Characterization and Measurement of Flavor Compounds - American

10 Automated Analysis of Volatile Flavor Compounds Robert G. Westendorf

Downloaded by NORTH CAROLINA STATE UNIV on December 27, 2017 | http://pubs.acs.org Publication Date: November 8, 1985 | doi: 10.1021/bk-1985-0289.ch010

Tekmar Company, Cincinnati, OH 45222-1856

Volatile organic compounds present in foods have a significant impact on flavor quality. The analysis of these compounds can be quite difficult, since the sample is often not amenable to direct GC injection, and the volatiles may be present in very low concentrations while still being important to flavor. The technique of dynamic headspace sampling was used for the isolation and concentration of volatiles prior to analysis by gas chromatography. Samples, which may be heated, are purged with an inert gas, sweeping any volatile compounds present out of the sample. The volatiles are trapped on Tenax, which is then thermally desorbed and backflushed to inject the sample into the GC. Using new instrumentation, this method was fully automated. Samples run include fruits, fruit products, edible oils, and oil-based foods. Detection limits in the low part-per-billion range were obtained with 2-8% reproducibility. The i n s t r u m e n t a l a n a l y s i s o f f l a v o r i n a food m a t e r i a l can be an e x t r e m e l y d i f f i c u l t t a s k . There a r e many f a c t o r s t h a t i n f l u e n c e f l a v o r . Of t h e s e , one o f the more i m p o r t a n t , y e t a l s o most d i f f i c u l t t o a n a l y z e , i s the p r o f i l e o f v o l a t i l e o r g a n i c compounds p r e s e n t . The d i f f i c u l t y a r i s e s from the f a c t t h a t t h e r e may be many v o l a t i l e s p r e s e n t a t v e r y low c o n c e n t r a t i o n s i n a c o m p l i c a t e d m a t r i x . A v a r i e t y o f methods have h i s t o r i c a l l y been used f o r t h i s a n a l y s i s . The m a j o r i t y o f these methods have u t i l i z e d gas chromatography (GC), d i f f e r i n g i n chromatographic parameters and sample p r e p a r a t i o n t e c h n i q u e s . Chromatographic systems have e v o l v e d tremendously i n r e c e n t y e a r s . Column t e c h n o l o g y has advanced t o a v e r y h i g h l e v e l o f s e p a r a t i o n power. Sample p r e p a r a t i o n t e c h n i q u e s , on the o t h e r hand, have not e v o l v e d as r a p i d l y as GC t e c h n o l o g y . A number o f d i f f e r e n t p r e p a r a t i o n t e c h n i q u e s have been used f o r the a n a l y s i s o f f l a v o r 0097-6156/85/0289-0138$06.00/0 © 1985 American Chemical Society

Bills and Mussinan; Characterization and Measurement of Flavor Compounds ACS Symposium Series; American Chemical Society: Washington, DC, 1985.


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v o l a t i l e s , i n c l u d i n g s o l v e n t e x t r a c t i o n , steam d i s t i l l a t i o n , e q u i l i b r i u m headspace s a m p l i n g , and dynamic headspace s a m p l i n g . Of these methods, dynamic headspace sampling i s p r o b a b l y the l e a s t w e l l known, y e t i t has a number of advantages over o t h e r t e c h n i q u e s i n use. Methods u t i l i z i n g dynamic headspace t e c h n i q u e s were r e p o r t e d as e a r l y as 1960 (_1 ). These methods g e n e r a l l y u t i l i z e d c r y o g e n i c t r a p s , or c r y o g e n i c a l l y c o o l e d t r a p s c o n t a i n i n g column p a c k i n g materials (2) or molecular sieves ( 3 ) . Dynamic headspace a n a l y s i s (DHA), a l s o know as purge and t r a p a n a l y s i s , u t i l i z i n g new porous polymers as t r a p p i n g agents and i n c o r p o r a t i n g m u l t i - p o r t v a l v e s f o r f l o w s w i t c h i n g was i n t r o d u c e d f o r the a n a l y s i s of o r g a n i c c o n t a m i n a n t s i n water i n 1974 ( 4 ) . Various forms of DHA have been used f o r a v a r i e t y of food samples. F r u i t s and j u i c e s have been i n v e s t i g a t e d by a number of r e s e a r c h e r s . Schamp and D i r i n c k (_5) found over f o r t y compounds i n a study of s t r a w b e r r y v a r i e t i e s , as w e l l as f i n d i n g 22 compounds i n Golden D e l i c i o u s a p p l e s . Keenaghan and Meyers ( 6 ) r e p o r t e d on the GC/MS i d e n t i f i c a t i o n of over twenty d i f f e r e n t compounds i n s e v e r a l v a r i e t i e s of a p p l e s and a p p l e p r o d u c t s . A d d i t i o n a l work w i t h a p p l e s has been r e p o r t e d by Westendorf (]_) » T h i s same paper r e p o r t e d the a n a l y s e s of d a i r y p r o d u c t s , v e g e t a b l e o i l s , and a r t i f i c i a l flavors. E d i b l e o i l s have been e x t e n s i v e l y s t u d i e d s i n c e the p r e s e n c e of v o l a t i l e s was f i r s t r e c o g n i z e d as an i n d i c a t o r of o i l q u a l i t y ( 8 ) . C o n s i d e r a b l e work w i t h o i l s and o i l - b a s e d foods u s i n g manual p r o c e d u r e s has been r e p o r t e d by J a c k s o n e t a l . ( 9 , H ) ) , Dupuy e t a l . (JUL ,_12), and S e l k e (_13). I n 1983 R o b e r t s ( 1 4 ) f i r s t r e p o r t e d the use o f an automated DHA procedure f o r o i l v o l a t i l e s . P r i n c i p l e of O p e r a t i o n DHA i s based on the p a r t i t i o n i n g of v o l a t i l e compounds between a sample and the vapor phase above the sample a t a r a t e dependent on a v a r i e t y of f a c t o r s . These i n c l u d e the v o l a t i l i t y of the s u b j e c t compound, i t s s o l u b i l i t y i n the sample m a t r i x , homogeneity of the m a t r i x , t e m p e r a t u r e , and sample c o n t a i n e r configuration. I n e q u i l i b r i u m headspace a n a l y s i s , the sample i s s e a l e d i n a c l o s e d v e s s e l and the v o l a t i l e s a r e a l l o w e d t o e q u i l i b r a t e between the sample and vapor phase. An a l i q u o t of the vapor phase i s then i n j e c t e d i n t o a GC f o r a n a l y s i s . In DHA, the sample i s purged w i t h an i n e r t gas, sweeping the v o l a t i l e s out of the sample c o n t a i n e r . The purge gas i s t h e n passed through a s h o r t column c o n t a i n i n g a porous polymer a d s o r b e n t which s e l e c t i v e l y r e t a i n s the sample compounds w h i l e a l l o w i n g the purge gas and any water vapor t o pass t h r o u g h . By p u r g i n g i n t h i s manner, the e n t i r e o r g a n i c c o n t e n t s o f the vapor phase can be s u b j e c t e d t o GC a n a l y s i s , not j u s t an a l i q u o t . In a d d i t i o n , s i n c e the purge gas i s c o n t i n u a l l y b e i n g removed from the sample the c o n c e n t r a t i o n of o r g a n i c s i n the vapor above the sample remains e s s e n t i a l l y z e r o . T h i s s i g n i f i c a n t l y enhances r e c o v e r y by p r o m o t i n g f u r t h e r p a r t i t i o n i n g of the v o l a t i l e s i n t o the vapor s t a t e . A f t e r the purge s t e p i s c o m p l e t e d , the a d s o r b e n t column i s h e a t e d t o r e l e a s e the o r g a n i c s and b a c k f l u s h e d v i a a 6 - p o r t v a l v e t o sweep the sample t o the GC. When u s i n g c a p i l l a r y


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columns, a c r y o g e n i c f o c u s i n g s t e p i s employed t o sharpen the i n j e c t i o n p r o f i l e ( 1_5). S e p a r a t i o n and d e t e c t i o n are c a r r i e d out i n the GC, n o r m a l l y under temperature programmed c o n d i t i o n s . A diagram o f the f l o w scheme i s i l l u s t r a t e d i n F i g u r e 1. G o a l s . The p r i m a r y g o a l o f t h i s work was t o adapt new i n s t r u m e n t a t i o n t o the f u l l y automated a n a l y s i s o f v o l a t i l e f l a v o r compounds i n foods w i t h o u t compromising any o t h e r aspect of a n a l y t i c a l c a p a b i l i t y . The g o a l s o f t h i s work, i n a d d i t i o n t o automation, i n c l u d e d : 1. Recover the maximum range o f compounds p o s s i b l e , from v e r y v o l a t i l e gases t o h i g h e r b o i l i n g , l e s s v o l a t i l e compounds. 2. A c h i e v e the maximum s e n s i t i v i t y p o s s i b l e , p r e f e r a b l y t o p a r t - p e r - b i 1 1 i o n (ppb) l e v e l s , s i n c e many compounds have important o r g a n o l e p t i c q u a l i t i e s a t low l e v e l s . 3. A c h i e v e the b e s t r e p r o d u c i b i l i t y p o s s i b l e . 4. Keep a r t i f a c t f o r m a t i o n t o a minimum, t o ensure t h a t the v o l a t i l e compounds found are those a c t u a l l y p r e s e n t i n the sample. 5. E l i m i n a t e c r o s s - c o n t a m i n a t i o n between samples. Achievement o f g o a l s 1-5 was c o n s i d e r e d n e c e s s a r y f o r the g o a l , t o t a l automation, t o be o f any p r a c t i c a l v a l u e .

primary

I n s t r u m e n t a t i o n f o r Automated A n a l y s e s . A l l samples were run u s i n g c o m m e r c i a l l y a v a i l a b l e automated DHA equipment i n t e r f a c e d to a m i c r o p r o c e s s o r GC. The DHA apparatus c o n s i s t s o f t h r e e p a r t s : the b a s i c c o n c e n t r a t o r (TEKMAR Model 4000), i n c o r p o r a t i n g the purge system, s w i t c h i n g v a l v e , and adsorbent; a t e n - p o s i t i o n automatic sampler (TEKMAR Model 4200); and a c a p i l l a r y column c r y o g e n i c t r a p (TEKMAR Model 1000). Under normal c o n d i t i o n s t h e o p e r a t o r f i r s t s e t s a l l o p e r a t i n g c o n d i t i o n s , loads the samples, r a i s e s the sample h e a t e r s ( h i g h performance e l e c t r i c m a n t l e s ) , and p l a c e s the i n s t r u m e n t s i n a u t o m a t i c mode. The automatic sampler advances t o the f i r s t p o s i t i o n and s i g n a l s the c o n c e n t r a t o r t o s t a r t . The prepurge, p r e h e a t , and purge steps are performed by the c o n c e n t r a t o r , which then sends a ready s i g n a l t o the c a p i l l a r y c r y o t r a p . When the c r y o t r a p a l s o r e c e i v e s a ready s i g n a l from the GC, i t w i l l c o o l t o a p r e s e t temperature and s i g n a l the c o n c e n t r a t o r t o s t a r t d e s o r b i n g the sample. When d e s o r p t i o n i s complete the c r y o t r a p w i l l heatup t o i n j e c t the sample and s i m u l t a n e o u s l y output s i g n a l s t o s t a r t the temperature program o f the GC and t o s t a r t data a c q u i s i t i o n on an i n t e g r a t o r . The i n t e g r a t o r w i l l a l s o read the sample p o s i t i o n number from the a u t o m a t i c sampler v i a a BCD i n t e r f a c e . While t h e GC run c o n t i n u e s the c o n c e n t r a t o r w i l l r e c o n d i t i o n and then c o o l the adsorbent t r a p . When the t r a p has c o o l e d the automatic sampler w i l l a g a i n advance and the above c y c l e w i l l r e p e a t . Experimental M a t e r i a l s . Samples were o b t a i n e d from a v a r i e t y o f s o u r c e s . F r u i t s and j u i c e s were purchased l o c a l l y . A l l o i l s , peanut b u t t e r , and food s t a r c h samples were s u p p l i e d by food p r o c e s s o r s . F l a v o r compounds used i n the p r e p a r a t i o n o f standards were



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Injection Port

F i g u r e 1:

Gas f l o w scheme of dynamic headspace gas chromatography system.


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purchased from A l d r i c h (Milwaukee, WI) and used w i t h o u t f u r t h e r purification. C o n c e n t r a t o r C o n d i t i o n s . The c o n c e n t r a t o r system used was a TEKMAR Model 4000 w i t h a TEKMAR Model 4200 t e n p o s i t i o n a u t o m a t i c sampler. A TEKMAR Model 1000 C a p i l l a r y I n t e r f a c e was used f o r i n t e r f a c e t o the GC. The o p e r a t i n g c o n d i t i o n s were as f o l l o w s : sample s i z e s - peanuts, f r u i t s : 1.00g, peanut b u t t e r , food starch: lOOmg, o i l s : 0.5ml, j u i c e s : 5.0ml. Sample temperatures - f r u i t s , j u i c e s , food s t a r c h : ambient (23 C ) , peanuts, peanut butter: 100 C, o i l s : 150 C. Prepurge 3.5 min. a t 50ml/min. n i t r o g e n , preheat 3 min. f o r 100 samples, 5 min. f o r 150 samples, purge 10 min. a t 50ml/min. n i t r o g e n . The t r a p was 12" X 1/8" s t a i n l e s s s t e e l packed w i t h 24cm (approx. 150mg) Tenax TA (Chrompack, B r i d g e w a t e r , N J ) . The t r a p was p r e h e a t e d at 175°C and then desorbed a t 180°C f o r 4 min. The t r a p was baked a t 225 C f o r 10 min. between runs. A l l v a l v e l i n e s and sampler mounts were c o n t i n u o u s l y heated t o 100 C. The C a p i l l a r y I n t e r f a c e was c o o l e d w i t h l i q u i d n i t r o g e n f o r c r y o t r a p p i n g t h e desorbed sample and heated f o r 10 seconds f o r GC i n j e c t i o n . Gas Chromatograph C o n d i t i o n s . The GC was a VARIAN 6000 w i t h a flame i o n i z a t i o n d e t e c t o r . The i n j e c t i o n p o r t was heated a t 200°, the d e t e c t o r a t 250°. The d e t e c t o r range was 10-12 AFS, a t t e n u a t i o n 64 except as noted. The column was 25m X 0.32mm f u s e d s i l i c a DB5 w i t h a 1.0 m i c r o n f i l m t h i c k n e s s (bonded SE54, J&W S c i e n t i f i c , Rancho Cordova, CA). The c a r r i e r gas was hydrogen a t 47 cm/s. The column was temperature programmed from an i n i t i a l temperature o f 35 C, h e l d f o r 4 min., t o a f i n a l temperature o f 200 C a t 4°C/min. R e s u l t s and D i s c u s s i o n V o l a t i l i t y Range. The range o f f l a v o r compounds r e c o v e r e d depends on a v a r i e t y o f f a c t o r s . The two p r i m a r y f a c t o r s a r e t h e sample m a t r i x and temperature. The m a t r i x can a f f e c t r e c o v e r y i n two ways. The f i r s t i s the s o l u b i l i t y o f t h e f l a v o r compounds i n the m a t r i x . Compounds t h a t have a poor s o l u b i l i t y w i l l be purged more e f f i c i e n t l y than compounds o f h i g h s o l u b i l i t y . The second i s t h a t v o l a t i l e s may be p h y s i c a l l y bound i n the sample. Flavor compounds may be p r e s e n t p r i m a r i l y i n t h e i n t e r i o r o f a sample, as i s the case w i t h c o f f e e beans. I n t h i s case t h e mass t r a n s f e r r a t e o f the v o l a t i l e s through t h e m a t r i x becomes a l i m i t i n g f a c t o r . F o r samples o f t h i s t y p e , g r i n d i n g o r o t h e r w i s e homogenizing the sample can s i g n i f i c a n t l y i n c r e a s e the r e c o v e r y . However, c a r e must be taken t o a v o i d the p o s s i b l e l o s s o r adulteration of v o l a t i l e s during t h i s process. A cryogenic g r i n d i n g procedure developed f o r p l a s t i c s (16) has been s u c c e s s f u l l y a p p l i e d t o food m a t e r i a l s i n the a u t h o r ' s laboratory. I n c r e a s i n g the temperature o f a sample s e r v e s t o i n c r e a s e r e c o v e r i e s by i n c r e a s i n g the vapor p r e s s u r e o f the v o l a t i l e compounds. The e f f e c t o f i n c r e a s i n g temperature on a c o r n o i l sample i s i l l u s t r a t e d i n F i g u r e 2. Temperature i s g e n e r a l l y the



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Recovery of c o r n o i l v o l a t i l e s w i t h i n c r e a s i temperature.


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p r i m a r y v a r i a b l e used i n o p t i m i z i n g r e c o v e r y . The r e c o v e r i e s o f a number o f r e p r e s e n t a t i v e compounds from c o r n o i l a r e l i s t e d i n Table I .


Table I . R e p r e s e n t a t i v e Compounds i n Corn O i l , 150 i t e n t i o n Time 1.03 min. 1.36 2.12 3.67 3.75 5.33 6.16 6.24 6.58 7.67 9.07 9.15 12.06 14.79 14.94 16.79 17.68 ND: n o t determined

Compound Butane Pentane Hexane Heptane 1-Butanal 1-Pentanal Octane 1-Hexanal 2-0ctene(a) trans-2-Hexenal Nonane 1-Heptanal Decane Undecane Amyl A l c o h o l ( a ) 1-Hexanol(a) l-Nonanol(a)

Recovery 847o

82 74 65 47 50 61 51 ND 59 55 43 50 41 ND ND ND a: t e n t a t i v e

Reproduc: 10.57o

2.9 5.3 3.6 3.6 4.9 4.0 4.9 7.9 7.8 3.4 3.4 1.9 6.0 4.9 8.3 2.8 identification

A p o t e n t i a l problem w i t h h e a t i n g some samples i s t h e presence o f l a r g e amounts o f water vapor. The Tenax adsorbent used i s h y d r o p h o b i c , and does n o t t r a p w a t e r . However, water vapor w i l l condense on a c o l d Tenax t r a p . Excess amounts o f water c a n i n t e r f e r e w i t h an a n a l y s i s i n two ways. I f the amount o f water p r e s e n t i s e x t r e m e l y h i g h , i t w i l l condense on t h e Tenax i n sufficent quantity to p h y s i c a l l y block a s i g n i f i c a n t portion of the a v a i l a b l e t r a p p i n g s u r f a c e . T h i s l e a d s t o reduced t r a p p i n g e f f i c i e n c y , d e g r a d i n g both s e n s i t i v i t y and r e p r o d u c i b i l i t y . F o r samples o f t h i s t y p e , t h e sample temperature must n o t be r a i s e d above 65 t o 95 , depending on the water c o n t e n t o f t h e sample. At 60 , however, even aqueous samples can be run w i t h o u t l o s i n g t r a p p i n g e f f i c i e n c y ( Γ7 ). A second way i n which water may i n t e r f e r e i s i n t h e GC s e p a r a t i o n and d e t e c t i o n . Water may degrade t h e column used, o r i n t e r f e r e i n t h e d e t e c t i o n p r o c e s s . Samples t h a t have i n t r o d u c e d s u f f i c i e n t water i n t o t h e column t o e x t i n q u i s h t h e flame i o n i z a t i o n d e t e c t o r have been encountered i n the a u t h o r ' s l a b o r a t o r y . S i n c e water i s n o t t r a p p e d on t h e Tenax, i t i s p o s s i b l e t o remove most o f i t by p a s s i n g d r y n i t r o g e n through t h e t r a p b e f o r e t h e d e s o r p t i o n s t e p (18). The v o l a t i l i t y range o f t h e compounds t h a t can be a n a l y z e d by dynamic headspace c o n c e n t r a t i o n extends from o r g a n i c compounds t h a t a r e gases a t room temperature up t o compounds c o n t a i n i n g about t e n t o t h i r t e e n carbon atoms p e r m o l e c u l e , depending on t h e number and n a t u r e o f any s i d e c h a i n s . Table I l i s t s a number o f r e p r e s e n t a t i v e compounds i n a c o r n o i l . Table I I l i s t s some


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a d d i t i o n a l compounds commonly encountered i n v e g e t a b l e o i l samples. Compounds t y p i c a l l y found i n f r u i t samples i n c l u d e p r i m a r i l y e t h y l e s t e r s w i t h a number o f aldehydes and a l c o h o l s i n the t h r e e t o n i n e carbon range. Table I I I l i s t s the GC/MS i d e n t i f i c a t i o n s o f the compounds encountered i n an apple sample (from r e f . 6 ) .


Table I I : A d d i t i o n a l Compounds Found i n V e g e t a b l e Benzaldehyde cis-2-trans-4-Decadienal 2-Decenal 2-Heptanone 1-Nonanal Octenal

Benzyl A l c o h o l trans-2-trans-4-Decadienal 2,4-Heptadienal M e t h y l E t h y l Ketone 2-Nonanone 1-Pentanol

Oils

1-Butenal 2-Decanone Diacetyl 1-Hexanol Octadiene

Table I I I : GC/MC I d e n t i f i c a t i o n o f Apple V o l a t i l e s (from r e f . 6) Compound

Compound

N-Propyl B u t y r a t e Butanal N-Butyl Propionate Ethyl Acetate N-Amyl A c e t a t e 1-Butanol M e t h y l Caproate N-Propyl A c e t a t e Ethyl-2-Methyl Butyrate M e t h y l Butanoate N-Butyl-N-Butyrate 2-Methyl B u t a n o l Ethyl-N-Caproate N-Hexanal N-Hexyl A c e t a t e Ethyl-N-Butanoate I s o p r o p y l Hexanoate N-Butyl A c e t a t e 1-Hexanol 2-Hexanal 2-Methyl B u t y l A c e t a t e N-Hexyl-N-Butyrate 2-Methyl-2-Methyl-Propyl Butyrate

S e n s i t i v i t y . The s e n s i t i v i t y o b t a i n a b l e depends p r i m a r i l y on the e f f i c i e n c y w i t h which a compound i s r e c o v e r e d from the sample. As w i t h the v o l a t i l i t y range, r e c o v e r y g e n e r a l l y i s most e f f e c t i v e l y i n c r e a s e d by r a i s i n g the sample temperature. A d d i t i o n a l f a c t o r s a f f e c t i n g s e n s i t i v i t y i n c l u d e t r a p p i n g and d e s o r p t i o n e f f i c i e n c i e s , column r e s o l u t i o n , i n t e r f e r e n c e s , and detector s e n s i t i v i t y . F o r o i l s the lower l i m i t o f d e t e c t i o n f o r the m a j o r i t y of the compounds l i s t e d i n T a b l e s I and I I i s on the o r d e r of 1 t o 100 ppb. F o r o i l samples, nonane, which i s o f t e n added as an i n t e r n a l s t a n d a r d , i s d e t e c t a b l e t o l e s s than 5ppb. R e p r o d u c i b i l i t y . R e p r o d u c i b i l i t y i s a t h i r d f a c t o r t h a t depends p r i m a r i l y on r e c o v e r y . As a g e n e r a l r u l e , r e p r o d u c i b i l i t y improves as r e c o v e r y i n c r e a s e s . For the m a j o r i t y o f compounds f o r which the r e c o v e r y i s g r e a t e r than 40%, the r e p r o d u c i b i l i t y w i l l be on the o r d e r o f 2 - 8% r e l a t i v e s t a n d a r d d e v i a t i o n (RSD). T h i s number i s s t r o n g l y a f f e c t e d by column r e s o l u t i o n , however, s i n c e many foods samples tend t o g i v e c o m p l i c a t e d chromatograms. As the r e c o v e r y drops below 40% the r e p r o d u c i b i l i t y r a p i d l y d e t e r i o r a t e s . For v e r y v o l a t i l e compounds, the purge e f f i c i e n c y


146


i s g e n e r a l l y h i g h , but t r a p p i n g e f f i c i e n c y may become a f a c t o r . The r e p r o d u c i b i l i t y o b t a i n e d f o r pentane i n c o r n o i l i s 2.97 RSD, but the r e p r o d u c i b i l i t y f o r butane i s 10.57 RSD. T h i s s u g g e s t s t h a t butane i s not q u a n t i t a t i v e l y t r a p p e d by Tenax. A number of r e s e a r c h e r s are c u r r e n t l y e v a l u a t i n g new s o r b e n t m a t e r i a l s , which may be used a l o n e or i n c o m b i n a t i o n w i t h Tenax to improve the a n a l y s i s of compounds w i t h lower m o l e c u l a r w e i g h t s . When h e a t i n g samples, an a d d i t i o n a l f a c t o r c o n c e r n i n g r e p r o d u c i b i l i t y i s i n t r o d u c e d . Not o n l y must the h e a t e r c o n t r o l be p r e c i s e , but a time must a l s o be a l l o w e d f o r the sample temperature to e q u i l i b r a t e w i t h the h e a t e r b e f o r e s t a r t i n g to purge. Samples p l a c e d i n a h e a t e r at 150 C do not i n s t a n t l y r e a c h and e q u i l i b r a t e at 150 C . The r a t e at which the a c t u a l sample temperature r i s e s i s e x t r e m e l y d i f f i c u l t to r e p r o d u c e . If p u r g i n g i s begun b e f o r e the sample has e q u i l i b r a t e d , the r e s u l t i n g r e p r o d u c i b i l i t y may be a c c e p t a b l e , but can be improved. By a l l o w i n g a p r e h e a t time s u f f i c i e n t f o r the sample temperature to e q u i l i b r a t e b e f o r e p u r g i n g , any temperature v a r i a n c e s r e s u l t i n g from h e a t e r v a r i a t i o n s , d i f f e r i n g samples or sample s i z e s ( i . e . d i f f e r e n c e s i n heat c a p a c i t y ) , or g e o m e t r i c v a r i a t i o n s i n the sample h o l d e r ( e . g . s o l i d chunks w i l l have d i f f e r e n t amounts of s u r f a c e a r e a c o n t a c t i n g the w a l l s of the v e s s e l ) can be m i n i m i z e d . 0


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A r t i f a c t Formation. The p o t e n t i a l f o r the f o r m a t i o n of a r t i f a c t s i s p r e s e n t i n v i r t u a l l y e v e r y a n a l y t i c a l method. An advantage of dynamic headspace a n a l y s i s i s t h a t no s o l v e n t i s u s e d , e l i m i n a t i n g the g r e a t e s t s i n g l e s o u r c e o f a r t i f a c t s . U n f o r t u n a t e l y , however, o t h e r mechanisms of a r t i f a c t f o r m a t i o n do exist. The i n s t r u m e n t a t i o n must be d e s i g n e d to m i n i m i z e any possible a r t i f a c t s . When r u n n i n g heated samples t h e r e are a number o f p o s s i b l e mechanisms f o r the f o r m a t i o n of a r t i f a c t s . F o r compounds t h a t are t h e r m a l l y l a b i l e the o n l y method o f p r e v e n t i n g t h e i r d e s t r u c t i o n i s to m a i n t a i n the sample temperature below any p o s s i b l e breakdown p o i n t . F o r samples s u b j e c t to o x i d a t i o n , an a d d i t i o n a l s t e p i s needed. The v o l a t i l e s p r e s e n t i n o i l s , f o r i n s t a n c e , are p r i m a r i l y formed through o x i d a t i o n r e a c t i o n s . At the temperatures n o r m a l l y used to run o i l s , the sample w i l l r a p i d l y r e a c t w i t h any t r a c e s o f oxygen p r e s e n t to form new compounds o r i n c r e a s e the c o n c e n t r a t i o n s of o x i d a t i o n p r o d u c t s already present. T h i s can be a g g r a v a t e d by the p r e h e a t i n g s t e p used to a c h i e v e temperature r e p r o d u c i b i l i t y , a l l o w i n g a g r e a t e r time f o r the hot sample to r e a c t w i t h any oxygen d i s s o l v e d i n the sample o r p r e s e n t i n the sample v e s s e l . To remove the oxygen a "prepurge" s t e p i s added i n which the sample i s purged f o r a s h o r t time b e f o r e heat i s a p p l i e d . Any oxygen p r e s e n t w i l l be removed from the sample and r e p l a c e d by n i t r o g e n . Note t h a t t h i s prepurge gas must a l s o be p a s s e d through the Tenax to t r a p any compounds t h a t may be purged at the low t e m p e r a t u r e . After p r e p u r g i n g , the sample h e a t e r i s t u r n e d on and the temperature a l l o w e d to r i s e . The e f f e c t s of the p r e p u r g i n g s t e p can be noted i n F i g u r e 3, which i l l u s t r a t e s a c o r n o i l sample run twice under i d e n t i c a l c o n d i t i o n s w i t h the e x c e p t i o n of the p r e p u r g e s t e p . For f u r t h e r c o n s i d e r a t i o n of i n t r o d u c i n g a r t i f a c t s , the p o s s i b i l i t y of c r o s s c o n t a m i n a t i o n between samples must f i r s t be examined. I t i s p o s s i b l e i n any i n s t r u m e n t a l method f o r p a r t of



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V a r i a n c e s i n c o r n o i l sample w i t h o u t ( t o p ) and w i t h (bottom) prepurge.

American Chemical Society Library 1155 16th St.,

N.W.

Bills and Mussinan; Characterization andD.C. Measurement Washington, 20036of Flavor Compounds ACS Symposium Series; American Chemical Society: Washington, DC, 1985.

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one sample t o " c a r r y o v e r " i n t o t h e next r u n , i n f l u e n c i n g t h e r e s u l t s o f the second run. With a c o n c e n t r a t o r system, the p o s s i b l e causes o f c a r r y o v e r a r e t h r e e f o l d : incomplete d e s o r p t i o n , c o n d e n s a t i o n , and a d s o r p t i o n . Any p o s s i b l e t r a c e s o f sample components remaining on t h e Tenax t r a p as a r e s u l t o f incomplete d e s o r p t i o n a r e g e n e r a l l y removed v i a a bake s t e p , i n which the t r a p i s heated t o a temperature above t h a t used f o r d e s o r p t i o n w h i l e p a s s i n g gas through the t r a p . Condensation and a d s o r p t i o n can occur i n the v a l v i n g and t r a n s f e r t u b i n g used. These can be e l i m i n a t e d by h e a t i n g the v a l v e and l i n e s . However, t h i s introduces the p o s s i b i l i t y o f a r t i f a c t formation v i a c a t a l y t i c decomposition on t h e h o t s u r f a c e s i n t h e i n s t r u m e n t . The e f f e c t o f temperature d i f f e r e n c e s i s i l l u s t r a t e d i n F i g u r e 4 on a soy o i l sample. There was no measurable change f o r any sample r u n between ambient and 100 C t e m p e r a t u r e s . However, above 100 C some r e a c t i o n s began t o o c c u r . For the m a j o r i t y of samples, 100 C was s u f f i c i e n t t o prevent any c a r r y o v e r . F o r samples c o n t a i n i n g l a r g e amounts o f v o l a t i l e components t h a t might have c a r r y o v e r a t 100 C, h i g h e r temperatures can be used f o r r a p i d cleanup.

A p p l i c a t i o n s . I n a d d i t i o n t o o i l s and a p p l e s , a number o f o t h e r samples were used t o e v a l u a t e t h e method. These i n c l u d e d a comparison o f an orange t o a food s t a r c h w i t h added orange o i l ( F i g u r e 5 ) . As i s t y p i c a l f o r many a r t i f i c i a l f l a v o r s o r f l a v o r - a d d e d samples, t h e p r i m a r y d i f f e r e n c e s occur i n t h e e a r l y , most v o l a t i l e , p o r t i o n o f t h e chromatogram. F r u i t samples c a n a l s o be e v a l u a t e d f o r v a r i e t a l d i f f e r e n c e s ( F i g u r e 6 ) , o r f o r e v a l u a t i o n of seasonal v a r i a t i o n s , r i p e n i n g s t u d i e s , o r d e t e c t i o n of s t o r a g e abuse. The e v o l u t i o n o f f l a v o r v o l a t i l e s through two d i f f e r e n t p r o c e s s e s f o r peanut p r o d u c t s i s i l l u s t r a t e d i n F i g u r e 7. The two chromatograms r e p r e s e n t p r o d u c t s made from t h e same l o t o f raw peanuts, which had v i r t u a l l y no v o l a t i l e s p r e s e n t p r i o r t o p r o c e s s i n g . There a r e a number o f a d d i t i o n a l sample types f o r which DHA i s c u r r e n t l y b e i n g used. These i n c l u d e d a i r y p r o d u c t s , such as m i l k and cheese, carbonated beverages, powdered d r i n k mixes, beer, wine, c o f f e e , meats, s p i c e s , g r a i n s , c e r e a l s , and v a r i o u s forms o f candy. S t u d i e s underway i n c l u d e e v a l u a t i o n of p r o c e s s i n g t e c h n i q u e s , s h e l f - l i f e , p a c k a g i n g , and q u a l i t y c o n t r o l o f p r o d u c t s and incoming m a t e r i a l s . Conclusion A method f o r t h e automated a n a l y s i s o f v o l a t i l e f l a v o r compounds i n foods i s d e s c r i b e d . V o l a t i l e compounds a r e removed from t h e sample and c o n c e n t r a t e d v i a t h e dynamic headspace t e c h n i q u e , w i t h subsequent s e p a r a t i o n and d e t e c t i o n by c a p i l l a r y column gas chromatography. With t h i s method, d e t e c t i o n l i m i t s o f low ppb l e v e l s a r e o b t a i n a b l e w i t h good r e p r o d u c i b i l i t y . T h i s method has e x p e r i e n c e d r a p i d growth i n r e c e n t y e a r s , and i s now i n r o u t i n e use i n a number o f l a b o r a t o r i e s .




WESTENDORF



150


F i g u r e 5: Comparison o f f l a v o r p r o f i l e s o f orange (bottom) and o r a n g e - f l a v o r e d food s t a r c h ( t o p ) .



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milium

Lit

1

ILL F i g u r e 6:

V a r i e t a l d i f f e r e n c e s between w h i t e ( t o p ) and r e d (bottom) grape j u i c e s .


151


152


F i g u r e 7:

Comparison o f r o a s t e d peanuts ( t o p ) and peanut b u t t e r (bottom) made from same l o t o f raw peanuts.


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Literature 1. 2. 3. 4. 5.


6.

7.

8. 9. 10. 11. 12. 13. 14.

15.

16. 17.

18.


Cited

Nawar, W.W., and Fagerson, I.S.; Anal. Chem., 1960, 32, 1534 Hornstein, I., and Crowe, P.F.; Anal. Chem., 1962, 34, 1354 Morgan, M.E., and Day, E.A.; J. Dairy Sci., 1965, 48, 1382 Bellar, T.A.; Lichtenberg, J.J.; Jour.AWWA 1974,66,739. Schamp, N.; Dirinck, P.; in "Chemistry of Foods and Beverages: Recent Developments"; Charalambous, G.; Inglett, G., Eds.; Academic; New York, 1982; p25. Keenaghan, J.; Meyers, M.C.; "Analysis of Volatile Organics in Foods and Beverages by Headspace Concentration-GC/MS"; presented at the 34th Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy; Atlantic City, N.J.; March, 1984. Westendorf, R.G.; "Trace Analysis of Volatile Organic Compounds in Foods by Dynamic Headspace Gas Chromatography"; presented at the 35th Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy; Atlantic City, N.J.; March, 1984. Scholz, R.G.; Ptak, L.R.; J.Am.Oil Chem. Soc. 1966,43,596. Jackson, H.W.; Giacherio, D.J.; J.Am.Oil Chem. Soc. 1977,54,458-460. Jackson, H.W.; J.Am.Oil Chem. Soc. 1981,58,227,231. Dupuy, H.P.; Fore, S.P.; Goldblatt, L.; J.Am.Oil Chem. Soc. 1971,48,876. Dupuy, H.P.; Fore, S.P.; Goldblatt, L.; J.Am.Oil Chem. Soc. 1973,50,340. Selke, E.; J. Am.Oil Chem. Soc. 1970,47,393 Roberts, J.; "Semiautomated Dynamic Headspace Analysis of Vegetable Oil Volatiles"; presented at the 74th Meeting of the American Oil Chemists Society; Chicago, IL; May, 1983. Capillary Column Use in Purge and Trap Gas Chromatography II, Use of the Model 1000 Capillary Interface"; Application Note B021684; Tekmar Company, Cincinnati, OH. "Plastic Sample Preparation"; Application Note B081882; Tekmar Company, Cincinnati, OH. Westendorf, R.G.; "Optimization of Parameters for Purge and Trap Gas Chromatography"; presented at the 32nd Pittsburgh Conference on Analytical Chemistry and applied Spectroscopy; Atlantic City, NJ; March, 1981. "Purge and Trap Analysis Using a Photoionization Detector: Removal of Water Interference"; Application Note B042281; Tekmar Company, Cincinnati, OH.

RECEIVED June 24, 1985


Characterization and Measurement of Flavor Compounds - American

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