2014 Volume 20 Issue 1 Pages 109-113
Aroma extract dilution analysis (AEDA) was performed on an extract prepared from chicken soup stock and 9 aroma-active compounds were selected. On the basis of high flavor dilution (FD) factors in combination with the results of the identification experiments, methylpyrazine, 2-ethyl-4-methylthiazole, 3-(methylthio)propanal, and (E,E)-2,4-decadienal were suggested as primary aroma compounds of chicken soup stock. Recombination and omission experiments of the identified aroma-active compounds in taste-reconstituted chicken soup stock revealed the main flavor profile of each compound as “roast”, “roast meaty”, “boiled meaty”, and “fatty”, respectively. A comparison of the overall flavor of the recombined mixture and the chicken soup stock revealed a high similarity, suggesting that these four compounds are important contributors to the aroma of chicken soup stock.
During food intake, flavor perception results from simultaneous activation of the gustatory, olfactory, and trigeminal sensation systems. Especially, olfactory stimulation through the retronasal pathway greatly contributes to the flavor perception of foods (Shephard, 2006; Verhagen & Engelen, 2006).
The characteristic aroma of meat products greatly contributes to their overall acceptance (Shahidi, 1994). It is known that volatile compounds determine the aroma attributes and contribute most to the characteristic of meat aroma (Mottram, 1998). However, a limited number of volatile compounds actually contribute to the overall food aroma (Schieberle, 1995).
Whereas the aroma compounds of cooked meat have been widely investigated using gas chromatography-olfactometry (GC-O). (Farmer & Patterson, 1991; Kerscher & Grosch, 1997; Kerscher & Grosch, 2000; Sollner & Schieberle, 2009; Christlbauer & Schieberle, 2009; Carrapiso et al., 2002a; Carrapiso et al., 2002b), there is a lack of research about the aroma-active compounds of chicken soup stock.
Due to its desirable meaty flavor profile, chicken soup stock is commonly used as a base for savory dishes all over the world. The aroma compounds from chicken soup stock have been investigated (Wilson & Katz, 1972), but little research has been conducted on the aroma-active compounds of chicken soup stock. The aroma-active compounds of cooked chicken have been investigated and ranked according to their relative aroma potency in relation to cooked chicken flavor (Gasser & Grosch, 1990; Kerler & Grosch, 1997; Farkas et al., 1997). However, in those studies, the volatiles were isolated from raw chicken meat or pressure-cooked meat by simultaneous distillation/extraction, or from boiled chicken meat by direct extraction. To our knowledge, no study published in the available literature investigated aroma-active compounds of chicken soup stock in which volatiles were isolated from chicken soup stock, not from chicken meat.
Therefore, the purpose of the present work was to identify the most aroma-active compounds in chicken soup stock and assess their contribution to the sensory characteristics.
Chemicals All chemicals were purchased from Sigma-Aldrich Japan Inc., Tokyo, Japan.
Preparation of chicken soup stock Whole chicken was purchased at a local market and cut into four pieces. The chicken was boiled for 2 min, followed by removal of the internal organs and scum. Pre-cooked chicken pieces (4000 g) were boiled in 6500 g of tap water for 4 h at low heat, which was then filtered to obtain clear chicken soup stock. The stock was stored at −21°C until use.
Preparation of the volatile fraction of chicken soup stock Chicken soup stock (20 mL) was extracted by steam distillation. Extraction was carried out in a boiling water bath for 30 min under normal pressure until the distillate became 100 mL. Cyclopentanol was added as an internal standard (1 ppm in chicken soup stock). The distillate was extracted with 100 mL of diethyl ether three times. The organic layer was combined and dried over anhydrous sodium sulfate. The extract was concentrated to 150 µl under a gentle stream of N2 prior to analysis.
Gas Chromatography-Mass Spectrometry (GC-MS) The volatile compounds of chicken soup stock were analyzed using an Agilent 6890 gas chromatograph and an Agilent 5973 mass spectrometer (Agilent Technologies Inc., Japan). One microliter of extract was injected into the TC-Wax column (60 m length × 0.25 mm i.d., 0.25 mm film thickness; GL Sciences Inc., Japan). Helium was used as the carrier gas at a constant linear velocity of 30 cm/s. Oven temperature was programmed from 40 to 220°C at a rate of 4°C/min, with an initial hold time of 5 min and a final hold time of 10 min. GC-MS conditions were as follows: splitless mode; injector temperature, 220°C; valve delay, 60 s; scan range, 30-300 amu; scan rate, 2.2 scans/s.
Aroma Extract Dilution Analysis (AEDA) The GC-O system consisted of a Gerstel ODP2 (Gerstel K.K., Japan). One microliter of the ether extract of chicken stock was injected into a TC-WAX column (60 m length × 0.25 mm i.d., 0.25 µm film thickness; GL Sciences Inc.). GC conditions were the same as for GC-MS described above. The FD factor for a compound was defined as the ratio of its concentration in the initial extract to its concentration in the most dilute extract in which aroma was detected by GC-O. The FD chromatogram presented herein was the average of three expert panelists.
Compound identification and quantification Compounds were identified based on comparisons of GC retention indices (RI), mass spectra to the Wiley mass spectral database (John Wiley and Sons, Inc.) and aroma properties of unknowns with those of authentic standard compounds. The quantification of identified compounds in chicken soup stock was carried out using the peak area ratio of each compound to cyclopentanol. Peak area ratio was corrected with response factors of each compound, and response factors were calculated from the intensity ratio of each authentic compound to cyclopentanol using peak area in the total ion chromatogram.
Sensory comparison of reconstituted aroma mixture with chicken stock Panel: The panel was composed of three assessors trained to describe and recognize the aroma qualities of chicken soup stock. Six aroma descriptors, given in parentheses, which show the characteristic aroma of chicken soup stock were selected by comparison between chicken soup stock and taste-reconstituted chicken stock (t-Rec) (Table 1): fatty flavor (fat or oily flavor at top to middle); boiled meaty flavor (boiled meat-like flavor at middle to after); roast meaty flavor (roast meat-like flavor at top to middle); roast flavor (roast flavor at top to middle); animalic flavor (raw meat flavor at middle to after); overall similarity (overall similarity to chicken stock).
| no. | compound name | concn(µmol/L) |
|---|---|---|
| 1 | L-Leucine | 440 |
| 2 | L-Tyrosine | 220 |
| 3 | L-Isoleucine | 460 |
| 4 | L-Tryptophan | 50 |
| 5 | L-Lysine hydrochloride | 2470 |
| 6 | L-Valine | 190 |
| 7 | L-Phenylalanine | 270 |
| 8 | L-Arginine | 520 |
| 9 | L-Histidine | 850 |
| 10 | Taurine | 6600 |
| 11 | L-Glutamic acid | 1830 |
| 12 | Sodium aspartate monohydrate | 640 |
| 13 | L-Glutamine | 580 |
| 14 | Succinate | 1300 |
| 15 | 5′-GMP2Na·7.5H2O | 20 |
| 16 | 5′-IMP2Na·7H2O | 350 |
| 17 | NaCl | 7020 |
| 18 | NH4Cl | 8500 |
| 19 | CaCl2·2H2O | 140 |
| 20 | MgCl2·6H2O | 400 |
| 21 | K2HPO4 | 15340 |
| 22 | Glucose | 27 |
| 23 | DL-Alanine | 760 |
| 24 | Glycine | 600 |
| 25 | L-Methinonine | 170 |
| 26 | L-Proline | 190 |
| 27 | L-Serine | 560 |
| 28 | L-Threonine | 300 |
| 29 | L-Lactate | 22600 |
| 30 | Citric acid | 140 |
| 31 | Sodium acetate | 300 |
| 32 | L-Carnosine | 2920 |
Samples: Taste-reconstituted chicken soup stock (t-Rec) was prepared based on analysis data in the literature. (Dunkel & Hofmann, 2009) (Table 1). A complete mixture of seven or four aroma compounds (CM7 or CM4, respectively) in which the FD factor was more than 16 or 64 was prepared by adding all seven or four aroma compounds to t-Rec at the same concentration as that in chicken soup stock.
Profiling: The evaluation was performed as follows: The solutions were presented to the panelists in small cups, and they were asked to rate the intensities of the above six aroma qualities in comparison to t-Rec and chicken soup stock on a scale of 1.0 (not perceivable, = t-Rec), 2.0 (weak), 3.0 (medium), 4.0 (strong), 5.0 (very strong, = chicken soup stock). The results obtained were averaged for each aroma note and plotted on a spider web diagram.
Sensory profiling by Omission Experiments (OM) Panel: Sensory evaluation was conducted as described above.
Samples: Four model mixtures that omitted the following odorants from CM4 were prepared: OM1 omitted methylpyrazine; OM2 omitted 2-ethyl-4-methylthiazole; OM3 omitted 3-(methylthio) propanal; OM4 omitted (E,E)-2,4-decadienal. Each model mixture was presented to the sensory panel in comparison to CM4.
Profiling: The evaluation was performed as described above.
Identification of aroma-active compounds To evaluate whether the overall aroma can be isolated by steam distillation, one small drop of the diethyl ether extract obtained was evaluated on a blotter card by the three trained panels. The aroma of the extract immediately evoked the characteristic aroma of chicken soup stock, thereby confirming that the key aroma compounds were successfully extracted/distilled by steam distillation. The heat of steam distillation seems to have little influence on the aroma of the extract, as the influence of 30 min of boiling via steam distillation is minimal compared to that of 4 h boiling via cooking chicken soup stock.
After isolation of chicken stock volatiles, the most aroma-active compounds in the extracts were identified by means of AEDA.
In chicken soup stock, nine aroma-active compounds were identified in which the FD factor was more than 16 (Fig. 1). Among these, the following seven compounds were identified and quantified: methylpyrazine, 2-ethyl-4-methylthiazole, 3-(methylthio)propanal, 2-ethyl-1-hexanol, 3-methyl-2-cyclopentenone, 3-methylbutanoic acid and (E,E)-2,4-decadienal (Table 2).
Flavor dilution (FD) chromatogram obtained by application of the aroma extract dilution analysis (AEDA) on an extract of chicken soup stock.
| no. | aroma compounda | aroma qualityb | RIc | FD factord | concentration (ppb)e |
|---|---|---|---|---|---|
| 1 | Methylpyrazine | burnt, roast | 1195 | 128 | 0.38 |
| 2 | 2-Ethyl-4-methylthiazole | roast, nutty | 1267 | 64 | 0.28 |
| 3 | 3-(Methylthio)propanal | cooked potato-like, green | 1396 | 128 | 3.79 |
| 4 | 2-Ethyl-1-hexanol | minty | 1429 | 16 | 9.02 |
| 5 | 3-Methyl-2-cyclopentenone | burnt | 1462 | 16 | 3.38 |
| 6 | 3-Methylbutanoic acid | blue cheeze-like | 1603 | 32 | 14.7 |
| 7 | unknown | roast, nutty | 1685 | 32 | - |
| 8 | (E,E)-2,4-Decadienal | fatty, chicken skin-like | 1727 | 128 | 19.49 |
| 9 | unknown | sweety, caramel-like | 1907 | 16 | - |
a Identification was performed by comparing the following criteria: retention index; aroma quality and aroma threshold perceived at the sniffing port; mass spectra with reference compounds. b Aroma property perceived at the sniffing port. c Retention index determined in comparison to a homologous series of n-alkanes. d Flavor dilution factor. e Concentration in chicken stock.
Comparison of reconstituted aroma mixture with chicken tock To confirm the sensory impact of the identified aroma compounds on chicken soup stock flavor, the complete mixture of seven aroma compounds (CM7) in which the FD factor was more than 16 (except two unknown compounds) and four odor compounds (CM4) in which the FD factor was more than 64 were compared with t-Rec and chicken soup stock by sensory panel. Compared to t-Rec, the sensory score of CM7 and CM4, especially “fatty flavor” (1.0→4.4) and “animalic flavor” (1.0→4.6), were found to be strongly increased and reached almost the intensity of 5.0 observed for chicken soup stock (Fig. 2). Additionally, there were no significant differences between the scores of CM7 and CM4 in all flavor descriptors. These data indicate that methylpyrazine, 2-ethyl-4-methylthiazole, 3-(methylthio)propanal and (E,E)-2,4-decadienal, in which the FD factor was more than 64, were the most aroma-active compounds in chicken soup stock. These compounds were successfully identified as key aroma compounds of chicken soup stock in this study.
Comparative flavor profile analysis of taste-reconstituted chicken soup stock (t-Rec) (gray), chicken soup stock (dashed), a complete mixture of 7 (CM7) (gray dot), and 4 (CM4) (black) odor compounds dissolved in t-Rec.
Sensory profiling of the identified compounds Omission experiments were performed to study sensory profiling of the individual identified compounds.
Compared to CM4, the intensities of several flavor descriptors of OM1 to OM4, especially OM3 and OM4, were found to be lower (Fig. 3). The intensities of “roast” (2.7) of OM1 were much lower compared to that of CM4 (4.0) (p < 0.001). This indicates that methylpyrazine contributes to the “roast” flavor profile in chicken soup stock. In the same way, the intensity of “roast meaty” (2.1) and “roast” (2.9) of OM2 were much lower compared to those of CM4 (3.9, 4.0) (p < 0.001), respectively. This indicates that 2-ethyl-4-methylthiazole contributes to “roast meaty” and “roast” flavor profiles. The intensity of “boiled meaty” (2.2) of OM3 was much lower compared to that of CM4 (4.2) (p < 0.001). 3-(Methylthio)propanal contributes to “boiled meaty” flavor profiles. Also, the intensity of “fatty” (2.1) and “animalic” (2.4) and “boiled meaty” (3.2) of OM4 were much lower compared to that of CM4 (4.4, 4.6, 4.2) (p < 0.001), respectively. (E,E)-2,4-decadienal contributes to “fatty” and “animalic” and “boiled meaty” flavor profiles.
Comparative flavor profile analysis of complete mixture (CM4) (black) consisting of 4 odorants dissolved in t-Rec and (a) OM1 (dashed) (Methylpyrazine omission), (b) OM2 (dashed) (2-Ethyl-4-methylthiazole omission), (c) OM3 (dashed) (3-(Methylthio)propanal omission), (d) OM4 (dashed) ((E,E)-2,4-Decadienal omission)(*p < 0.001).
On the basis of the data obtained, it can be concluded that methylpyrazine and 2-ethyl-4-methylthiazole contribute to “roast”, whereas 2-ethyl-4-methylthiazole contributes to “roast meaty” in addition to “roast” flavors. 3-(Methylthio)propanal and (E,E)-2,4-decadienal have similar flavor profiles of “boiled meaty” but (E,E)-2,4-decadienal exhibits “fatty” and “animalic” flavor in addition to “boiled meaty” flavor. Additionally, they are important contributors to each characteristic flavor of chicken soup stock. This is the first study to confirm the key aroma compounds of chicken soup stock using aroma recombination and omission experiments and to investigate the roles of each aroma compound in chicken soup stock. However, the aroma-active compounds from cooked chicken have been investigated and ranked according to their relative aroma potency in relation to cooked chicken flavor (Gasser & Grosch, 1990; Kerler & Grosch, 1997; Farkas et al., 1997). In those studies, the volatiles were isolated from boiled chicken meat or pressure-cooked meat by simultaneous distillation/extraction, or from boiled chicken by direct extraction. A number of differences in key odorants between chicken meat and soup stock were expected in this study; methylpyrazine and 2-ethyl-4-methylthiazole were found as primary odorants of chicken soup stock for the first time. These odorants might be formed by reactions between aqueous components during the cooking of chicken soup stock, and further study is necessary to confirm the mechanisms responsible for their generation, as well as differences in key odorants of chicken products.
