Flavor Chemistry - American Chemical Society

Chapter 3

Downloaded by UCSF LIB CKM RSCS MGMT on November 28, 2014 | http://pubs.acs.org Publication Date: February 21, 1989 | doi: 10.1021/bk-1989-0388.ch003

Aroma Development

in Ripening Fruits

P. Dirinck, H. De Pooter, and N. Schamp Laboratory for Organic Chemistry, Faculty of Agricultural Sciences, State University of Ghent, Coupure Links 653, B-9000 Ghent, Belgium Instrumental aroma analysis, involving headspace sampling, analysis and identification of aroma compounds by gas chromatography-mass spectrometry allows evaluation of the different flavor determinating parameters in fruits. In apples e.g. aroma compounds, mainly esters, are formed gradually during ripening in a manner which parallels the respiration climacteric towards a maximum. Headspace gas chromatography permits following the complete ripening process. Measurement during the early picking period, allows prediction of the earliest acceptable harvest date for storage apples. After storage in controlled atmosphere (CA-storage) the normal ripening pattern is disturbed and ester production diminishes as a function of storage time. Volatile analysis by non-destructive headspace techniques is also an interesting tool for flavor formation studies e.g. by treatment of intact apples with ester precursors (carboxylic acids, aldehydes, alcohols). It is generally recognised t h a t i n f r u i t and v e g e t a b l e p r o d u c t i o n more a t t e n t i o n s h o u l d be g i v e n to the h i d d e n s e n s o r y q u a l i t y paramet e r s , such as f l a v o r and t e x t u r e . These q u a l i t y a t t r i b u t e s a r e the r e s u l t o f a number o f p r e - and p o s t - h a r v e s t f a c t o r s and a r e c l o s e l y related with fruit ripening. P a i l l a r d d i s t i n g u i s h e s e x t e r n a l and i n t e r n a l f a c t o r s i n f l u e n c i n g aroma f o r m a t i o n in fruits (1). The first ones are associated with the c u l t u r e o f the p l a n t and p o s t h a r v e s t t r e a t m e n t s , the second are i n connection with the m e t a b o l i c r e g u l a t i o n o f the f r u i t . A s u r v e y i s g i v e n i n F i g u r e 1. It i s not p o s s i b l e to c o v e r these d i f f e r e n t aspects f o r a v a r i e t y of f r u i t products. T h e r e f o r e we s e l e c t e d to d i s c u s s the knowledge of the i n f l u e n c i n g parameters on a p p l e f l a v o r and to f o c u s on some r e c e n t a n a l y t i c a l a c h i e v e m e n t s , which a r e o f importance f o r the study of these e f f e c t s . E x c e l l e n t work has a l r e a d y been done c o n c e r n i n g the s e p a r a t i o n and identification of aroma g i v i n g compounds. The s t a t e o f the a r t up to 1982 on a p p l e f l a v o r has been c l e a r l y r e v i e w e d by D i m i c k and H o s k i n ( 2 ) , c o v e r i n g i m p o r t a n t s t u d i e s by 0097-6156/89/0388-0023$06.00/0 ° 1989 American Chemical Society

In Flavor Chemistry; Teranishi, R., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1989.

24

FLAVOR CHEMISTRY: TRENDS AND

DEVELOPMENTS

EXTERNAL FACTORS


PRE-HARVEST : SOIL/HYDROPONIC CULTURE FERTILIZATION CLIMATE/IRRIGATION PICKING DATE-MATURITY POST-HARVEST TECHNOLOGY : DURATION OF STORAGE STORAGE CONDITIONS : TEMPERATURE HUMIDITY GAS COMPOSITION INTERNAL FACTORS GENETIC CONTROL : CULTIVAR METABOLIC REGULATION : ETHYLENE RESPIRATION Figure

1 . Factors influencing

f l a v o r formation

in fruits.

Drawert ( 3 ) , F l a t h ( 4 ) , Guadagni ( 5 ) , W i l l i a m s (6) and many o t h e r s . Some compounds have been claimed to be important c o n t r i b u t o r s to apple aroma : e.g. ethyl 2-methylbutanoate, n . h e x a n a l , t r a n s - 2 h e x e n a l and 4 - m e t h o x y a l l y l b e n z e n e . However i t seems t h a t a p p l e aroma i s not a m a t t e r o f a l i m i t e d number o f c h a r a c t e r impact compounds but i s due to complex m i x t u r e o f a l c o h o l s , aldehydes, C^-C^ esterified a c i d s , e s t r a g o l and t e r p e n e s (_7). R e c e n t l y Y a j i m a e t a l . (8) i d e n t i f i e d 22 new apple flavor components i n the steam d i s t i l l a t e of Kogyoku a p p l e s , which c o r r e s p o n d to American J o n a t h a n a p p l e s . In an i n t e r e s t i n g approach u s i n g "Charm" a n a l y s i s on f o r t y d i f f e r e n t apple c u l t i v a r s Cunningham e t a l . i n d i c a t e d the importance to a p p l e odour o f 3-damascenone, b u t y l - , 3 - m e t h y l b u t y l - and h e x y l hexanoates, a l o n g w i t h e t h y l , p r o p y l and h e x y l b u t a n o a t e s . Flavor formation in fruit p r o d u c t s has a l s o e x t e n s i v e l y been reviewed ( 1 0 ) . A d i s t i n c t i o n can be made between the p r i m a r y aroma components, which are b i o s y n t h e s i z e d by the whole f r u i t and secondary aroma compounds (e.g. h e x a n a l , 2 - h e x e n a l ) , formed a f t e r d i s r u p t i o n o f the c e l l s during processing or chewing (11) . The importance o f the p e e l f o r aroma f o r m a t i o n has a l s o been s t r e s s e d by s e v e r a l a u t h o r s (12). An e x t e n s i v e l i t e r a t u r e on the r e s p i r a t i o n c l i m a c t e r i c ( 1 3 ) , the r o l e o f e t h y l e n e (14) and the enzymes and s u b s t r a t e s r e q u i r e d f o r b i o s y n t h e s i s i s a v a i l a b l e (15). The t o p i c o f a p p l e f l a v o r s h o u l d a l s o be c o n s i d e r e d i n r e l a t i o n to the modern developments in isolation and p r e p a r a t i o n t e c h n o l o g y . The i m p o r t a n t i n f l u e n c e of the sample p r e p a r a t i o n t e c h n i q u e s on the c o m p o s i t i o n o f b i o l o g i c a l l y d e r i v e d aromas was r e c e n t l y reviewed by Parliment (16). The g o a l of t h i s a r t i c l e i s to p r e s e n t a p r o c e d u r e f o r aroma a n a l y s i s , c o n s i s t i n g o f dynamic headspace sampling of i n t a c t f r u i t s , f o l l o w e d by high r e s o l u t i o n gas chromatography - mass s p e c t r o m e t r y . This procedure allows e v a l u a t i o n of the i n f l u e n c e of c u l t i v a r , p i c k i n g date and c o n t r o l l e d atmosphere s t o r a g e on aroma development i n apples. The a n a l y t i c a l system i s a l s o o f importance for studying b i o g e n e s i s o f aroma components i n i n t a c t f r u i t s .


3.

DIRINCKETAL

Aroma Development in Ripening Fruits


A n a l y s i s Technology;

Dynamic Headspace Gas

Chromatography

Dynamic headspace sampling on Tenax GC, by t r a p p i n g v o l a t i l e s , l i b e r a t e d d u r i n g d i s i n t e g r a t i o n o f f r u i t s , has been p r e v i o u s l y d e s c r i b e d by the a u t h o r s (17,18). This p r o c e d u r e a l l o w s the i s o l a t i o n o f an i n s t a n t aroma and i m i t a t e s the e a t i n g mechanism. Dynamic headspace sampling also offers the possibility f o r i s o l a t i o n of v o l a t i l e s , r e l e a s e d from i n t a c t a p p l e s , which i n the case o f a p p l e aroma, a l l o w s following the complete r i p e n i n g process. A t y p i c a l apparatus f o r i s o l a t i o n o f v o l a t i l e s from i n t a c t a p p l e s i s presented i n F i g u r e 2. The d e s s i c a t o r s are c o n t i n u o u s l y f l u s h e d w i t h a i r and a t r e g u l a r i n t e r v a l s f o r sampling a Tenax a d s o r b e r i s a t t a c h e d to the o u t l e t of the d e s s i c a t o r . F o r q u a n t i t a t i v e a d s o r p t i o n h i g h amounts o f Tenax GC a r e used : 5 g o f Tenax GC 60/80 mesh, packed i n g l a s s tubes ( i . d . 1.6 cm, l e n g t h 10 cm). Sample r e c o v e r y can be performed by heat d e s o r p t i o n and c o l l e c t i o n in a cold t r a p , which allows a sharp injection on 0.5 mm i.d. capillary columns. T h e r e f o r e the gas chromatographs a r e m o d i f i e d w i t h a s e l f c o n s t r u c t e d i n j e c t i o n system, c o n s i s t i n g of a d e s o r p t i o n oven and a t h e r m o s t a t e d t w o - p o s i t i o n s i x port, high temperature injection valve. The f u l l experimental d e t a i l s of the a n a l y s i s system have been d e s c r i b e d b e f o r e ( 1 9 ) . As an i l l u s t r a t i o n o f the r e s u l t s o b t a i n e d by t h i s system i n F i g u r e 3 the chromatogram of Spartan variety a p p l e v o l a t i l e s , i s o l a t e d by dynamic headspace sampling of the intact fruits, is presented t o g e t h e r w i t h the i d e n t i f i e d compounds. I n f l u e n c e o f P i c k i n g Date As aroma i s one o f the key f a c t o r s i n f l a v o r q u a l i t y , i t can be used as a c r i t e r i o n f o r e v a l u a t i o n o f f l a v o r q u a l i t y o f a p p l e s . By f o l l o wing the e v o l u t i o n o f the v o l a t i l e c o m p o s i t i o n a complete p i c t u r e o f the dynamic f l a v o r q u a l i t y p r o c e s s can be o b t a i n e d . In F i g u r e 4 the sum o f e s t e r s i s p r e s e n t e d as a f u n c t i o n o f days o f r i p e n i n g i n s t a n dard c o n d i t i o n s (18°C) f o r Golden D e l i c i o u s a p p l e s . L a t e p i c k e d a p p l e s show an immediate aroma development and r e a c h a c o n s i d e r a b l e h i g h e r maximum compared to e a r l y picked apples. From t h i s p i c t u r e one can c o n c l u d e that f o r optimum a p p l e q u a l i t y the p i c k i n g date s h o u l d be r e l a t e d t o the consumption p e r i o d . Figure 4 also i l l u s t r a t e s the w e l l known f a c t t h a t too e a r l y p i c k e d a p p l e s w i l l never d e v e l o p t h e i r f u l l f l a v o r . On the c o n t r a r y for apples picked a f t e r the optimum h a r v e s t date s t o r a g e a b i l i t y i s reduced. E s t a b l i s h i n g the p r o p e r time t o h a r v e s t apple c u l t i v a r s i n r e l a t i o n to s t o r a g e d i s o r d e r s and i n o r d e r to ensure h i g h q u a l i t y a f t e r l o n g - t e r m CA storage requires special consideration. Post harvest r i p e n i n g i n apples i s a s s o c i a t e d w i t h c o l o r changes, s o f t e n i n g o f the f r u i t f l e s h , changes o f the sour-sweet b a l a n c e and a r i s e of e t h y l e n e . S e v e r a l o f t h e s e parameters have been used f o r d e t e r m i n a t i o n o f the optimum h a r v e s t d a t e . Unfortunately these h a r v e s t i n d i c e s have no p r e d i c t i v e v a l u e . As aroma i s an e x p o n e n t i a l l y e v a l u a t i n g parameter d u r i n g the e a r l y p i c k i n g p e r i o d , we have used the s e n s i t i v i t y of the d e s c r i b e d a n a l y t i c a l p r o c e d u r e f o r p r e d i c t i o n o f optimum h a r v e s t date f o r s t o r a g e a p p l e s . At regular intervals i n the premature stage the sum of e s t e r s or b u t y l a c e t a t e (for faster a n a l y s i s ) i s measured a f t e r 2 days o f r i p e n i n g i n s t a n d a r d c o n d i tions. The optimum h a r v e s t time f o r s t o r a g e a p p l e s can be p r e d i c t e d


25


26

FLAVOR CHEMISTRY: TRENDS AND

DEVELOPMENTS

Figure 2. Apparatus f o r dynamic headspace s a m p l i n g o f v o l a t i l e s , r e l e a s e d from i n t a c t apples. 1 = high purity purge air, 2 » dessicators, 3 = thermostated w a t e r b a t h (18°C), 4 - f i n e m e t e r i n g v a l v e , 5 = Tenax a d s o r b e r , 6 = f l o w meter, 7 " w e t - t e s t m e t e r .



7?

1

1

1

β

° ί ,?ί.14

«

F i g u r e 3. Chromatogram o f the aroma o f S p a r t a n : 30mx0.5mm RSL-150 FS capillary column, linear t e m p e r a t u r e programming 20°C + 220°C, FID-detection. 1 = e t h y l acetate, 2 = butanol, 3 = e t h y l propionate, 4 = propyl acetate, 5 2-methylbutanol, 6 = 2-methylpropyl acetate, 7 = e t h y l butanoate, 8 = p r o p y l p r o p i o n a t e , 9 = b u t y l a c e t a t e , 10 = e t h y l 2 - m e t h y l b u t a n o a t e , 11 2-methylbutyl acetate, 12 = propyl b u t a n o a t e , 13 = b u t y l p r o p i o n a t e , 14 = p e n t y l a c e t a t e , 15 = p r o p y l 2m e t h y l b u t a n o a t e , 16 = 2 - m e t h y l p r o p y l b u t a n o a t e , 17 = b e n z a l d e h y d e , 18 = b u t y l b u t a n o a t e , 19 - e t h y l h e x a n o a t e , 20 = p e n t y l p r o p i o n a t e , 21 = hexyl acetate, 22 = b u t y l 2-methylbutanoate, 23 = 2-methylbutyl b u t a n o a t e , 24 = p r o p y l h e x a n o a t e , 25 = h e x y l p r o p i o n a t e , 26 - h e p t y l a c e t a t e , 27 = p e n t y l 2 - m e t h y l b u t a n o a t e , 28 = hexyl 2-methylpropionat e , 29 = b u t y l hexanoate + h e x y l b u t a n o a t e , 30 = 4-methoxyallylbenzene, 31 - h e x y l 2 - m e t h y l b u t a n o a t e , 32 - 2 - m e t h y l b u t y l h e x a n o a t e , 33 = h e x y l p e n t a n o a t e + p e n t y l h e x a n o a t e , 34 = h e x y l h e x a n o a t e , 35 = α-farnesene.

UUJJ

3„4


>3

s-

η

S

28

FLAVOR CHEMISTRY: TRENDS AND DEVELOPMENTS

by l i n e a r r e g r e s s i o n between the l o g a r i t h m o f the b u t y l a c e t a t e c o n c e n t r a t i o n and the p i c k i n g d a t e . The r e s u l t s f o r Golden D e l i c i o u s a p p l e s i n the season 1986 a r e p r e s e n t e d i n F i g u r e 5. As e v a l u a t e d by s t o r a g e e x p e r i m e n t s and subsequent aroma a n a l y s e s optimum h a r v e s t c o r r e s p o n d s t o a b u t y l a c e t a t e c o n c e n t r a t i o n o f 0.4 ug/kg a p p l e s / 6 1 dynamic headspace.


400

F i g u r e 4. I n f l u e n c e o f date o f g a t h e r i n g on the e v o l u t i o n o f the sum of Golden D e l i c i o u s e s t e r s as a f u n c t i o n o f days o f r i p e n i n g i n standard c o n d i t i o n s .


3.

Postharvest



DIRINCKETAL

29

P r e s e r v a t i o n Technology

In most c o u n t r i e s an i m p o r t a n t p a r t o f the a p p l e p r o d u c t i o n i s s t o red. E a r l y s t u d i e s showed t h a t h i g h carbon dioxide levels and low oxygen atmospheres could delay r i p e n i n g . Choosing the s t o r a g e c i r cumstances seems l a r g e l y t o have been a q u e s t i o n of t r i a l and e r r o r and an i m p r e s s i v e amount o f work has been done about s t o r a g e i n h i g h carbon d i o x i d e and/or low oxygen atmospheres ( 2 0 ) , under h y p o b a r i c o r low e t h y l e n e c o n d i t i o n s (21,22). Aroma a n a l y s i s can be used as an o b j e c t i v e c r i t e r i o n f o r measurement o f the e v o l u t i o n o f f l a v o r q u a l i t y as a f u n c t i o n o f c o n t r o l led-atmosphere s t o r a g e time. Some y e a r s ago we measured the e v o l u t i o n o f aroma development i n Golden D e l i c i o u s a p p l e s a f t e r removal from CA-storage f o r s i x p e r i o d s d u r i n g the s t o r a g e p e r i o d by means o f a headspace t e c h n i q u e , which i s o l a t e d the v o l a t i l e s r e l e a s e d d u r i n g m a c e r a t i o n o f the f r u i t s (23). I n a l a t t e r experiment we used the non-destructive headspace sampling technique with i n t a c t apples. Because the same f r u i t s a r e sampled throughout the complete r i p e n i n g experiment t h i s t e c h n i q u e has s e v e r a l advantages, such as c o n v e n i e n ce, f a s t e r a n a l y s i s and b e t t e r r e p r o d u c i b i l i t y . An i l l u s t r a t i o n o f the aroma development after removal from v e n t i l a t e d controlled atmosphere (Temperature : 0,5°C, 16% carbon d i o x i d e , 5% oxygen) i s shown i n f i g u r e 6. R e s u l t s o b j e c t i v e l y i n d i c a t e an i m p o r t a n t d e c r e a se i n aroma q u a l i t y a f t e r l o n g CA-storage. Postharvest p r e s e r v a t i o n technology i s s t i l l i n p r o g r e s s and more and more a t t e n t i o n i s g i v e n t o low oxygen and t o low carbon d i oxide levels (ULO-storage) and t o e t h y l e n e s c r u b b i n g . Our f u t u r e r e s e a r c h i n t h i s f i e l d w i l l be d i r e c t e d t o the e v a l u a t i o n o f these r e c e n t developments i n o r d e r t o m a i n t a i n f l a v o r q u a l i t y a f t e r l o n g storage. The p r o p o s e d technique can be u s e d t o g e t h e r w i t h p a n e l e v a l u a t i o n s t o compare d i f f e r e n t s t o r a g e c o n d i t i o n s . 250

Early

picking

Sept

23

250

Medium

picking

10

ft storage until Jan 12

ft storage until Jan 12 * storage until March 10 ifc storage until May 25

Oct

+ storage until March 10 200-

$

storage until May 25

150

100

days 10

15

20

25

30

35

10

15

20

25

F i g u r e 6. Aroma development a f t e r removal from v e n t i l a t e d atmosphere. I n f l u e n c e o f s t o r a g e time and p i c k i n g d a t e .

30

35

controlled


30

FLAVOR CHEMISTRY:

TRENDS AND

DEVELOPMENTS


Genetic Control : C u l t i v a r s One o f the important s t r a t e g i e s o f the r e c e n t q u a l i t y p o l i c y o f the a p p l e i n d u s t r y c o n s i s t i n d i v e r s i f i c a t i o n o f the number o f c u l t i v a r s , which are o f f e r e d to the consumer. For i n s t a n c e the B e l g i a n p r o d u c t i o n , which around 1980 c o n s i s t e d of 75% Golden D e l i c i o u s has been d i v e r s i f i e d to about 33% Golden D e l i c i o u s , 24% J o n a g o l d , 18% Boscoop, 6.5% Cox's Orange P i p p i n , 4.3% G l o s t e r and o t h e r s . As apple c u l t i v a r s can d i f f e r markedly i n aroma c h a r a c t e r , the d e c i s i o n which c u l t i v a r s s h o u l d be grown i s o f extreme economic importance. Classifi c a t i o n o f a p p l e c u l t i v a r s a c c o r d i n g to t h e i r aroma c o m p o s i t i o n can be helpful for cultivar selection and f o r d e t e c t i o n of r e l a t i o n s h i p s between c u l t i v a r s . Information on the aroma p a t t e r n s o f d i f f e r e n t a p p l e c u l t i v a r s i s s c a r c e (24,25). In a r e c e n t p r o j e c t we have determined the aroma development o f 25 a p p l e c u l t i v a r s by means o f dynamic headspace i s o l a t i o n o f the v o l a t i l e s r e l e a s e d by i n t a c t f r u i t s , f o l l o w e d by h i g h r e s o l u t i o n gas chromatography and mass s p e c t r o m e t r i c i d e n t i f i c a t i o n . Data a c q u i s i tion resulted i n 25 tables with q u a n t i t a t i v e data f o r about f o r t y compounds a t 5 to 8 r i p e n i n g s t a g e s . For c l a s s i f i c a t i o n purposes the c o m p o s i t i o n near the maximum o f aroma development was used and 12 parameters were selected : hexanol, b u t y l a c e t a t e , hexyl a c e t a t e , b u t y l propionate, h e x y l propionate, e t h y l butanoate, p r o p y l butanoa t e , b u t y l 2-methylbutanoate, h e x y l 2-methylbutanoate, h e x y l hexanoa te, 4-methoxyallylbenzene and α-farnesene. Parameter s e l e c t i o n was guided by v i s u a l comparison o f the chromatogram and a l s o some a n a l y t i c a l and b i o c h e m i c a l i n s i g h t s were taken i n c o n s i d e r a t i o n . Hexanol was s e l e c t e d as a r e p r e s e n t a t i v e o f a l c o h o l s , which a r e c h a r a c t e r i s t i c o f some a p p l e c u l t i v a r s . The d i f f e r e n t e s t e r i f i e d C^ to C5 a c i d s were r e p r e s e n t e d by the h i g h c o n c e n t r a t i o n b u t y l and h e x y l e s t e r s , except f o r the b u t a n o a t e s . As respectively b u t y l butanoate/ethyl hexanoate and h e x y l b u t a n o a t e / b u t y l hexanoate were not w e l l s e p a r a t e d on the RSL-150 (SE-52) c a p i l l a r y column, we used e t h y l and p r o p y l butanoate for representing the butanoates and disregarded b u t y l hexanoate. Furthermore 4 - m e t h o x y a l l y l - b e n z e n e , r e s p o n s i b l e f o r the s p i c y note i n a p p l e s , and α-farnesene as the main s e s q u i t e r p e n e were used f o r c l a s s i f i c a t i o n . Data p r o c e s s i n g was performed by P r i n c i p a l Component A n a l y s i s on a p e r s o n a l IBM computer. In a f i r s t s e t the p r o c e n t u a l v a l u e s o f the s e l e c t e d parameters of the 25 apple cultivars were used. A projection of the 12-dimensional into a 2-dimensional space i s p r e s e n t e d i n F i g u r e 7. The reduced space p r e s e n t s i n an o p t i m a l way (75.5% o f the o r i g i n a l v a r i a n c e ) the r e l a t i o n s between the o b j e c t s . In the same plane the r e l a t i o n s between the v a r i a b l e s are g i v e n and can be r e l a t e d to the o b j e c t s . F i g u r e 7 shows t h a t a l a r g e group o f c u l t i v a r s i s c h a r a c t e r i s e d by h i g h c o n c e n t r a t i o n o f a c e t a t e s . High c o n c e n t r a t i o n s o f h e x a n o l and e t h y l b u t a n o a t e are t y p i c a l f o r a n o t h e r group, composed of Nico, Granny Smith, Summerred and P a u l a r e d . Boscoop and Jacques L e b e l a r e o u t l i e r s and a r e c h a r a c t e r i s e d r e s p e c t i v e l y by h i g h α-farnesene and h i g h h e x y l 2-methylbutanoate. In a second data set the relationships between the c l o s e l y related "acetate -type c u l t i v a r s was examined. The r e s u l t s f o r 17 c u l t i v a r s a r e p r e s e n t e d i n a 3 - d i m e n s i o n a l space (88% o f the o r i g i n a l variance) i n Figure 8. From t h i s p i c t u r e i t i s c l e a r that f . i . J o n a g o l d (Golden D e l i c i o u s χ Jonathan) en Golden D e l i c i o u s have v e r y l,


3.


DIRINCK ET AL.

31

Farnesene

Boscoop

• Butyl

acetate

•

*


• · PC 2

• Nico • • Granny _ . , Smith

Karmijn * Hexanol

• **· •

Jersey

Hexyl

*r acetate

• •· · •

mac

•Summered

Flavor Chemistry - American Chemical Society

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