmenta samplepresented very low racemization ratios, with theexception only of Alo/Ile. This result has beeninterpreted as a diagenetic effect and was alsodiscarded prior to performance of the final clus-ter analysis (Fig. 5). All the analyses were car-ried out on the basis of four amino acids (Alo/lie, leucine, aspartic acid and glutamic acid)since some results for phenyla lanine were missing.When the clustering was accomplished usingcomplete results, phenylalanine included, the

grouping obtained was the same (Fig. 6), thusreinforcing the validity of the previous clusteranalysis.

Final age calculation was accomplished accord-ing to the first set of models for seven samplesgrouped into Opercula,and according to the sec-ond set of models for nine samples included in theGastropodagroup (Fig. 7 and Table 3). The uni-

dentified fragmenta was included in the Operculagroup, this giving rise to a slight difference withrespect to previously published data (Torres et al.,1995).

Taking into account the fact that the resultsobtained are independent measurements of thesame parameter over a time interval, it is possibleto calculate a global dating for each group(Table 3); it may be observed that age calculationsfor the fragmenta group show a higher scatteringthan those for the Operculagroup. Prior to comput-ing a "general average dating", the former globalresults from (Table 3) were analysed for each

sample group, Operculaan d Gastropoda.The test ofthe homogeneity of variances for the two groupsshows that both are different, according to the

Table 3. D/L average values and age calculations of Venta Micena samples

A/I LEU ASP PHE GLU GLOBAL

OperculaO/L 1.160 + 0.031 0.893 + 0.017 0.922 + 0.012 0.896 0.013 0.835 0.033Age (Ky) 778 66 1174 149 947 18 909 78 1062 138 976 65

GastropodaD / L 0 8 7 3 0 . 0 5 5 0 . 8 8 1 + 0 . 0 4 3 0 . 8 0 3 0 . 0 2 3 0 . 7 0 6 0 . 0 4 3 0 , 6 1 3 0 . 0 2 5

Age (Ky) 1511 318 1111 4-258 591 77 809 144 1012 107 1009 +_ 130


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94 T. Torreset aL

torres et al. (1997) - amino acid chronology of the lower pleistocene deposits of venta micena...

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