The Importance of the Vacuum Headspace Method for the Analysis

Chapter 4

The Importance of the Vacuum Headspace Method for the Analysis of Fruit Flavors 1

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Downloaded by STANFORD UNIV GREEN LIBR on September 29, 2012 | http://pubs.acs.org Publication Date: September 10, 1998 | doi: 10.1021/bk-1998-0705.ch004

Matthias Güntert , Gerhard Krammer , Horst Sommer , and Peter Werkhoff

1Flavor Division and2CorporateResearch, Haarmann and Reimer GmbH, 37603 Holzminden, Germany

The Vacuum Headspace Method is a very gentle work-up procedure especially suited for the analysis of fruits. Comparison of the Simultaneous Distillation-Extraction Method, the Dynamic Headspace Method, and the Vacuum Headspace Method shows clearly the superior performance of the latter with respect to the sensory impression of the resulting extracts. Sensory evaluation of the vacuum headspace extracts of various fruits (e.g., passion fruit, strawberry, pear, peach, raspberry, cherry) leads to very fruit-typical descriptions. Consequently, the qualitative and quantitative flavor patterns of the analyzed fruits represent the genuine fruit flavors very well. The comparison of the different work-up procedures is explained in detail. Moreover, the volatile constituents of various analyzed fruits are shown and discussed. The importance of individual flavor compounds for the respective fruit flavor is indicated. It is explained how these results can serve as the basis for the composition of a new type offresh-fruitflavors. The analysis of food flavors has been dramatically improved over the past 30 years, mainly through the invention of new work-up procedures, modern separation techniques (high resolution gas chromatography on fused silica capillaries, high performance liquid chromatography), and more sensitive spectroscopic methods (nuclear magnetic resonance, mass spectrometry, fourier transform infrared spectroscopy) some of them being available as hyphenated techniques (HRGC-MS, HRGC-FTIR). This continuous development has, over the years, led to results and publications about virtually every food flavor. Thousands of flavor compounds have been identified in the different foods and are collected in libraries (7). Though there have always been groups that have worked this way, a significant trend in the nineties is for flavor research to focus on the few volatiles of a food flavor that are important character impact compounds and that dominate the smell and taste

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©1998 American Chemical Society In Flavor Analysis; Mussinan, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1998.


39 of a food flavor. In order to identify these active flavor compounds and moreover the right qualitative and quantitative compositional picture, the applied work-up procedures have to be selected and screened very carefully. Only if the extract, produced by the applied work-up procedure, shows the sensory properties of the flavor from the analyzed food are the chances good that the analytical results represent a true picture. Flavor analysis of non-thermally treated foods, which are eaten in the raw state (e.g., fruits), needs to be done in a very gentle way. In order to avoid cooked notes and artifacts, fruits have to be worked-up in a way that only the genuine flavor compounds in the right proportions are picked up. Because of the importance of the work-up conditions, we have carried out a comparison between three of the most commonly used procedures: the simultaneous steam distillation-extraction according to LikensNickerson (LN), the dynamic headspace (DHS), and the vacuum headspace (VHS). Our goal was to produce an extract that resembles the sensory quality of a freshly picked ripe fruit. Therefore, fruits were first subjected to very thorough sensory evaluation before the application of the work-up procedures. Subsequently, sensory evaluation was run on the extracts in addition to GC and GC/MS analyses. In this way we have elaborated the analysis of numerous fruits. Results of passion fruit, strawberry, peach, pear, raspberry, and cherry are described here. The analytical results were used as a tool to formulate synthetic (natureidentical) and natural fruit flavors. The main feature of these flavors is their typical strong smell and taste that resembled the freshly picked fruits. Experimental Procedure Investigated Fruits. Sensory evaluation, by our flavorists, was run on all fruits. The target was to analyze the fruits at the height of ripeness in their most typical state of taste. The fruits analyzed are listed below. • Yellow passion fruit (Passifloraceae, Passiflora edulis var. flavicarpa) harvested in 1995 in Columbia (South America) • Strawberry (Rosaceae, Fragaria x ananassa var. Senga sengand) harvested in 1995 in the environment of Holzminden (Germany) • Raspberry (Rosaceae, Rubus idaeus) harvested in 1993 • Pear (Rosaceae, Pints communis var. Bartlett, Williams Christ) harvested in 1995 • White peach (Rosaceae, Prunus persica var. Maria Bianca) harvested in 1995 • Sweet Cherry (Rosaceae, Prunus avium) harvested in 1993 • Sour Cherry (Rosaceae, Prunus cerasus var. "Schattenmorelle ") harvested in 1993 Sample Preparation. For the analysis of the fruits four different work-up procedures were employed. These were the Simultaneous Distillation-Extraction Method at ambient pressure (SDE) and the Simultaneous Distillation-Extraction Method under vacuum (SDEV), the Dynamic Headspace Method (DHS), and the Vacuum Headspace Method (VHS). Drawings of the physical principles of these methods are shown in Figure 1. Simultaneous Distillation-Extraction Method at Ambient Pressure (SDE) and the Simultaneous Distillation-Extraction Method under Vacuum (SDEV). This method is one of the most widely used sample preparation procedures (2-6). Water steam volatile flavor compounds from the fruits (between 0.5 kg and 3 kg

In Flavor Analysis; Mussinan, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1998.


Figure 1: Isolation Procedures for the Flavor Volatiles of Fruits



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chopped fruits or fruit pulp depending on the flavor strength) are continuously distilled off and simultaneously extracted from the gas phase using an organic solvent (200 mL pentane/diethyl ether 1:1). The procedure under reduced pressure (80 mbar/50 °C) was conducted in the same way by using methyl ter/-butyl ether as an extraction solvent. In this way, the flavor compounds are enriched in the organic phase. Afterwards, the extracts are gently concentrated, using a Vigreux distillation column, to a final volume of about 0.2 mL. Dynamic Headspace Method (DHS). The physical principle of this sample preparation method is to strip the volatile flavor compounds from the headspace above the foods by a stream of air or inert gas (7,8). This flavor isolation procedure has been used successfully by Buttery and co-workers (9,10) for the analysis of tomato paste and tortillas .The method has also been used quite often for the analysis of blossoms. Its big advantage is it non-destructive and can be used to isolate volatiles from living nature. In a recent publication Roman Kaiser described the evolution of the analysis of blossom scents in nature (77). DHS has also been applied for investigation of fruits on the branch or vine (12-14). In our experiments, we used between 1 kg and 3 kg chopped fruits or fruit pulp (depending on the flavor strength), worked at ambient temperature, and used purified (passed through a molecular sieve) helium as a purge gas (150 rnL/rnin flow). The •'extracted" flavor compounds were adsorbed onto a polymer carrier material (in our case a glass tube packed with 3.1 g of Tenax TA) and can then be desorbed in different ways (thermal desorption directly into the GC, thermal desorption into a glass desorption trap, desorption with an organic solvent). In our experiments, we ran the adsorption from 5 h to 45 h depending on the fruits. The desorption was carried out with a mixture of pentane/diethyl ether 1:1 (150 mL). Afterwards, the combined extracts were gently concentrated, using a Vigreux distillation column, down to a volume of about 0.2 mL. Vacuum Headspace Method (VHS). This work-up procedure was first applied by various scientists for the investigation of blossom scents. The first publication appeared in 1986 by Daniel Joulain (75). Since then it has become an established method for analyzing blossom scents (16-20). More recently, the vacuum headspace technique has been used for flavor analysis of various fruits, such as strawberries, peaches, and cupua$u (21-24). Very recently, Tarantilis and Polissiou reported on the isolation and identification of the flavor compounds of saffron using the vacuum headspace method (25). It also has to be mentioned, a similar method for isolating flavor volatiles was described in the literature many years ago under the name "high vacuum distillation" (26-28). The vacuum headspace method is, in principle, a vacuum steam distillation that takes place with the water of the respective fruit. The volatile flavor compounds distill off and are condensed in the cooled traps. The contents of the traps are then combined and are cold extracted with an organic solvent. In our experiments we used between 0.5 kg and 5 kg chopped fruits or fruit pulp (depending on the flavor strength) and ran the vacuum headspace from 4 h to 8 h, depending on the fruits (approximately 1-10 mbar). The three cooling traps were cooled with ice-water, dry ice-acetone, and liquid nitrogen. The liquid-liquid extraction was carried out with a mixture of pentane/diethyl ether 1:1 in a rotatory extractor. Afterwards, the extracts were gently concentrated, using a Vigreux distillation column, down to a volume of about 0.2 mL.


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The aroma of the flavor concentrates, obtained from fruits by the different isolation methods, were compared with the aroma of fresh fruits by our flavorists. The results are summarized in Table 1. In order to study the influence of the four different isolation methods on the chemical composition of the fruit volatiles all solvent extracts were subsequently analyzed using completely identical chromatographic and spectroscopic conditions. Instrumental Analysis. Instrumentation (capillary gas chromatography, spectro scopy) as well as analytical and preparative conditions were described in previous publications (29,30) and in a very recent paper from our work group where we described the flavor chemistry of yellow passion fruits in detail (Werkhoff, P.; Guntert, M ; Krammer, G.; Sommer, H . ; Kaulen, J. JAgric. Food Chem., accepted for publication) Component Identification. Flavor compounds from the fruits investigated were identified by comparison of the compound's mass spectrum and Kovats indices with those of a reference standard. In some cases reference compounds were synthesized in our laboratory. In all cases the respective structures were confirmed by N M R , IR, and M S spectroscopy. Gas Chromatography-Olfactometry. G C with simultaneous FID and odor port evaluation was carried out using a Carlo Erba Type 5360 Mega Series gas chromatograph. Separations were performed using a 60m x 0.32 mm (ID) capillary column coated with DB-1 (df = 1 um) and D B - W A X (df = 0.5 um) stationary phases. The flow rate of the helium carrier gas was 3-4 mL/min. The column effluent was split 1:50 with a glass-cap-cross (Seekamp, Achim, Germany). The temperature program used was 60 °C-220 °C at 3 °C/min. Injections were made in the on-column as well as in the split/splitless mode. The injector temperature was 220 ° C and the detector temperature was 250 °C Results and Discussion Analysis of the Fruits. The main target of this study was to find the sample preparation method that gives an extract with the sensory impression most typical of the ripe, freshly picked fruit. Subsequently, the qualitative and quantitative flavor patterns, identified in this study, should be the basis for formulating a range of new and unique fruit flavors. Therefore, comparison of the different work-up procedures applied to the various fruits was very important. A second aspect of this study was to compare the dynamic headspace method with the vacuum headspace method. We wanted to determine whether the dynamic headspace method, which has been applied to blossoms, could be successfully applied to fruits. Several reviews on sample preparation and isolation procedures have recently been published (31-35). The influence of these different techniques on the resulting flavor extracts was discussed by several scientists (36-40). These publications all demonstrate that the isolation procedure is of critical importance for the sensory quality of the resulting flavor extracts. In our study, the results of the different work-up procedures applied on the



estery,fruity,green, juicy, pear skin, typical

fruity, green, woody, ionone note, violet note, sweet, typical

acidic, cinnamon note, sweet, cooked note

fruity, hay note, juicy, typical

Pear

Raspberry

Sweet cherry

Sour cherry

-

-

-

fruity, fatty, rancid, weak

estery,fruity,sweet, amine-like, weak

fruity, green, acidic, sweet, earthy, ripe, typical

Strawberry

herbaceous, tropical, weak

lactoney,fruity,green, white lactoney,fruity,juicy, overripe,fruitflesh, peach skin very volatile typical

hi

White peach

t)

estery, tropical fruit, green, sulfury, juicy, fresh, typical

J

Yellow passion fruit

(VHS^IlsSj^

-

-

-

-

-

green, estery, tatty, sweaty, juicy, typical

overripe, cooked

-

green, cinnamon note, cooked, artificial

green,fruity,lactoney, cooked, canned

green, buttery, weedy, marmelade, cooked

lactoney,fruity,tea note, cooked note

sulfury, fruity, tropical fruit, sulfury, tropical, sweet, burnt, acidic, somewhat somewhat typical typical

Table I: Sensory Evaluation of the Fruit Concentrates Obtained by the Various Flavor Isolation Procedures Simultaneous D istmitioi^ Vacuum,Hea

The Importance of the Vacuum Headspace Method for the Analysis

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