i r «mm of mm m mm mmB u d a p e s t , S e p te m b e r 4 -11 , 1963

F ree M ethio n in e , O ccurence and S ig n i f ic a n c e i n Anim al T is s u e s ,

O lle DahlS c a n 's C e n t r a l la b o r a to r iu m , Malmö, Sweden.

S ? s

2 6

An a p p re c ia b le .a m o u n t o f th e m e th io n in e . fo u n d in m uscle i s p r e s e n t in a f r e e s t a t e , v i z . a b o u t 10 $ o f th e t o t a l , 1 »2 ) The re s id u e i s bound

as a p r o t e in c o n s t i t u e n t . Under a s e p t i c c o n d i t io n s , th e c o n te n t o f f r e e

h io n in e in m uscle i s c o n s ta n t f o r a t l e a s t th r e e weeks a t te m p e ra tu re s to s t above z e r o , 1 ’ 2 )

As e v id en c e d by s e v e r a l i n v e s t i g a t i o n s , th e r ô le p la y e d by m e th io n in e

iQ 'b io lo g ic a l sy stem s i s a s a m e th y la t in g a g e n t and aa a m e ta b o lic p r e -

°ur s o r o f c y s te in e , To be a b le to a c t in t r a n s m e th y la t io n r e a c t i o n s ,

however, m e th io n in e must be c o n v e r te d to S -a d e n o s y lm e th io n in e , a p h o sp h a te -

ee su lphonium com pound,5 5^ The c o n v e rs io n i s p e rfo rm ed by means o f an

A c tiv a t in g enzyme, a d en o s in e t r i p h o s p h a te , Mg++ , and red u ced g lu t a th io n e .

^he t r a n s m é th y la t io n s a re c a ta ly z e d by s p e c i f i c m e th y lp h e ra s e s . The

P roduct o f t r a n s m e th y la t io n i s u s u a l l y th e N -m ethyl d e r iv a t iv e o f th e

a c c e p to r w h ile S -a d e n o sy lm e th io n in e i s c o n v e r te d to S -ad en o sy lh o m o p y a te in « .Example s o f su ch t r a n s m é th y la t io n s a r e g l y c i n e ^ s a r c o s i n e and b e t a i n e .Sh+ • 7a n o la m in e * ^ c h o l in e , c a r n o s in e — * a n s e r in e , dopa am ine a d r e n a l in e ,Sbycocyam ine—* c r e a t i n e .

The f u n c t io n a l s i g n i f i c a n c e o f m e th io n in e e x c i t e s a t t e n t i o n to th e

° ccu ren ce o f f r e e m e th io n in e in v a r io u s t i s s u e s . The p re s e n c e o f f r e e

a e th io n in e m igh t i n d ic a te t r a n s m e th y la t io n a c t i v i t y a n d /o r th e c o n v e rs io n M eth io n in e in to c y s te in e .

^££a rim e n ta l

F ree m e th io n in e was d e te rm in e d in a w a te ry e x t r a c t o f v a r io u s a n im a l

SBUes a f t e r p r e c i p i t a t i n g p ro te in o u s m a te r i a l w ith p h o s p h o tu n g s tic a c id

^ th e n a p p ly in g th e m ethod o f M cCarthy and S u l l iv a n 5 ) a s m o d if ie d by 0rn-> Jo n e s and B lum ^),

x The g-x t r a c t wa s p re p a re d by h e a t in g 50 g f i n e l y g round m a te r ia l +

w a te r f o r 20 m in u te s in a b o i l i n g w a te r b a th u s in g r e f l u x . A f te r

c° ° l in g to room te m p e ra tu re th e m ix tu re .w a s a g i t a t e d in a m achine f o r 30

te 3 and th e n c e n t r i f u g e d . Between 0 ,5 and 4 g p h o s p h o tu n g s tic a c id

^ added to 100 ml e x t r a c t , th e amount o f p h o s p h o tu n g s tic a c id b e in g

P enden t on th e m a te r i a l ; e x t r a c t s o f i n t e s t i n e s , stom achs and c e r t a i n

^ g a n s r e q u i r e a l a r g e r a d d i t io n th a n th o s e o f s k e l e t a l m u sc le s . A f te r

av in g b een k e p t in a r e f r i g e r a t o r o v e r n ig h t th e e x t r a c t was f i l t e r e d

2

to glVe a c l e a r s o lu t io n . Ground p ig s k in (T ab le I , o an p le Ho* 76 ) was

E r e c t e d w ith o u t p re c e d in g h e a t t r e a tm e n t .

B r i e f ly , th e d e te r m in a t io n v/as c a r r i e d o u t by f i r s t ad d in g 1 ml

^ NaOH to 4,5 ml e x t r a c t and th e n by a d d in g 0 ,5 ml o f a 2 $ N a - n i t r o -

P ru ss id e . A f te r e x a c t ly 10 m in u te s a t room te m p e ra tu re 2 ml o f a 3 fo

° lu t io n o f g ly c in e was added and , a f t e r a n o th e r 10 m in u te s p e r io d , 2 ml

c°n c e n tra te d p h o sp h o ric a c id . Ten m in u te s t h e r e a f t e r th e e x t in c t io n was

111 a Beckman s p e c tro p h o to m e te r model B a t . 510 m y /, .A b la n k t e s t and

Sei“i e s o f s ta n d a rd s o lu t io n s o f m e th io n in e ( 0.25 to 1 ,0 0 ml o f a s o lu t io n

100 m e th io n in e in 95 ml w a te r + 5 ml 6N HC1 were u se d ) w ere m easured f 0

•C a lib ra tio n . Two a n a ly s e s w ere p e rfo rm ed on s e p a r a te e x t r a c t s . A d d itio n

known am ounts o f m e th io n in e to v a r io u s e x t r a c t s y ie ld e d com ple te r e ­g i o n .

Bor c a l c u l a t i o n th e c o n te n t o f w a te r and p r o te in o f th e m a te r ia l

S ^e ^enm ined. The c o n te n t o f m e th io n in e was com puted on a c ru d e p r o te in

'sas> assum ing t h a t th e f r e e m e th io n in e a f t e r a g i t a t i n g was e v e n ly

s ln ib u te d in th e added w a te r and t h a t c o n ta in e d in th e t i s s u e ,

The m ethod u sed i s s im i l a r to th e m o d if ic a t io n by Csonka and D e n to n ^

k n has b een found to g iv e r e s u l t s in good ag reem en t w ith th o s e o b ta in e d

^ ^ke m ic r o b io lo g ic a l m e th o d ,8 ^

E^ • ^ L gL and d is c u s s io n

B able I shows th e r e s u l t s o b ta in e d . The d e te r m in a t io n s w ere p erfo rm edon

Ihe

iht,

the v a r io u s t i s s u e s 1 to 4 days a f t e r s l a u g h t e r o f th e a n im a ls . A lthough

c o n te n t o f f r e e m e th io n in e i s s u r p r i s i n g l y c o n s ta n t in th e r e f r i g e r a t e d

a c t m uscle d u r in g a lo n g p e r i od p o s t mortem - a s was a l r e a d y rem arked

"khe in t r o d u c t io n - i t was o b se rv ed t h a t th e c o n te n t g e n e r a l ly in c re a s e d

Sld e ra b ly when g round sam ples w ere s to r e d r e f r i g e r a t e d o r f r o z e n . T h isnas^ s p e c i a l l y t ru e f o r th e o rg a n s . In c re a s e d enzyme a c t i v i t y due to c e l l

Srh p tio n i s , how ever, a w ell-know n phenomenon.

Table I

The h ig h c o n te n t o f f r e e m e th io n in e in s k e l e t a l m u sc les - on an ^ age 200 mg-fo o f th e c ru d e p r o t e in - i s v e ry s t r i k i n g . Assum ing a t o t a l

2 , 5 fo m e th io n in e in th e m uscle p r o t e in (s e e r e f , ^ ) , 6 to 11 /a (a v e ra g eV3' ° f the m e th io n in e i s p r e s e n t in a f r e e s t a t e . Of c o u rs e , t h i s f a c t i s

hoam biguous p ro o f o f an o u ts ta n d in g f u n c t io n a l s ig n i f i c a n c e o f f r e e

- 3 -

aeth io n in e in th e s k e l e t a l m u sc le s , even l e s s c o u ld c o n c lu s io n s be draw n

a t°u t which f u n c t io n s i t e x e r t s . B ut th e f a c t t h a t th e c o n te n t o f f r e e

aeth io n in e in s k e l e t a l m uscle i s e s s e n t i a l l y h ig h e r th a n in o rg an s and

^ h e r t i s s u e s i n v e s t i g a t e d makes an i n t e r e s t i n g s t a r t i n g p o in t f o r f u r t h e r 8 tud ies on t h i s p a r t i c u l a r s u b je c t .

Sam ples No. 4 , 8 , 9 , 1 3 , and 14 w ere ta k e n from d i f f e r e n t p a r t s o fthe cj

ame c a r c a s s . E v id e n t ly th e c o n te n t o f f r e e m e th io n in e i n v a r io u s b e e f

slcel e t a l m u scles i s f a i r l y c o n s ta n t . The ran g e was 181-202 mg-$ f r e e

^ th io n in e i n the c ru d e p r o t e in and t h i s c o n te n t i s o b v io u s ly in d e p e n d e n t

the c o n te n t o f c o n n e c tiv e and f a t t y t i s s u e ( s e e b e lo w ). These c o n c lu s io n s

6e® to be t r u e a l s o f o r h o rs e m u sc le s . Thus, th e sam ples No. 29 , 3 1 , and

* ?,rith c o n te n ts o f f r e e m e th io n in e ra n g in g be tw een 200 and 225 mg-$ of

Crude p r o t e i n , were ta k e n .f ro m one h o rse c a r c a s s and th e sam ples No. 30, > and 34 Wi t h c o n te n ts o f 152-183 mg-$ w ere ta k e n from a n o th e r . I n

addi t i o n , th e s e r e s u l t s i n d i c a t e t h a t th e r e .m ig h t be i n d iv id u a l d i f f e r e n c e s d^h re g a rd to th e l e v e l o f f r e e m e th io n in e .

As a l r e a d y p o in te d o u t , th e c o n te n t o f f r e e m e th io n in e in s k e l e t a l_ «1

th seems n o t to be r e l a t e d to th e c o n te n t o f c o n n e c tiv e t i s s u e o f

m u sc le s , T h is i s a l s o e v id e n t from F ig . 1 in w hich th e c o n te n t o f f r e e

lo n in e i s p l o t t e d a g a in s t th e c o n te n t o f h y d ro x y p ro lin e , th e l a t t e ra r\ 1 i ^

a s an in d ex o f th e p re s e n c e o f c o n n e c tiv e t i s s u e . * ' One p e r c e n t

0xy p r o l in e c o rre sp o n d s to a b o u t 7lj 5® c o n n e c tiv e t i s s u e p r o t e in .

F ig . 1

I t h as n o t b een d e m o n s tra te d w h e th e r c o n n e c tiv e t i s s u e c o n ta in s

S o c i a b l e am ounts o f f r e e m e th io n in e o r n o t ( s k in h a s a low c o n te n t ,

^ ^ • 7 6 ) . I f i t s c o n te n t o f f r e e m e th io n in e i s low , an e x p la n a t io nOf* +.T-e h ig h l e v e l s o f f r e e m e th io n in e i n s k e l e t a l m u sc les r i c h in

ct i v c t i s s u e c o u ld be t h a t th e s e m u sc le s have a la r s r e r workin/? °apacl+ s

ATy an d , c o n se q u e n tly , a l s o a g r e a t e r m e ta b o lic a c t i v i t y . T h is c o u ld p° ss ib ithe 17 t e PUt l n c o n n e c tio n w ith a h i &h er c o n te n t o f f r e e m e th io n in e in

P ro p e r s t r i a t e d t i ssu e o f su ch m u sc le s .

fof*

U

'hhd inAmong th e o rg a n s p a r t i c u l a r l y low c o n te n ts o f f r e e m e th io n in e were

u d d e r , rum en, lu n g s , h e a r t , k id n e y , and s k in ) b lo o d and i t s

t io n s (sa m p le s No, 6.1 —63 and 77-79) w ere a l s o found to have v e ry low

s o f f r ee m e th io n in e . S u r p r i s in g ly enough s t r i a t e d m u sc le s from th eaead (

sam ples No, 16 and 1 7 ) showed e x tre m e ly low v a lu e s o f _ f r e e m e th io n in e ,^ S° th ~0 s t r i a t e d m uscle o f d iaph ragm (sam p les No. 15 and 2 7 ) was fo u n d .

°nt a i n s i g n i f i c a n t l y l e s s f r e e m e th io n in e th a n th e s k e l e t a l m u sc le s .

- 4 - 3

A • 121W ording to e a r l i e r i n v e s t i g a t i o n s ' head m eat has an e s s e n t i a l l y lo w er

c° r te n t o f c r e a t in e th a n s k e l e t a l m u sc le s . T h is f a c t i n d i c a t e s some

rela t io n s h ip betw een th e c o n te n t o f f r e e m e th io n in e . and c r e a t in e and , w ith

refe ren ce to w hat was m en tioned in th e in t r o d u c t io n , may r e f l e c t th e s i g n i -f i canoe o f m e th io n in e a s a m e th y la t in g a g e n t in th e fo rm a tio n o f c r e a t in e

f r °m g ly co cy am in e . In o rd e r to g e t a more c l e a r c u t c o n c e p tio n o f th e

E l a t i o n betw een f r e e m e th io n in e and c r e a t i n e , th e c o n te n t o f th e s e

c° n s t i tu e n ts was a n a ly z e d f o r a number o f sam ples o f s k e l e t a l m u sc le s ,

r ia te d m uscle from head and d iaphragm a s w e ll a s o f o rg a n s . The r e s u l t s

a r° P re se n te d in P ig . 2.

S ix s e p a r a te g roups can be d i s t in g u i s h e d . Group I h a s a h ig h c o n te n t

"both f r e e m e th io n in e and c r e a t i n e ; i t in c lu d e s s k e l e t a l m u sc le s . Groups

and m have a m odera te c o n te n t o f f r e e m e th io n in e and a m o dera te to

c o n te n t o f c r e a t i n e ; t h i s group i s r e p r e s e n te d by s t r i a t e d m uscle from aPhragm and to n g u e , Group IV (sam p les No. 24 and 28) h as a low c o n te n t

f^ee m e th io n in e and a m o d era te to h ig h c o n te n t o f c r e a t i n e ; i t i s r e -

f i e son ted by th e c a r d ia c m u sc le . Group V i s c h a r a c te r iz e d by a low c o n te n tOf f

re e m e th io n in e and a m o d era te c o n te n t o f c r e a t i n e ; t h i s g roup c o n s i s t s

s t r i a t e d m u sc les from th e h ead , F i n a l l y , Group VI h as a low to m o dera te contn *

Gnt o f f r o e m e th io n in e and a low c o n te n t o f c r e a t i n e ; t h i s group in -

fudes re d o rg ans (e x c e p t h e a r t and to n g u e ) , smooth ( u n s t r i a t e d ) t i s s u e s ,I olciri> and b lo o d .

freeHo’

The c ru d e p r o te in o f th e mucosa from b e e f bungs i s much r i c h e r in .

m e th io n in e th a n t h a t o f th e bung t i s s u e (T ab le I , sam ples No, 5 7 -6 0 ) ,

we v e r , th e m ucosa p r o t e in was found to make o n ly 22 to 26 $ o f th e sum

^ 8 mucosa p r o te in + bung t i s s u e p r o t e in . Thus, i t can be c a l c u la te d t h a t

° ru d e p r o t e in o f th e bung w ith m ucosa c o n ta in e d betw een 80 and 90 mg-$

iGe m e th io n in e .

A lso th e c ru d e p r o t e in o f th e m ucosa from p ig s tom achs h as a h ig h e r

^ fo n t o f f r e e m e th io n in e th a n t h a t o f th e u n s t r i a t e d m uscle t i s s u e to

kieh i t i s a t ta c h e d (T ab le I , sam ples No, 6 8 -7 1 ) . About 7 i° o f th e sum ttlUCn O

ba p r o te in + stom ach t i s s u e p r o te in was mucosa p r o t e i n . On th e o th e r th e p reG moth io n in e c o n ta in e d in th e c ru d e p r o t e in o f th e mucosa

0la o h i t t e r l i n g was found to be b u t a l i t t l e h ig h e r th a n t h a t in th e

e P r o te in o f th e c h i t t e r l i n g t i s s u e (T ab le I , sam ples No, 7 2 -7 5 ) .

- 5 -

From those results it seems evident that, among other things, the aucosa of beef bung and pig's stomach are sites for intense metabolic turnover. Whether the high values of free methionine are merely a sign of Proteolytic activity or of transmethylation, or both, must be disclosed V separate studies.

Earlier analyses of characteristic constituents of various animal tissues have revealed that the content of tryptophan and particularly of kydroxyproline in the tissue protein can be used as an index of high and

quality animal protein, respectively. ’ 1 Blood and its fractionsare exceptions; although their proteins are high in tryptophan, they are ^iicient in the essential amino acids isoleucine and methionine. To distinguish striated muscular tissue from organs and smooth muscles the c°utent of creatine, calculated on a crude protein basis, is very useful, ■■ho present investigations have disclosed that the content of free Elethionine in the crude protein is also a feature of skeletal muscle. However, as far as a suitable index of the quality of meat products is concerned, the determination of the content of free methionine in the Crude protein offers no advantage over establishing the content of Hydroxyproline and/or creatine. In addition, nothing is hitherto known atout changes of the content of free methionine which may occur during ’the processing of meat products and this may cause erroneous conclusions,

SummaryBetween 6 and 11 c/° o f th e t o t a l m e th io n in e p r e s e n t in s k e l e t a l

Muscle was fo u n d to be f r e e m e th io n in e , The c o n te n t o f f r e e m e th io n in e

in o th o r an im al t i s s u e s a s w e ll a s in b lo o d and i t s f r a c t i o n s i s low ,

d h te s t in a l mucosa i s r i c h in f r e e m e th io n in e . The f in d in g s a r e d is c u s s e d

ih view o f th e f u n c t io n a l s i g n i f i c a n c e o f m e th io n in e . The d e te rm in a tio n

°£ th e c o n te n t of f r e e m e th io n in e o f f e r s no ad v an tag e o v e r th e a n a ly s i s

°£ th e c o n te n t o f h y d ro x y p ro lin e a n d /o r c r e a t in e f o r th e pu rp o se o f

S e t t in g an in d e x o f th e q u a l i t y o f moat p ro d u c ts .

RésuméDe 6 h 11 c!° de l a m é th io n in e t o t a l e du m uscle s q u e le t t iq u e se tro u v e

8ous form e de m é th io n in e l i b r e , La te n e u r en m é th io n in e l i b r e des a u t r e s

d is su s de même que c e l l e du san g e t de ses f r a c t i o n s e s t f a i b l e , La A q u e u se i n t e s t i n a l e e s t r ic h e en m é th io n in e l i b r e . Les r é s u l t a t s s o n t

- 6 -

d is c u té s q u an t à l a s i g n i f i c a t i o n f o n c t io n n e l l e de l a m é th io n in e . La dé­

te rm in a tio n de l a te n e u r en m é th io n in e l i b r e n 'o f f r e p as d 'a v a n ta g e , p a r

ra P p o rt à l a d é te r m in a t io n de l a te n e u r en h y d ro x y p ro lin e o u /e t en c r< éa tin e ,

Pour é t a b l i r un in d e x de l a q u a l i t é des p r o d u i t s c a r n é s .

Zusam m enfassung

Zw ischen 6 und 11 cfo vom G esam tm eth ion in im S k e le ttm u s k e l wurden a l s

i:re ie s M e th io n in g e fu n d en . D er G e h a lt an f r e ie m M eth io n in i n a n d e ren

t i e r i s c h e n Geweben sowie im B lu t und in B lu t f r a k t io n e n i s t n i e d r i g . Die

S chleim haut d e r E ingew eide i s t r e i c h an f re ie m M eth io n in . D ie B efunde w er-

H in b l ic k a u f d ie f u n k t i o n e l l e B edeu tung des M eth io n in s e r ö r t e r t .

t>ie E rm it t lu n g des G e h a lts an f r e ie m M eth io n in b i e t e t k e in e n V o r te i l ü b e r

Bestimmung des G e h a lts an H y d ro x y p ro lin u n d /o d e r K re a t in , um e in Q u a lltä tsm a ss f ü r F le is c h w a re n zu e r h a l t e n .

R e fe re n c e s ;

1.

2.

3.4.

5.6.

? ,

8.

9.

10,

U .

1 2 .

0» D ah l, Svensk Kem, T id s k r . 75 _ ( l9 6 l) , 614.

0 . D ah l, D ie Nahrung 6 ( 1962) , 492,

G«L, C a n to n i, J , B io l , Chem, 204 (1 9 5 3 ) , 403.

G .L. C a n to n i, "Onium compounds and t h e i r b io lo g ic a l s ig n i f ic a n c e " in C om parative B io c h e m is try , V ol, I , i 960, p , 181} E d ite d by M. F lo r k in and H .S , Mason, Academic P r e s s , New York and London.

Î .E . M cCarthy and M.X. S u l l iv a n , J , B io l , Chem. 141 ( l 9 4 l ) . 871.

M .J. H orn, D.B, J o n e s , and A .E, Blum, J , B io l , Chem, 166 ( 1946) , 313.

F »A, Csonka and C.A, D enton , J , B io l , Chem, 163 ( 1946) , 329 .

B*A, Greenwood, ■ H.R.- K r a y b i l l , and B .S , S c h w e ig e r t, J . B io l , Chem«121 (1 9 5 1 ) , 25 .

h . J . B lock and K.W, W eiss, Amino A cid H andbook. 'C h a r l e s C, Thomas, P u b l i s h e r , S p r in g f i e ld , 1 1 1 ,, U .S .A ,, 1956,

0 , D ah l, A cta Chem, S cand, 14 ( I 960) , 227.

D ah l, " E s tim a tin g p r o t e i n q u a l i t y o f m eat p ro d u c ts from th e c o n te n to f t y p i c a l amino a c id s and c r e a t in e " in P ro c e e d in g s o f th e 1 s t I n t e r n a t i o n a l C ongress o f Food S c ie n ce & T ech n o lo g y . London ( E n g lan d ) , I 8 th - 2 1 s t S ep tem ber, 1962} in p r e s s .

D ah l, J , A g ri , Food Chem, 11 ( 1963)} in p r e s s ,

ir\M 3

O«•H3O•H 3

h:

M«:

¡r'H 'O'

+»! O ' U

\a«j

o'

"ö•h ; «I

e;

a!

5¡®lSI

ti1

So«!°lti!O4»cj3ÜMC3Oo!q•hlis;Q)gO*+>qa§ü

Cr-c-.ttf-T-tf'íO lf\ e-— 3 ITS CD VO CD CM LTV 3 •fOYC—rr\0

CM U"V CD -vf- CD -CD O IAO KMAINCjT\ CT\ Ñ*\ CT\ MO Lpv ON 0\ CD C7\ O O CD O N*\ CM 3 tÇV C-—

fc\ cr\ VO CD CD C- "sf 3VO CM CO O CO CM CD\

CM H OJ i—1 i—1 H rl i—1 i—It—l<MC\Ji—[C\ii 1 CVirH

CMCM3333CM3CM CM 3 CM 3 CM 3

o3CdCd■rt

Es

©3%

&o¡2»

L «

©3Cd : ca •rt PS

P : Ptí«Ht

J 3

:

tí©

>

■rt P O

:

©Oo©

I—I

3•rt

M•rt

>O3raratíorttío©rrtO3ÈSSort«Hraora 3a•rtrara•rt .

.t*C tíc

fp

o

©

rSI 05s

itó

pt

í :

«H©rrt3•rt

Cd•

rt

>O3p33©3ram•rt

P©¡>•rt

PO©gOo3p•rt

§m

r

ca •H

•P•P-P

r

dcp1

=3

3mm•rt

p©t>•rt

PO©gOo3p

tí tirt

tí 3

©

S

33

d•rt

©•3

,3

P

©3

aP

o

rt

<H 3

Ora

©

©

r—I rrtO

O

ra ra

3

3

3

3

•3

O

P

tí s

•

rt

rt P

ra

•rtm

rt0

3ra1•rtra m•Ö

js

f-

o

3 o

s

©

m

CM

Ptí =«H

©©

P3

3ca

=z

•H

~ca

pra

•Hd

=

z•H

*\p

3>

s©

o =

p

=Z

a

=r

3p

od

caP

©3

3

-3

P3

caP

3

rr

ca3

r

—d

©•H

fc3

3P

ora

o3

ca r

©>

ca•H

>•H

ca r

rp

•Hp

•HP

O

rp

oo

©i>

©p

o•rt

2>■

p§

o•H

o

=o

Op

©o

o

sr:

©O

23

s1—

1d

Op

32

O•H

•Ho

Esca

rz

o3

•HP

r

! o>

3•H

ra3

p

=z:

Esca

rao

=

Z•H

dra

3Es

CMo

•rrtCQ

rtP

»vca

dra

do

Oca

o

=>

d

nrt

©« *H

og

dp

prt

5)

oo

; OCo

d3

©o

rtO

is3

32

©P3

o

s©3P3Ort©sm©rrtOra3rrtrtp©©3ra

o

3o

o

tí•

rt

>3

3i—l —do

tc\ •st- tr\ VO

C— CD

O

V O

3

CM

N"\

© LTV v

û C—

CD

OY

3

31

33

33

33

33

©

Esa

3O

•He

zz

z3

a

3ca

zZ

rt•rl

OP

rt3

CQ rt

o

v i

f

3P

caO

om

ca3

, o

©©

3O

zz

z3 3

Pa

cao

O•H

3•V

caCO

caÖ

; m

33

raz

z©

aa

•HP

3h£

tíO 3

P3

Pm

tí

oo

©z

zz

3

Pp

PP

a ©

ZZ

n

n¡I

d©

•H*•

a

©rt

SZ

zCQ

rt Xp

3

©ca

©ca{£

rtz

zn

zz

oAn

3

O P

C\JCTV •'*’ CPvVO

e-

CDOY o

PCM ny

CM CM CMGM CM CMCM C\JCM CMrA rA

ro

3ts

©3ra : ra•rtPi>>

P

P

: tí

333 :tí

-3©g•rtPdo©

hhI *phhh 'hh

iina

<b3CQCQ'N&hO "■

|á

Of

flON

c— o

CM VO

.

OV

^-M

-O

O V

f"tf- O

O

CO M

3 oN

N VO

C

r- ON

Q

1

oC

" ^ N

NO

O

NV

O

H

HC—

NN

VOLfN

NN

^CM

rHCM

IfV

M-V

jD C

AH

N

N1--1

■ CM

rH

1 c3

~*1cao

zz

a-p1

ci*

oPCn

-Pti60

}Ö

r

•»C¡

/--N

pcrt

c3ca

pg

g

a> .

oO

s

m

-p

,o

= V

ti

i>>ö

Vt

rHo

ofr

o<0

ö'—

/ti

fH3

-P

M0)

0)Ö

î>*CQ

fHf!

CQ r

j T

ÍPC

5)

UG)

o r

ti +>

?H O

G)t>.0

O0

63

O

O

O-P

c

c

=!>

r

7-&

G)

0)=

Cl

ö

>d =

r

Ö=

O

r

O

O

î>>O

<D

P<tí

Cu ^

>

r «H

C¡ "O

Cici

H rH

M

!-P

^

CQo o

•H•rl

rH Pl

Hpc

G

pc

pc<0

-p P3i—

1 CQ

M

<HV

t<u

- =

r- —

Tl Tl

63

0)•H

Z.

Za>

mfr

pq

Vf>V

O

C— CD

ON

orH

CM

tC\

ur\K

O C~~ <P

ON

OH

.CM

C*N N

T LfN VO

C—

GO ON

O

H

CMK

N^

LfN LfN

LfN LfN

LfN tfN

LfN LfN

LfN LfN

VO VO

VO VO

•

C o n ten t o f f r e e n e th io n in e and t o t a l h y d roxypro li.no

in th e c ru d e p r o t e i n .o f s k e l e t a l m uscle from b e e f ,

c a l f , p ig , and h o r s e , The f i g u r e s r e f e r to sam ple

num bers g iv e n in T ab le I .

CMS

htuChoUaofidto•H«HQ>a•Haddd*H(1)POfHft0)ftoddoft4)ftdH3Oftdo0)d•HftdofHOXJ§<D Öft d o■Hd

-p g a

M

daa■HEh

aaH&

E30)<DU«H

to5

ft6

O

0) d a

a

: •H -P!>> -p -P :d«Hx( d :da>•Hft

ftd<HOI—I

ftaft>Odp>d¿2

a

d na

ft-P-P-Pa

d

d

ft aa

xi

•H d

a

d a

a :

•H ft-P -P : d

ftxl d

:da>*H

•H ft

ch ft o

o

d a,-p _pdo

o

O -p

lafH0

:xJ: a1Í2Vidoftaaft

o E

P

dP

OE

aCD

0a

d

pï>

>

do

•Hft

oo

pp

o

O r

oa

aa

a

rHd

£h rHd

d5

s•H r

cO

ft ECQ

oO

CQ•rH

>rd

d

f>-P r

pO

s•H

•HO

Ed

&*

0

•»«K

mCQ

CQCQ

c£c3

O

r—

O r

rfH

fHCÖ

c3o

o0

arH

rHo

Er

rO

E E

dd

&is

aa

o E

“ “

*rlO

E E

fHCQ

fHft

fHft

oCQ

TjCQ

aa

ftCQ

1—1

o r

= s

dO

E E

CQa

CQd

•Hd

aCQ

aCQ

dd

pd

1 P>a

sE

E O

a

e e

rHrH

rHa

ad

•X

CQs

CQ

ft•»

~ r

~ —

rHe

toC3

•H r

ftdftiaaoaddodaa•Hpaî>ft-POaddood-pft&

•HI

s5

Oft

a;

ira|•2toda

•> a

U

: oW

-PdftarHd•Ha•HOd

a

d

a

02 : •H -P

-Pd¿2 ft -P :

a

d

d ft

aa xf

•H d

: -P

dat*•HftOa

= :

oordft•H&aadofHdoai—iodisaO :

d

ftaai—iO :adaPd

P a

:i—iaXoo

daoodp•H|Sad•HPaapd•Hd

a

atofHd

adaa•HPai>•HPÜaoo

rdp to''-'•h d X)£ ft d •r ft ft

rd fH fHo

a

a c3 P

xf

to a p d od

O

-H ft od

P

rd rM ft

ft a o a ¿o

1

-P•H

-

s

r: s

<3fH«V

d

E

End

&d

s

r

r

=

do

•Hft0

r

aÄ

=

r

r

=

<"H _

_

»—H

—

—-P

PaÖo

=

=

r

=

O

E

EP

fHftCQ

a0

i—i

rHO

E

E

o

-

—

-

—00

0d

3a

a

fta0

p-P£T>

M

Mci

n

n •H

•HU

P-P

-PCQ

m•s

♦> Q

)»o

r, *

\ Û

)ft

•>

oo ft

»

a

a

to

fno

e

e

to

fn

a

-n

oa

ft

o

fp

ft

w

po ft

w

rfN-d-LTW

d c~- CD o>

o ft

P^N^N^fOvfOvfOvirsi-'Si-

aP

p

d

d0

d

d

!>>fn

to a0

pd

fn

a

ad

d

>

rHfH

to a a

d

a

o ft

P<d

d

i> ft ftd

p

ftra

aO

ft ftft

dp

ft a

•s4H

•V•*

•• f.

0 =

r2:

to0

•HE

E E

EPQ

Ph

aft Olí§

*CO

I

cm tA-d’ift'd o- cp ctno ft cm if\oft\o t— go o\o tft-CM n>ftftftftftftftftftftCM<NCMCM

■d-lftVptftCpONOftCMC

MO

JCM

CM

CvJC

Mfftfftfft

3 s v

2

co#\

1*

i____(.

24

/ * , / / 18 16

9 / 3 / • •

/ Z i t t

2 * 1 1 • •

I ,

_____ 14 /

/////

!

---7--- 1--/ /

/IV // // /

¡22/25'S'¡29 ! h i

f i T >s j t ^ '

n 1

/ I J # 415 1 2 * * IL

5 % /> X

, . 1 9 .

\ 23' i i

, - " " 3 8 • v~ ------------ ^

- - ' " • 3 5 ►34 32

33 , f i¿1 ® 3 ^ *1 36 4

V I 39

. * 2 0 31 4

b * 5 ^

— J----------------------

11

7 /

m g-7» FREE METHIONINE

F iy . %