Acople de Gauss-Bonnet (l´ımite k=0)

S eve r a l an a l yt i ca l methods u s e d i n t he eva luat i on o f fat s are we l l e s t ab l i shed . The methods u s e d in the course o f t h i s wo rk are de s c r i bed be l ow .

3 . 1 . EXTRAC T I ON OF MI LKFAT

S ome ana l y s e s requ i re pure mi l k f at . Mi l k fat was e x t racted by me l t ing a but t e r s amp le at 5 0 °C . The fat layer w a s cen t r i fuged and the separated fat pha se f i l t e red t h rough Whatman fi l t e r paper .

3 . 2 . SEC T I L I T Y HARD NE S S

S amp l e s f o r hardne s s me asureme nt s were h e l d for t h i rt y days at 4 °C t o a l l ow fu l l s etting pri o r t o anal y s i s . Sect i l i t y

ha rdne s s a t 1 0 °C wa s de termined u s ing a modi fied F I RA / N I RD e x t rus i on apparat u s according t o the method o f T a y l o r e t a l

( 1 9 7 1 ) .

S amp l e s we re cont a i ned in 4 8 mm d i amet e r s t a i n l e s s s t e e l rings . Longi tudi n a l s l o t s on oppo s i t e s i de s o f the rings a l l owed the c y l inder o f but t e r to be cut by a s t a i n l e s s s t e e l w i re . The wire was mounted on a bow ca r r i e d by t he l ongi t u d i n a l rod cont act ing the me a su r ing spr ing o f the i n s t rument . A cut t i ng speed o f 4 0 mm /min was used . S amp l e s p l u s s amp l e h o l d e r s we re he l d in a wat e r bat h at 1 0 °C f o r a l e a s t f i ve h o u r s be fore me a surement . Dup l i c ate s amp l e s we re me a s u red .

The i n s t rument i s c a l ibrated i n t e rms o f load; hence s e ct i l i ty ha r dne s s i s quoted i n units o f grams . The s e un i t s are

arb i t r a r y a s they d epend on the w i re d imens i on s , cut t ing speed and s amp l e r i n g dimens i on s .

3 . 3 . FAT T Y AC I D COMPO S I T I ON

F a t t y a c i d c ompo s i t i on wa s de t e rmined by gas chroma t og r aphy o f fat t y a c i d met h y l e s t e r s prepa red by di rect t ra n s

e s t e r i f i c a t i o n o f a m i l k fat s amp l e w i t h s o d i um meth o x i de / methan o l .

A Hew let t - P ackard 5 8 4 0A gas chromat ograph w i t h a s t a i n l e s s s t e e l c o l umn 2 m l ong , 0 . 1 2 5 i n ch di amet er packed w i t h 1 0 - 1 5% d i et h y l e ne g l y c o l succinate (An a l ab , In c . ) on Chromo s o s rb W-HP

8 0 / 1 0 0 (Va r i an Ae rograph ) wa s u s e d to an a l y s e t he fat t y acid met h y l e s t e r s . N i t rogen , flowing at a rate of 15 m l / mi n , was u s e d as a c a r r i e r gas .

A st anda rd anhydrous m i l k fat wa s u s e d t o c a l i b r a t e t he gas chromat ograph . The r e s u l t s obt a i ned shou ld agree w i t h

e s t ab l i shed m e a n v a l u e s ( T ab l e 3 . 1 ) within t w o s t andard dev i at i o n s . W e i ght correct ion fact o r s ( we i ght / a r e a of peak ) a re g i ven i n Tab l e 3 . 2 .

S amp l e s were prepared by me lt i n g t he mi l k fat a t 6 0 °C . When me l t e d , e i ght drops were t r ans ferred to a s c rew t op t ube and 2 m l o f the e s t e r i fying re agent ( s odium methox i de / me t hano l in a m i xt u r e o f e t hy l ether and pet r o l eum ethe r ) w a s a dded . S odium methox i de / methan o l wa s prepa red b y t he method de s cr ibed by S hehat a et a l , 1 9 7 0 . Aft e r 2 - 4 mi nut e s 5 ml o f h e xane was a dde d . A furt he r 2 minutes l at e r 5 ml of saturated s o di um c h l o r i de s o l ut i on was added . The t ube wa s i nve r t e d s eve r a l t i me s t o m i x the cont ent s t h e n cent r i fuged b r i e f l y . Four m l s o f the upp e r hexane l ayer were w i t hdrawn a n d t ra n s ferred t o a v i a l cont a i n i n g 5 0 0 mg o f anhydrous s odium s u l p h at e . The vial wa s shaken and a l l owed t o s e t t l e . 1 - 2 � 1 o f s amp l e was

i n t ro duced to t h e gas chroma t og r aph . The s o l ut i on of e s t e r s is s t ab l e f o r a few days i f kept r e f r i ge rated .

The i n j e ct o r tempe rature and det e c t o r base were ma i nt a i ned at 2 4 0 °C and 2 5 0 °C re spect i ve l y . A ft e r i n j e ct i on o f t he s amp le

T able 3 . 1 : Comp o s i t i o n o f st andard anhydrous m i l k fat .

FATTY AC I D MEAN STD . DEV .

( we ight % ) 4 : 0 3 . 5 8 0 . 3 6 : 0 2 . 0 7 0 . 1 5 8 : 0 1 . 2 2 0 . 1 1 0 : 0 2 . 4 8 0 . 1 1 2 : 0 3 . 2 4 0 . 1 1 4 : 0 1 0 . 0 0 . 3 1 6 : 0 2 5 . 0 0 . 3 1 8 : 0 1 0 . 4 0 . 4 1 8 : 1 2 7 . 5 0 . 7

T able 3 . 2 : Norma l i z i ng fact o r s for fat t y acid we i ght pe rcent age s , re l at i ve t o c 1 6 : 0 · FAT TY AC I D 1 6 : 0 4 : 0 6 : 0 8 : 0