Olfactometric characterization of aroma active compounds in fermented fish paste in comparison with fish sauce, fermented soy paste and sauce products

Food Research International 43 (2010) 1027–1040

Contents lists available at ScienceDirect

Food Research International journal homepage: www.elsevier.com/locate/foodres

Olfactometric characterization of aroma active compounds in fermented fish paste in comparison with fish sauce, fermented soy paste and sauce products Anupam Giri a, Kazufumi Osako a, Akira Okamoto b, Toshiaki Ohshima a,* a b

Department of Food Science and Technology, Tokyo University of Marine Science and Technology, Tokyo 108-8477, Japan Nagasaki Prefectural Institute of Fisheries, Taira, Nagasaki 851-2213, Japan

a r t i c l e

i n f o

Article history: Received 11 November 2009 Accepted 19 January 2010

Keywords: Aroma active compounds Olfactometry Organoleptic Fish miso Soy miso Fish sauce Soy sauce

a b s t r a c t Aroma active compounds in commercial fermented fish meat paste product (fish miso), fermented soy paste (soy miso), fish sauce and soy sauces were characterized by using a dynamic headspace method for volatile isolation and GC olfactometry for odor perception. A total of 123 volatile compounds consisting mainly of aldehydes, alcohols, esters, ketones, furans, sulfur and nitrogen-containing compounds, aromatics and acids were consistently identified. A major 16 odor-active compounds were distinguished to contribute as key aroma compounds for the miso and sauce products. Olfactometric and sensory findings clearly differentiated miso products with caramelic, fruity aroma notes, whereas fish sauce products were characterized by ammoniacal, fishy, nutty and cheesy odor note. Soy sauce products, however, were dominated by nutty and cheese aroma. Use of koji for fish miso production was found effective to enhance sweet aroma to the product with a reduction of nutty, meaty and rancid nuance. Principal component analysis employed for statistical interpretations clearly elucidated the relationship among different types of fermented products. Ó 2010 Elsevier Ltd. All rights reserved.

1. Introduction Soy sauce, fermented soy paste (soy miso) and fish sauces are widely consumed as seasoning liquid or pastes mostly in Southeast Asian countries such as Japan, Korea, China, Vietnam, Cambodia and Thailand. Considering the importance of aroma for those products several researches have been carried out on their volatile profile. The characteristic flavor–aroma formation in the soy sauce depends on the manner of production employed, as well as raw materials and strains of microorganism used (Saisithi, Kasermsarn, & Dollar, 1966). Most of the studies on the volatile compounds in traditional soy sauces which were produced from the regions including Japan, Korea, China and Indonesia, have been carried out (Apriyantono, Husain, Lie, Jodoamidjojo, & Puspitasari-Nienaber, 1999; Kim et al., 1996; Kobayashi & Sugawara, 1999; Seo et al., 1996). However, the comparative study on olfactometric and sensory analysis on aroma profile of soy sauce still needs to be well conducted. The volatile composition of soy miso aroma has been investigated by several research groups (Itoh & Ebine, 1970; Iwabuchi & Shibasaki, 1973; Yasuhira, Itoga, & Mochizuki, 1974). The components reported so far, however, were limited to volatile acids and neutral compounds such as alcohols, esters and carbonyl compounds. In spite of the possible importance of the major * Corresponding author. Tel.: +81 3 5463 0613. E-mail address: [email protected] (T. Ohshima). 0963-9969/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.foodres.2010.01.012

components in miso flavor, there has been a scarcity of investigations in this field. Fish sauce plays a major role in the utilization of pelagic fish in Southeast Asia. It has been reported that the aroma of fish sauce is composed of a blend of ammoniacal, cheesy and meaty notes. A variety of amines, ammonia and other nitrogen-containing compounds have been attributed to the ammoniacal note as described by Dougan and Howard (1975). Van-Chom (1958) considered volatile fatty acids such as formic, acetic and n-butanoic acids as contributory factors in the aroma of fish sauce. Investigation of Nouc-mam, a Vietnamese fish sauce, revealed that it contains a large amount of methyl ketones which probably accounts for the cheesy odor. n-Butanoic acid was found to be the most abundant acid in fish sauce whereas Mclver, Brooks and Reineccius (1982) reported that acetic acid is a prominent acid in Nampla, a Thiland fish sauce. Because of their fishy flavor and characteristic unpleasant odor due to volatile amines such as trimethylamine, fish sauces have limited consumer acceptability and are usually used as ‘‘hidden flavors” in Japan (Fukami, Funatsu, Kawasaki, & Watabe, 2004). On the other hand, preliminary organoleptic study on fish miso, a Japanese fermented fish meat paste, which was comparably a newer product than other traditional fermented fish products described above, revealed that though they are produced from fish meat origin, have overall sweet aroma in contrast to those of fish sauce products. Very few works related to its nutritional and taste aspects are available (Giri, Osako, & Ohshima, 2009). However, no

1028

A. Giri et al. / Food Research International 43 (2010) 1027–1040

literature has been available on its olfactometric characterization. Lack of knowledge regarding the distinct clarification of odor-active compounds among the fermented fish miso, soy miso, fish sauce and soy sauce often creates confusion about their aroma profile. To characterize and investigate the aroma profile of fish miso in comparison with other fermented fish and soy fermented products, commercial fish miso prepared with koji (rice malt inoculated with Aspergillus oryzae) and without koji are considered in the present study. Although, the varieties of commercial fermented fish and soy products differ in the material ratio, ageing time, pasteurization treatment and soy bean or fish cooking condition, it could be expected that a comparison of the results for different types of products would give clear indication of each production process. The purpose of selecting wide ranges of fermented paste and liquid products from fish meat and soybean was to get a comprehensive as well as comparative view of aroma profile of diverse traditional and newly developed seasonings for Asian food and cooking. 2. Materials and methods

and 4.5 in. length, Supelco, Bellefonte, PA) filled with 175 mg of 60/88 mesh Tenax TA (Supelco) fitted to a bigger neck by a connector. Dry nitrogen purge was carried out for 10 min at a flow rate of 50 mL/min in a direction opposite to the isolation to remove the residual moisture. The Tenax TA trap was preconditioned at 300 °C for 30 min with nitrogen gas at 50 mL/min by a Model 10 tube conditioner (Dynatherm Analytical instruments, Inc., Kelton, PA) before each sampling. 2.4. Thermal desorption of volatiles to gas chromatography Dry purged tube with isolated volatiles was placed onto a Model ACEM 900-FF/EPC automated thermal desorption system (CDS Analytical, Inc., Oxford, PA), devised with an additional focusing trap with a low internal volume. Volatiles trapped in dry purged Tenax TA tube were desorbed for 3 min at a temperature of 300 °C and were absorbed by focusing trap. Subsequent rapid increase in focusing trap temperature to 300 °C in 15 s and subsequent hold for 5 min allowed efficient injection of the collected volatiles onto a capillary column in a narrow-band plug through a short path transfer line (0.5 m inert column kept at 250 °C).

2.1. Materials 2.5. Conditions of gas chromatography–olfactometry Ten different kind of commercial fermented fish miso, soy miso, fish sauce and soy sauce were used for the present study. Details of them are presented in Table 1. Products were purchased from local markets in Tokyo, Japan. Miso and sauce products were stored at 80 °C and 5 °C respectively to minimize the changes of aroma profile throughout the experimental period.

All the volatile standards used for identification and other analysis purposes were of GC-analytical grade and were purchased from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan).

Analysis of volatiles were carried out on a Shimadzu model 14A gas chromatograph (Kyoto, Japan) equipped with a SupelcowaxTM – 10 fused silica capillary polar column (0.32 mm i.d.  60 m, 0.25 lm in film thickness). The outlet of the column was channeled into a flame ionization detector and an olfactometry port (OP275, GL Science, Tokyo, Japan) at a ratio of 1:1. High purity helium was used as a carrier gas at a flow rate of 1.5 mL/min with a column inlet pressure of 3 kg/cm2. The column temperature was held at 40 °C for 3 min and subsequently programmed to 200 °C at a rate of 3 °C/min. Injector and detector temperatures were set at 250 °C.

2.3. Isolation of volatiles using Tenax TA trap

2.6. Conditions of gas chromatography–mass spectrometry

Samples of 3 g with 0.01 mL of 1000 ppm 2,4,6-trimethylpyridine as an internal standard were taken into 500 mL dual-necked round bottom glass flask and was placed over hot water bath maintained at 75 °C. High purity nitrogen gas was passed through the smaller neck of the flask at a flow rate of 100 mL/min and headspace volatiles emanating from the sample were trapped for 30 min on a glass-fritted absorption trap (4 mm I.D., 6 mm O.D.

Analysis of mass spectrum of volatile compounds were carried out on a Hewlett Packard 5890 Series II gas chromatograph (SupelcowaxTM – 10 fused silica capillary polar column: 0.25 mm i.d.  60 m, 0.25 lm in film thickness; Supelco) equipped with a Tekmar 7000 headspace auto-sampler (Cincinnati, OH, USA) and an ion source from Automass (JEOL, Japan). The ionization energy, scan range and scan rate applied for the analysis were 70 eV,

2.2. Chemicals

Table 1 Details of commercial fermented fish miso, soy miso, fish sauce and soy sauce with major ingredients and producing country. Fish miso Fish miso (with koji)a (without koji)

Soy miso (light)b

Product type

Fermented fish paste

Fermented fish paste

Fermented Fermented soy paste soy paste

Color

Dark brown Anchovy, salt, koji

Dark brown

Light yellow Soy, rice koji, salt

Dark brown Soy, rice koji, salt

Nagata Shokuhin, Japan

Nagata Shokuhin, Japan

Takeya, Japan

Marukome, City Food Japan Co. Thailand

Major ingredients Producer name and country a b c d e

Anchovy, salt

Soy miso (dark)c

Koji, rice malt innoculated with A. oryza as fermentation starter. Light colored product. Dark colored product. 18 Months ripned product. Nine months ripened product.

Nampla Nampla Nouc-mam (premium)d (standard)e

Oyster sauce

Liquid fermented fish Dark brown

Liquid fermented fish Brown

Liquid fermented fish

Liquid Liquid fermented fermented soy fish Dark brown Light brown

Anchovy, salt, sugar

Anchovy, salt

Anchovy, salt

Youki, Thailand

Hung Thanh, Phu Quoc, Vietnam

Yellow

Oyster, sugar, salt, starch Rainbow Shokuhin, Japan

Soy sauce (light)b

Soy, flour, salt, sugar

Soy sauce (dark)c Liquid fermented soy Dark brown Soy, flour, salt, sugar

Guangdong Guangdong Foodstuffs Corp. Foodstuffs Corp. China China

1029

A. Giri et al. / Food Research International 43 (2010) 1027–1040 Table 2 Identification, odor description, threshold, linerity and precision of volatiles detected in different commercial miso and sauce samples. Peak no

Volatile compounds

RIa

Identificationb

Odor description

Threshold (lg/L)

Standard curve (r)

Validation range

RSD%

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17

Trimethylamine Acetaldehyde Propanal 2-Propanone 2-Methylpropanal Ethyl acetate 2-Butanone 2-Methylbutanal 3-Methylbutanal 2-Propanol Ethanol 2-Ethylfuran Ethyl isobutyrate 2-Pentanone Pentanal 2,3-Butanedione 2-Methylpropyl acetate 2,3-Pentanedione Ethyl butanoate 2-Butenal 1-Propanol Ethyl-2methylbutanoate Ethy l-3 methylbutanoate Dimethyl disulfide Butyl acetate Hexanal 3 -Methyl-2butanol 2-Methylpropanol 2-Methyl-2butenal (E) Isobutyl isobutanoate Ethyl benzene 3-Pentanol Isoamyl acetate 2-n -Butylfuran p-Xylene Ethyl pentanoate 1-Butanol 1-Penten-3-ol o-Xylene Heptanal Cymene 3-Hexanol Propyl benzene 2-Methyl-1butanol 3 -Methyl-1 butanol 2-Hexenal(Z) 2-Hexanol 2-Pentylfuran Ethyl hexanoate 4-Heptenal Methyl pyrazine 3-Octanone 1-Pentanol 3-Methylbutyl butanoate p-Cymene 2-Methylbutyl 2methylbutanoate 2-Octanone Octanal Isoamyl isovalerate 3-Methyl-1 -pentanol

679 692 803 810 814 890 903 912 915 922 924 956 970 981 982 985 1026

MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS,

RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI,

Std, Std, Std, Std Std, Std, Std Std, Std, Std Std Std, Std, Std Std Std, Std,

Rotten fish, ammoniacal Apple skin, fruity Solvent, pungent Glue, fruity Nutty, malty, burnt, green Fruity, buttery, orange Ethereal, cheese, chemical Nutty, almond, burnt, malty Almond, nutty, herbaceous Ethereal, alcoholic Strong alcoholic Rubber, pungent, musty Strawberry, fruity, sweet Sweet, fruity, ketone like Pungent, almond like, rubber Creamy, caramel, butter scotch Banana, pear note, fruity

0.1 25.1 15.1 45000.3 1.5 5.0 35400.2 1.0 0.2 8500.8 950000.0 2.3 0.2 75005.9 2501.3 0.1 25.0

y = 0.8812x + 0.0007 (0.9823) y = 0.6879x 0.0013 (0.9908) y = 0.7802x 0.001 (0.9744) y = 0.0325x + 0.0001 (0.9901) y = 0.3705x 0.0009 (0.9758) y = 0.6991x 0.0011 (0.9921) y = 0.0241x + 0.0003 (0.9669) y = 0.8982x 0.0011 (0.9909) y = 1.0672x 0.0012 (0.9918) y = 0.6374x 0.0015 (0.9824) y = 0.5314x 0.0 02 (0.9924) y = 1.0504x 0.0011 (0.9931) y = 0.5803x 0.0006 (0.9973) y = 1.4749x 0.0022 (0.9939) y = 1.045x 0.0015 (0.9898) y = 0.0329x + 0.0001 (0.9986) y = 1.1822x 0.0018 (0.9931)

10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng

-1000 ng - 300 ng - 300 ng -1000 ng - 300 ng - 300 ng -1000 ng - 300 ng - 300 ng - 300 ng - 300 ng - 300 ng - 300 ng - 300 ng - 300 ng -1000 ng - 300 ng

2.123 1.560 1.871 1.590 1.990 1.487 1.980 1.697 2.135 1.540 1.123 2.111 1.111 1.896 1.456 0.987 1.458

1052 1054 1058 1064 1067

MS, MS, MS, MS, MS,

RI, RI, RI, RI, RI,

Std Std, O Std, O Std Std, O

Butter scotch, almond, fruity Apple, fruity, pear drops Pungent, ethereal Plastic, pungent, musty Sweet caramel, grape

5505.6 1.0 0.4 8505.7 0.2

y = 0.0448x + 0.0002 (0.9669) y = 0.342x 0.0008 (0.9956) y = 0.6821x 0.0007 (0.9982) y = 0.2812x 0.0002 (0.9864) y = 0.8176x 0.0008 (0.9953)

10 ng 10 ng 10 ng 10 ng 10 ng

-1000 ng - 300 ng - 300 ng -1000 ng - 300 ng

1.254 1.569 0.896 1.889 2.102

1071

MS, RI, Std, O

Cashew, apple, sweet fruity

0.2

y = 0.689x

10 ng - 300 ng

2.210

1081 1083 1091 1095

MS, MS, MS, MS,

Cooked cabbage, onion Pineapple note, banana, fruity Fishy, grassy, leafy, green Green apple, solvent

1.1 58.0 5.0 1259.9

y = 0.3913x y = 0.8986x y = 0.1895x y = 1.8776x

0.0011 0.0014 0.0007 0.0019

10 ng 10 ng 10 ng 10 ng

- 300 ng - 300 ng -1000 ng - 300 ng

1.589 1.469 1.458 1.998

1103 1115

MS, RI, Std MS, RI, Std, O

Solvent like, glue, leek, biller Cocoa, coffee like

6505.3 459.0

y = 2.1909x y = 1.6982x

0.0034 (0.9951) 0.0022 (0.9916)

10 ng - 300 ng 10 ng - 300 ng

1.456 1.002

1117

MS, RI, Std, O

Fruity, sweet

30.1

y = 1.1168x

0.0006 (0.9953)

10 ng - 300 ng

1.203

1126 1131 1133 1136 1139 1143 1169 1181 1191 1199 1209 1214 1215 1223

MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS, MS,

Ethereal, floral, sweet Green Banana, fresh, pear odor Noncharacteristics, weak Cold meat, oily Orange, fruity, mint Fragrant, solvent, pungent Burnt, meaty, grassy, green Geranium, oily, pungent Dry fish, green, rancid, bily Strong aromatic Alcoholic, solvent Moth-ball like odor, like toluene Fusel oil, ripe onion, malty

2205.3 4125.0 1.6 5.0 68.6 5.9 459.2 358.1 450.2 2.9 6.5 820.5 177.1 16.0

y = 0.5172x 0.0005 (0.9787) y = 1.8693x 0.0025 (0.9910) y = 2.9385x 0.0025 (0.9950) y = 1.3099x + 0.001 (0.9733) y = 1.1625x 0.0021 (0.9554) y = 2.5398x 0.0023 (0.9946) y = 2.1129x 0.0012 (0.9981) y = 2.2005x 0.0029 (0.9939) y = 1.6817x 0.0025 (0.9756) y = 0.1859x 0.0032 (0.9737) y = 1.027x 0.0002 (0.9988) y = 1.7385x 0.0025 (0.9931) y = 1.5522x 0.0001 (0.9722) y = 0.8985x 0.0011(0.9481)

10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng

300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng 300 ng

2.031 1.369 1.121 1.932 1.568 1.987 1.777 2.102 2.111 1.030 1.784 2.469 1.362 1.362

1226

MS, RI, Std, O

Rancid, pungent, balsamic

4.0

y = 2.4585x

10 ng - 300 ng

2.362

1235 1237 1238 1241 1255 1261 1265 1263 1272

MS, MS, MS, MS, MS, MS, MS, MS, MS,

Stink bug, bitter, almond Fatty, fruity, wine like Green bean like, pungent Fruity, strawberry, wine like Biscuit, creamy, fatty Fishy, nutty, ammoniacal Mashroom, harbal, resinous Green, pungent, fruity, wax Sweet, apricot, banana like

19.2 1508.2 5.9 2.3 4.2 56564.6 21.5 150.3 15.0

y = 1.0751x 0.0013 (0.9800) y = 0.5401x 0.0007 (0.9954) y = 0.2607x + 0.001 (0.9835) y = 2.787x 0.0026 (0.9986) y = 1.2125x 0.0015 (0.9812) y = 0.0457x 0.0001 (0.9012) y = 1.0908x 0.0022 (0.9818) y = 2.1786x + 0.0002 (0.9918) y = 1.157x 0.002 (0.9798)

10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng

- 300 ng - 300 ng - 300 ng - 300 ng - 300 ng -1000 ng - 300 ng - 300 ng - 300 ng

1.560 1.784 1.224 0.987 1.556 2.985 1.256 1.236 1.563

1274 1283

MS, RI, Std, O MS, RI, Std, O

Gasoline, spicy, harbal, lemon Berry note, fruity, apple

5.0 75.0

y = 0.0098x + 0.0009 (0.7869) y = 0.9719x 0.0027 (0.9692)

10 ng -1000 ng 10 ng - 300 ng

3.654 1.254

1295 1298 1303

MS, RI, Std, O MS, RI, Std, O MS, RI, Std, O

Soapy, floral, musty, cheese Orange peel, fatty, pungent Fruity, sweet

50.3 0.6 19.9

y = 1.1695x 0.0009 (0.9786) y = 0.8453x 0.0027 (0.9549) y = 0.366x + 0.0002 (0.9889)

10 ng - 300 ng 10 ng - 300 ng 10 ng - 300 ng

2.123 1.989 1.785

1318

MS, RI, Std, O

Winey, whisky, green note

7.6

y = 1.2212x

10 ng - 300 ng

1.236

18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

RI, RI, RI, RI,

RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI, RI,

RI, RI, RI, RI, RI, RI, RI, RI, RI,

O O O O O O O

O O

O O

Std, O Std, O Std, O Std

Std Std Std, Std, Std, Std, Std Std, Std Std, Std, Std Std Std,

Std, Std Std, Std, Std, Std Std, Std, Std,

O O O O O O O

O

O O O O O O O

0.0014 (0.9800) (0.9841) (0.9765) (0.9344) (0.9933)

0.0011 (0.9987)

0.0003 (0.9865)

-

(continued on next page)

1030

A. Giri et al. / Food Research International 43 (2010) 1027–1040

Table 2 (continued) Peak no

Volatile compounds

RIa

Identificationb

Odor description

Threshold (lg/L)

Standard curve (r)

Validation range

RSD%

61

2,6-Dimethyl pyrazine 2,4-Dimethyl pyrazine Ethyl pyrazine 2,3-Dimethyl pyrazine 2-Ethyl-1-butanol Cyclopentanol 2-Heptanol 2-Penten-1-ol(E) 3 -Methy l-3 buten-1 -ol Ethyl heptanoate 6-Methyl-5hepten 2-one 1-Hepten-3-ol Hexanol Phenyl ethyne 2,4,5-Trimethyl thiazole Dimethyl trisulfide 3-Octanol Nonanal 2-Hexen-1-ol(E) 2-Hexen-1-ol(Z) 2-Ethyl 3-methyl pyrazine 2,3,5-Trimethyl pyrazine 3-Ethyl- 2,3dimethyl pyrazine Ethyl octanoate Acetic acid 1-Octen-3-ol Heptanol 3-(Methylthio) propanal 1,3-Dimethyl 1Hpyrazole 2,4Heptadienal(EZ) 2-Furaldehyde Tetramethyl pyrazine 2-Ethyl hexanol 3,5-Diethyl 2methyl pyrazine 2,4-Heptadienal (E,E) 2-Acetylfuran 2-Nonanol Benzaldehyde 2-(Methylthio) ethanol 2,3Butanediol(levo) Octanol 2-Methyl propanoic acid Ethyl 3(methylthio) propanoate 3,5-Octadien-2one (EE) 5-Methylfurfural 2,3-Butanediol (meso) 2,6Nonadienal(EZ) 2,4Nonadienal(EZ) Benzene acetaldehyde

1319

MS, RI, Std, O

Rosted nuts, fried potato

157.6

y = 0.1584x + 0.0001 (0.9939)

10 ng -1000 ng

1.023

1320

MS, RI, Std

Chocolate, earthy, nutty

578.6

y = 0.0857x

0.0001 (0.9766)

10 ng -1000 ng

1.456

1321 1322

MS, RI, Std MS, RI, Std

Peanut, buttery, chocolate Green nutty, coffee, caramel

5550.5 2225.3

y = 0.0451x y = 0.0336x

0.0001 (0.9451) 0.0001 (0.9777)

10 ng -1000 ng 10 ng -1000 ng

2.120 1.568

1324 1327 1330 1337 1339

MS, MS, MS, MS, MS,

O O O O

Sweet, musty odor Pleasant, amyl alcohol like Mashroom, acrid, herb Green, plastic, rubber Herbaceous

75.2 125.1 65.2 89.2 547.1

y = 1.3187x 0.0015 (0.9983) y = 2.1904x 0.0035 (0.9828) y = 0.5261x + 0.0002 (0.9875) y = 1.3589x 0.0022 (0.9900) y = 1.2119x 0.0001 (0.9841)

10 ng 10 ng 10 ng 10 ng 10 ng

300 ng 300 ng 300 ng 300 ng 300 ng

1.002 1.563 1.446 1.889 1.456

1341 1345

MS, RI, Std, O MS, RI, Std, O

Banana, strawberry, fruity Sweet, fruity

2.0 68.0

y = 0.338x 0.0007 (0.9687) y = 0.2825x 0.001 (0.924)

10 ng -1000 ng 10 ng -1000 ng

1.779 2.365

1347 1367 1376 1389

MS,RI MS, RI, Std, O MS MS, RI, Std, O

Green, grassy, fatty, leafy

5.7

y = 1.0605x

0.0014 (0.9733)

10 ng - 300 ng

1.895

Cocoa, dark chocolate, green

45.0

y = 0.2474x

0.0013 (0.8482)

10 ng -1000 ng

2.999

1397

MS, RI, Std, O

Fishy, sulfury, cooked onion

0.0

y = 0.571x

10 ng -1000 ng

1.890

1402 1405 1417 1422 1434

MS, MS, MS, MS, MS,

Mashroom, herbaceous Green, tallowy, fatty, lavender Brandy nuance Walnut, medicinal, green, leaf Nutty, earthy, rosted, potato

35.0 1.1 232.0 359.4 0.4

y = 1.7598x y = 1.4646x y = 2.2742x y = 2.3725x

10 ng 10 ng 10 ng 10 ng

300 ng 300 ng 300 ng 300 ng

2.123 1.898 2.135 1.469

1438

MS, RI, Std, O

Bread, burnt, musty, peanut

350.1

y = 0.201x

10 ng -1000 ng

1.456

1440

MS, RI

1444 1451 1460 1465 1476

MS, MS, MS, MS, MS,

Sweet, soapy, apple, leafy Vinegar, acidic, cheesy Mushroom, fishy, grass, fatty Fresh, light green, nutty Baked potato, meaty, onion

19.4 5.5 1.5 5.5 0.5

y = 0.3361x y = 0.5411x y = 1.9798x y = 0.5548x y = 0.0874x

0.0014 0.0025 0.0057 0.0005 0.0004

10 ng 10 ng 10 ng 10 ng 10 ng

-1000 ng - 300 ng - 300 ng - 300 ng -1000 ng

2.454 1.632 1.856 1.002 2.547

1484

MS

1483

MS, RI, Std, O

Fried, fatty, nutty

94.8

y = 0.7101x

0.0021 (0.9145)

10 ng - 300 ng

2.101

1484 1487

MS, RI, Std MS, RI, Std

Wood, almond, flowery

9562.0 2525.0

y = 1.4018x y = 0.6033x

0.0027 (0.9626) 0.0012 (0.9763)

10 ng - 300 ng 10 ng - 300 ng

1.445 1.369

1497 1498

MS, RI, Std MS, RI

Green rosy

25482.2

y = 1.1605x

0.0037 (0.8791)

10 ng - 300 ng

2.569

1508

MS, RI, Std, O

Fatty, hay, fishy odor

15.4

y = 0.7548x

0.0017 (0.9664)

10 ng - 300 ng

1.889

1510 1525 1544 1550

MS, RI, Std MS, RI, Std MS, RI, Std MS

Smoky, tobacco, narcotic Coconut, waxy, cucumber Bitter almond, woody, burnt

15025.2 8598.8 750.9

y = 1.3368x y = 0.1207x y = 1.7699x

0.0035 (0.9717) 0.0001 (0.9642) 0.0016 (0.9971)

10 ng - 300 ng 10 ng -1000 ng 10 ng - 300 ng

1.432 1.888 1.023

1559

MS, RI, Std, O

Fruity, onion

95.1

y = 0.5044x

0.0018 (0.9205)

10 ng - 300 ng

1.560

1566 1576

MS, RI, Std, O MS, RI, Std

Herbaceous, fatty, green Cheese, phenolic, fatty, sweety

125.9 6550.6

y = 0.4074x y = 0.7014x

0.0011(0.9384) 0.0011(0.9458)

10 ng - 300 ng 10 ng - 300 ng

1.668 1.478

1582

MS, RI, Std, O

Cheese, tropical fruit, pineapple

8.5

y = 0.3213x

0.0001 (0.9901)

10 ng - 300 ng

1.556

1588

MS, RI, O

Woody, mushroom, hay, fresh

1590 1593

MS, RI, Std, O MS, RI, Std, O

Almond, spicy, caramelic Fruity, onion

6.0 95.1

y = 1.0011x y = 0.3125x

0.0021 (0.9556) 0.0011 (0.9012)

10 ng - 300 ng 10 ng - 300 ng

1.568 2.121

1599

MS, RI, Std, O

Cucumber, wax, green

0.8

y = 0.5418x

0.0011 (0.9964)

10 ng - 300 ng

2.551

1606

MS, RI, Std, O

Fried fat, rancid, oily, soapy

0.1

y = 0.6418x

0.0015 (0.9154)

10 ng - 300 ng

1.235

1608

MS,RI

62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102 103

104 105 106 107 108 109

RI, RI, RI, RI, RI,

RI, RI, RI, RI, RI,

RI, RI, RI, RI, RI,

Std, Std, Std, Std, Std

Std, Std, Std, Std, Std,

Std, Std, Std, Std, Std,

O O O O O

O O O O O

0.0001 (0.9414) 0.0012 0.0059 0.0065 0.0067

(0.9254) (0.9230) (0.9600) (0.9533)

0.0005 (0.9503)

(0.8578) (0.919) (0.9154) (0.9970) (0.8650)

-

-

1031

A. Giri et al. / Food Research International 43 (2010) 1027–1040 Table 2 (continued) Peak no

Volatile compounds

RIa

Identificationb

Odor description

Threshold (lg/L)

Standard curve (r)

Validation range

RSD%

110 111 112 113 114 115 116 117

2-Undecanone Butanoic acid Acetophenone Ethyl decanoate 1-Nonanol Ethyl benzoate Furfuryl alcohol 2-Methyl butanoic acid 3-Methyl butanoic acid 2-Ethoxy thiazole Ethyl 3 (2-furyl) propanoate 4-Ethyl benzaldehyde 3-(Methylthio) propanol Ethylphenyl acetate 2-Phenylethyl acetate p-Guaiacol Benzyl alcohol Phenylethyl alcohol Benzene acetaldehyde aethylidiene 2-Acetyl pyrrole Phenol 4-Vinyl guaiacol 3-Ethoxy benzaldehyde 4-Ethyl guaiacol 2-Methyl phenol 4-Ethyl Phenol

1610 1630 1644 1649 1671 1683 1689 1690

MS, MS, MS, MS, MS, MS, MS, MS,

Tallow, musty, green, ketone Rancid butter, cheesy Glue, musty Grape, Dusty, oily, green, floral Chamomile, flowery, musty Fermented burnt sugar, sweet Sweet, cheese, rancid

5.5 0.2 65.9 5.0 45.5 55.6 4500.6 0.7

y = 0.3372x y = 0.8001x y = 0.5332x y = 0.0411x y = 0.2219x y = 0.4309x y = 0.5691x y = 0.9022x

0.0018 (0.7436) 0.0015 (0.9812) 0.0015 (0.8512) 0.0002 (0.8797) 0.0008 (0.9237) 0.0011(0.9569) 0.0009 (0.9776) 0.0001 (0.9145)

10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng 10 ng

- 300 ng - 300 ng - 300 ng -1000 ng -1000 ng - 300 ng - 300 ng - 300 ng

3.335 2.123 3.336 2.125 1.564 1.123 1.111 1.871

1693

MS, RI, Std

Over ripe fruit, sweaty, cashew

1560.9

y = 0.5849x

0.0001 (0.9228)

10 ng - 300 ng

1.235

1695 1697

MS, RI, Std, O MS

Strong, meaty, nutty, roasted

51.2

y = 0.4057x

0.0015 (0.9506)

10 ng - 300 ng

1.568

1728

MS, RI, Std, O

Fruity, anisic, minty, burnt

123.2

y = 1.5359x

0.0015 (0.9892)

10 ng - 300 ng

1.112

1735

MS, RI, Std, O

Raw potato, garlic, vegetable

856.1

y = 0.0929x

0.0004 (0.8500)

10 ng -1000 ng

3.568

1799

MS, RI, Std, O

Honey, rose, yeast nuance

155.6

y = 0.1768x

0.0001 (0.9094)

10 ng -1000 ng

1.698

1834

MS, RI, Std, O

Honey, rosy with green nectar

249.6

y = 0.148x

10 ng -1000 ng

2.569

1883 1893 1931

MS,RI MS, RI, Std MS, RI, Std, O

Sweet floral, aromatic, fruity Honey, rose, lilac, spicy

2546.2 564.2

y = 1.2361x y = 0.2596x

0.0024 (0.9756) 0.0003 (0.9512)

10 ng - 300 ng 10 ng -1000 ng

1.445 1.231

1955

MS

1993 2025 2033 2039

MS, RI, Std MS, RI, Std MS, RI, Std, O MS,RI

Herbal, nutty, anisic, sweet Medicinal, acrid, tarry Woody, amber, ceder, peanut

58585.3 5501.2 12.0

y = 0.0425x y = 0.5068x y = 0.6413x

0.0002 (0.9294) 0.0023 (0.9173) 0.0011 (0.9215)

10 ng -1000 ng 10 ng - 300 ng 10 ng - 300 ng

1.669 1.998 1.568

2046 2103 2195

MS, RI, Std, O MS, RI, Std MS, RI, Std

Clove, spicy with vanilla note Bitumen, manure, leather Shoe polish, leather, smoky

89.3 550.6 1010.1

y = 0.02145x 0.0001 (0.8989) y = 0.0814x 0.0002 (0.9010) y = 0.0564x 0.0002 (0.9513)

10 ng -1000 ng 10 ng -1000 ng 10 ng -1000 ng

2.365 1.556 1.568

118 119 120 121 122 123 124 125 126 127 128

129 130 131 132 133 134 135 a b

RI, RI, RI, RI, RI, RI, RI, RI,

Std, Std, Std, Std, Std, Std, Std Std,

O O O O O O O

0.0004 (0.8855)

RI, retention index. MS, mass spectrum; RI, retention index; Std, confirmed by authentic standards; O, olfactometrical confirmation.

40–350 m/z and 500 m/s, respectively. The column temperature was initially held at 40 °C for 3 min and subsequently programmed to 200 °C at a rate of 3 °C/min.

was introduced to the 3 g of sample in the flask just before the subsequent flavor isolation process. 2.8. Threshold testing and odor descriptions of volatile compounds

2.7. Identification and quantification of volatile compounds The volatiles were identified based on comparison of their mass spectra and relative abundances. Each compound was further confirmed by comparing its mass spectra, linear retention index (LRI), and retention times with those obtained for authentic standards. Kovats retention index (RIs) were determined for both GC-FID-O and GC–MS by using a series of n-alkanes (C5–C24). To improve the detection limits, the selected ion monitoring (SIM) mode was used. Olfactometric identification was also carried out to achieve flavor perception and relative intensities of the volatiles. Sniffers (n = 17) were well introduced about the flavor perceptions of the preliminarily identified volatiles by means of authentic standards. Quantification of the major volatile compounds was carried out by standard curves obtained by each compound at least from five different concentrations in methanol. The standard curves achieved were presented in Table 2, where ‘‘y” represents the peak area ratio (peak area of volatile standard/ peak area of IS) and ‘‘x” represents the concentration ratio (concentration of volatile standard/concentration of IS). The levels of the volatile compounds were normalized by 2,4,6-trimethylpyridine equivalents (assuming all of response factors to the FID were 1). The methanolic solution of internal standard 2,4,6-trimethylpyridine (0.01 mL of 1000 ppm)

Orthonasal detection thresholds of volatiles in water were determined using the forced choice ascending concentration series method of limits (ASTM, 1992). Compounds were diluted in methanol (for the water threshold) before addition to the deodorized water. The deodorized water was prepared by boiling deionized water to two-thirds of its volume. The compound concentrations were serially diluted by a factor of 5 for 7 concentration series for the threshold test. Blank samples in each set were adjusted with the same concentration of methanol to eliminate any bias due to the solvent used. Each 10 mL volume screw capped test tube was filled up to 5 mL and was allowed to equilibrate for 1 h before testing. All sample preparation and testing was carried out with the lights off to minimize compound degradation during this time. Each concentration in the series was presented to the panels (n = 15) with 2 blank in a randomized order and were asked to choose the different sample containing the testing compound based on odor perception. The individual best estimated threshold was calculated by taking the geometric mean of the last concentration, which was incorrect, and the 1st concentration that was correct with no further samples missed. The group threshold was calculated as the geometric mean of the individual best estimate thresholds. The most common odor descriptions of volatile standard compounds over threshold concentrations obtained from

Peak no.

Volatile compounds

Fish miso with koji

Fish miso without koji

Soy miso (light)

1032

Table 3 Concentration (xg/kg of sample) and OAVs (in parenthesis) of volatiles detected in different commercial miso and sauce samples. Soy miso (dark)

Nampla (premium)

Nampla (standard)

Nouc-mam

Oyster sauce

Soy sauce (light)

Soy sauce (dark)

Aldehydes Acetaldehyde Propanal 2-Methylpropanal 2-Methylbutanal 3-Methylbutanal Pentanal 2-Butenal Hexanal 2-Methyl-2-butenal (E) Heptanal 2-Hexenal (Z) 4-Heptenal Octanal Nonanal 2,4-Heptadienal (EZ) 2,4-Heptadienal (EE) 2,6-Nonadienal (EZ) 2,4-Nonadienal (EZ) Subtotal

6.50 1.59 7.35 12.84 2.05 0.08 0.08 0.49 0.09 3.35 0.10 0.06 0.62 0.15 5.06 0.11 0.11 0.43 41.05

(0.3) (0.1) (4.9) (12.8) (12.9) (<0.1) (0.2) (0.1) (<0.1) (1.2) (<0.1) (<0.1) (1.0) (0.1) (0.1) (<0.1) (0.1) (3.4) (37.2)

2.12 9.00 33.38 33.87 10.22 0.66 1.54 2.25 0.14 4.97 0.28 0.10 3.68 0.35 13.90 0.19 0.79 0.64 118.09

(0.1) (0.6) (22.2) (33.7) (64.3) (<0.1) (4.3) (0.4) (<0.1) (1.7) (<0.1) (<0.1) (6.3) (0.3) (0.1) (<0.1) (1.0) (5.2) (140.4)

1.16 1.95 4.84 2.88 0.47 0.06 0.07 1.91 0.05 1.79 0.05 0.07 0.39 0.16 0.69 0.08 0.10 0.27 16.97

(<0.1) (0.1) (3.2) (2.9) (3.0) (<0.1) (0.2) (0.4) (<0.1) (0.6) (<0.1) (<0.1) (0.7) (0.3) (<0.1) (<0.1) (0.1) (2.1) (13.5)

1.37 1.57 11.76 1.64 0.39 0.09 0.09 0.96 0.05 1.34 0.05 0.06 0.42 0.14 0.62 0.08 0.09 0.26 20.99

(0.1) (0.1) (7.8) (1.6) (2.5) (<0.1) (0.3) (0.2) (<0.1) (0.5) (<0.1) (<0.1) (0.7) (0.1) (<0.1) (<0.1) (0.1) (2.1) (16.1)

0.21 0.34 11.67 7.15 0.38 0.05 0.72 0.80 0.07 19.78 0.07 0.08 0.11 0.19 0.39 0.19 0.07 0.31 42.57

(<0.1) (<0.1) (7.8) (7.1) (2.4) (<0.1) (2.0) (0.2) (<0.1) (6.9) (<0.1) (<0.1) (0.2) (0.2) (<0.1) (<0.1) (<0.1) (2.5) (29.4)

0.12 2.54 16.92 0.45 2.05 0.12 0.87 1.49 0.05 9.71 0.09 0.05 0.11 0.19 0.56 0.12 0.07 0.95 36.46

(<0.1) (0.2) (11.3) (0.4) (12.9) (<0.1) (2.5) (0.3) (<0.1) (3.4) (<0.1) (<0.1) (0.2) (0.2) (<0.1) (<0.1) (<0.1) (7.6) (39.0)

4.38 5.91 13.04 4.13 0.32 0.07 0.50 1.15 0.06 12.07 0.11 0.08 0.14 0.18 0.62 0.08 0.07 1.18 44.10

(0.2) (0.4) (8.7) (4.1) (2.0) (<0.1) (1.4) (0.2) (<0.1) (4.2) (<0.1) (<0.1) (0.2) (0.2) (<0.1) (<0.1) (<0.1) (9.4) (31.2)

0.69 0.34 1.77 0.53 0.17 0.10 0.04 0.64 0.05 6.12 0.04 0.08 0.16 0.17 0.41 0.08 0.08 0.87 12.33

(<0.1) (<0.1) (1.2) (0.5) (1.0) (<0.1) (0.1) (0.1) (<0.1) (2.1) (<0.1) (<0.1) (0.3) (0.2) (<0.1) (<0.1) (<0.1) (7.0) (12.7)

0.13 5.65 44.32 11.36 5.88 0.06 0.40 0.63 0.04 0.69 0.07 0.05 0.11 0.15 1.72 0.08 0.07 0.15 71.56

(<0.1) (0.4) (29.5) (11.3) (37.0) (<0.1) (1.1) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (<0.1) (<0.1) (1.2) (81.3)

9.03 1.56 80.90 0.74 0.15 0.06 0.13 1.00 0.05 1.08 0.05 0.06 0.14 0.15 0.25 0.08 0.07 0.08 95.58

(0.4) (<0.1) (53.9) (0.7) (0.9) (<0.1) (0.4) (0.2) (<0.1) (0.4) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (<0.1) (<0.1) (0.6) (58.1)

10 11 21 27 28 32 37 38 42 44 45 47 53 60 65 66 67 68 69 72 73 77 79 80 86 87 93 97 100 101 106 114

Alcohols 2-Propanol Ethanol 1-Propanol 3-Methyl-2-butanol 2-Methyl-propanol 3-Pentanol 1-Butanol 1-Penten-3-ol 3-Hexanol 2-Methyl-1-butanol 3-Methyl-1-butanol 2-Hexanol 1-Pentanol 3-Methyl 1-pentanol 2-Ethyl-1-butanol Cyclopentanol 2-Heptanol 2-Penten-1-ol (E) 3-Methyl-3-buten-1-ol 1-Hepten-3-ol Hexanol 3-Octanol 2-Hexen-1-ol (E) 2-Hexen-1-ol (Z) 1-Octen-3-ol Heptanol 2-Ethyl hexanol 2-Nonanol 2,3-Butanediol (levo) Octanol 2,3-Butanediol (meso) Nonanol Subtotal

0.30 361.69 262.67 453.73 1.43 0.70 44.58 0.24 0.08 1.64 10.78 2.70 0.69 1.83 4.37 0.42 1.51 0.07 0.93 0.95 28.86 0.83 3.61 0.13 0.82 0.42 19.50 16.81 3.63 0.48 240.96 0.83 1468.20

(<0.1) (<0.1) (<0.1) (0.4) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (0.1) (2.6) (<0.1) (<0.1) (0.2) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) NEa (4.9) (<0.1) (<0.1) (<0.1) (0.5) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (2.5) (<0.1) (11.6)

1.46 889.72 52.08 190.79 0.57 0.35 27.74 0.36 0.27 3.55 58.77 15.55 0.24 2.72 0.58 1.71 0.87 0.07 1.29 4.33 15.20 0.21 3.18 0.22 4.22 1.30 3.26 7.22 3.99 1.07 106.14 1.62 1400.64

(<0.1) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (0.2) (14.6) (<0.1) (<0.1) (0.4) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (2.7) (<0.1) (<0.1) (<0.1) (2.8) (0.2) (<0.1) (<0.1) (<0.1) (<0.1) (1.1) (<0.1) (22.4)

0.75 780.11 16.19 30.43 0.35 0.08 58.99 0.16 0.38 1.87 50.07 0.59 0.07 0.97 1.47 0.21 0.19 0.05 0.28 0.32 3.35 0.13 0.28 0.10 0.31 0.08 0.56 1.47 0.30 0.24 74.03 0.31 1024.66

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (0.1) (12.5) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.6) (<0.1) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.8) (<0.1) (14.5)

1.15 493.10 3.02 12.36 0.29 0.25 25.99 0.23 0.09 0.80 27.90 0.36 0.14 1.30 0.78 0.13 0.31 0.07 0.13 0.15 1.69 0.32 0.32 0.11 0.18 0.04 0.41 2.50 0.21 0.22 89.48 0.39 664.42

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (<0.1) (7.0) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.3) (<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.9) (<0.1) (8.6)

0.24 13.36 41.66 0.07 0.06 0.77 2.09 0.14 0.08 0.44 2.12 0.14 0.22 0.11 0.08 0.06 0.06 0.09 0.02 0.06 0.04 0.02 0.10 0.17 1.58 0.08 0.16 0.03 0.13 0.13 1.79 0.14 66.23

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.5) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) (<0.1) (<0.1) (1.0) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (1.7)

0.08 14.49 29.54 0.06 0.06 0.80 3.32 0.14 0.07 0.45 2.32 0.20 0.42 1.08 0.05 0.05 0.07 0.44 0.04 0.06 0.04 0.02 0.10 0.11 1.99 0.10 0.15 0.03 0.17 0.11 3.10 0.15 59.82

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) (<0.1) (<0.1) (1.3) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (2.1)

0.09 7.34 14.95 0.05 0.06 0.28 1.54 0.15 0.05 0.08 1.17 0.08 0.02 0.01 0.06 0.07 0.05 0.05 0.07 0.06 0.04 0.02 0.10 0.11 1.99 0.18 0.43 0.03 0.13 0.10 1.28 0.21 30.87

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) (<0.1) (<0.1) (1.3) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (1.7)

0.08 120.31 1.13 0.06 0.06 0.36 0.41 0.13 0.07 0.09 0.33 0.05 0.01 0.05 0.04 0.05 0.02 0.09 0.03 0.08 0.47 0.08 0.10 0.11 1.93 0.71 0.19 0.46 0.12 0.11 0.28 0.14 128.17

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.1) (<0.1) (<0.1) (<0.1) (1.3) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (1.6)

0.55 245.06 15.45 69.48 0.15 0.07 5.50 0.15 0.07 0.58 11.79 0.77 0.12 0.01 0.07 0.09 0.05 0.06 0.05 0.07 0.09 0.05 0.10 0.10 0.12 3.98 0.38 0.03 0.13 0.19 0.39 0.20 355.89

(<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (2.9) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (0.7) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (3.9)

0.18 179.98 5.23 0.85 0.15 0.20 1.73 0.19 0.08 0.10 1.51 0.35 0.04 0.33 0.12 0.43 0.11 0.08 0.07 0.10 0.16 0.09 0.10 0.10 0.13 0.55 0.20 0.03 0.13 0.19 2.17 0.35 196.00

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.4) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.7)

A. Giri et al. / Food Research International 43 (2010) 1027–1040

2 3 5 8 9 15 20 26 29 40 46 50 58 78 90 95 107 108

6 13 17 19 22 23 25 30 33 36 49 54 56 59 70 84 113

134.92 5.30 2.87 1.56 6.01 4.46 59.30 2.72 0.72 0.41 14.87 1.60 NDb

(26.9) (34.4) (0.1) (1.6) (37.8) (29.4) (1.0) (0.1) (0.5) (0.1) (4.6) (0.1) ND

8.56 0.57 11.84 0.99 0.69 0.36 2.23 0.17 0.37 0.06 4.79 0.60 ND

(1.7) (3.7) (0.5) (1.0) (4.3) (2.4) (<0.1) (<0.1) (0.2) (<0.1) (2.1) (<0.1) ND

25.65 1.58 0.43 1.57 0.58 1.15 0.59 1.06 0.39 0.12 0.15 0.62 ND

(5.1) (10.2) (<0.1) (1.6) (3.7) (7.6) (<0.1) (<0.1) (0.3) (<0.1) (0.1) (<0.1) ND

11.11 0.84 0.14 0.78 0.91 1.02 0.63 0.62 0.29 0.22 0.07 0.51 ND

(2.2) (5.5) (<0.1) (0.8) (5.7) (6.7) (<0.1) (<0.1) (0.2) (<0.1) (<0.1) (<0.1) ND

0.09 0.05 0.41 1.12 0.92 0.13 0.08 0.11 0.06 0.04 0.03 0.10 ND

(<0.1) (0.3) (<0.1) (1.1) (5.8) (0.9) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND

0.05 0.03 1.66 0.08 1.39 0.08 0.05 0.02 0.04 0.03 0.03 0.09 0.64

(<0.1) (0.2) (0.1) (0.1) (8.7) (0.5) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1)

0.06 0.11 13.34 0.08 0.17 0.10 0.05 0.03 0.03 0.04 0.03 0.07 0.14

(<0.1) (0.7) (0.5) (0.1) (1.1) (0.6) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1)

14.24 0.22 0.42 0.11 0.15 0.08 0.06 0.04 0.04 0.04 0.03 0.07 0.10

(2.8) (1.4) (<0.1) (0.1) (0.9) (0.5) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1)

5.47 0.33 0.46 0.81 1.12 0.54 5.40 0.16 0.11 0.37 0.52 0.32 0.09

(1.1) (2.2) (<0.1) (0.8) (7.0) (3.5) (0.1) (<0.1) (0.1) (0.1) (0.2) (<0.1) (<0.1)

0.96 0.69 0.59 0.60 0.17 0.62 5.88 0.12 0.13 0.08 0.99 0.59 0.12

(0.2) (4.5) (<0.1) (0.6) (1.0) (4.1) (0.1) (<0.1) (0.1) (<0.1) (0.4) (<0.1) (<0.1)

1.80 8.35 2.07 24.76 271.71

(0.1) (4.2) (0.1) (4.3) (145.2)

0.45 3.22 0.66 19.25 54.81

(<0.1) (1.6) (<0.1) (3.8) (21.5)

0.42 1.14 0.66 13.24 49.35

(<0.1) (0.6) (<0.1) (2.6) (31.9)

0.44 1.16 0.96 13.42 33.11

(<0.1) (0.6) (<0.1) (2.7) (24.6)

0.08 0.07 0.15 0.16 3.61

(<0.1) (<0.1) (<0.1) (<0.1) (8.3)

0.08 0.07 0.20 0.16 4.71

(<0.1) (<0.1) (<0.1) (<0.1) (9.8)

0.03 0.07 0.23 0.38 14.97

(<0.1) (<0.1) (<0.1) (0.1) (3.2)

0.03 0.14 0.16 0.28 16.21

(<0.1) (0.1) (<0.1) (0.1) (6.0)

0.29 1.08 0.38 2.36 19.80

(<0.1) (0.5) (<0.1) (0.5) (16.2)

0.48 0.44 0.63 0.16 13.24

(<0.1) (0.2) (<0.1) (<0.1) (11.4)

4 7 14 16 18 52 57 71 104 110

Ketones 2-Propanone 2-Butanone 2-Pentanone 2,3-Butanedione 2,3-Pentanedione 3-Octanone 2-Octanone 6-Methyl-5-hepten-2-one 3,5-Octadien 2-one (EE) 2-Undecanone Subtotal

0.51 ND 0.10 5.57 173.94 0.87 0.89 4.50 0.37 6.80 193.55

(<0.1) ND (<0.1) (94.5) (<0.1) (<0.1) (<0.1) (<0.1) NE (1.2) (95.9)

5.64 ND 0.26 1.65 36.01 6.78 0.39 3.18 0.06 10.15 64.12

(<0.1) ND (<0.1) (28.0) (<0.1) (0.3) (<0.1) (<0.1) NE (1.8) (30.2)

1.84 18.40 0.06 1.57 11.01 0.22 0.06 0.50 0.02 1.62 35.31

(<0.1) (<0.1) (<0.1) (26.7) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.3) (27.0)

3.36 7.54 0.05 0.56 10.41 0.26 0.17 0.35 0.02 1.08 23.82

(<0.1) (<0.1) (<0.1) (9.5) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.2) (9.7)

0.12 8.30 0.08 0.27 9.67 0.07 0.03 0.23 0.03 2.00 20.79

(<0.1) (<0.1) (<0.1) (4.6) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.4) (4.9)

0.07 ND 0.10 0.33 0.15 0.60 0.03 0.19 0.02 4.09 5.57

(<0.1) ND (<0.1) (5.6) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.7) (6.4)

0.09 0.58 0.06 0.18 0.33 0.14 0.03 0.22 0.02 0.30 1.94

(<0.1) (<0.1) (<0.1) (3.0) (<0.1) (<0.1) (<0.1) (<0.1) NE (0.1) (3.1)

0.05 1.38 0.08 0.62 0.25 0.08 0.03 0.19 0.02 0.18 2.88

(<0.1) (<0.1) (<0.1) (10.5) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (10.6)

9.34 2.51 0.06 1.11 5.32 1.36 1.48 0.46 0.02 0.44 22.11

(<0.1) (<0.1) (<0.1) (18.8) (<0.1) (0.1) (<0.1) (<0.1) NE (0.1) (18.9)

0.14 0.90 0.06 0.71 6.36 11.21 0.45 0.70 0.02 0.56 21.12

(<0.1) (<0.1) (<0.1) (12.0) (<0.1) (0.5) (<0.1) (<0.1) NE (0.1) (12.7)

12 34 48 91 96 105 116 120

Furans 2-Ethylfuran 2-n-Butylfuran 2-Pentylfuran 2-Furaldehyde 2-Acetylfuran 5-Methylfurfural Furfuryl alcohol Ethyl-3-(2-furyl) propanoate Subtotal

4.17 0.12 0.13 2.69 0.31 3.84 3.70 0.31 15.27

(1.8) (<0.1) (<0.1) (<0.1) (<0.1) (0.6) (<0.1) NE (2.5)

11.41 0.16 0.57 7.43 0.35 5.39 4.92 1.60 31.83

(4.9) (<0.1) (0.1) (<0.1) (<0.1) (0.9) (<0.1) NE (5.9)

1.82 0.10 0.13 2.19 0.14 0.90 6.90 0.02 12.20

(0.8) (<0.1) (<0.1) (<0.1) (<0.1) (0.1) (<0.1) NE (1.0)

0.40 0.07 0.16 1.72 0.13 1.28 9.17 ND 12.92

(0.2) (<0.1) (<0.1) (<0.1) (<0.1) (0.2) (<0.1) ND (0.4)

0.08 0.11 0.13 0.06 0.09 0.09 0.61 ND 1.19

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND (0.1)

0.03 0.09 0.13 0.06 0.11 0.16 0.62 ND 1.20

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND (0.1)

0.04 0.09 0.20 0.06 0.09 0.07 0.18 ND 0.74

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND (0.1)

0.06 0.07 0.30 0.76 0.09 2.66 0.31 0.05 4.30

(<0.1) (<0.1) (0.1) (<0.1) (<0.1) (0.4) (<0.1) NE (0.5)

0.03 0.03 0.28 1.11 0.10 1.04 14.78 0.06 17.44

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.2) (<0.1) NE (0.2)

0.09 0.05 0.39 9.35 0.13 8.98 2.00 0.10 21.09

(<0.1) (0.1) (0.1) (<0.1) (<0.1) (1.5) (<0.1) NE (1.6)

24 75 76 88 99 103 119 122

Sulfur-containing compounds Dimethyl disulfide 2,4,5-Trimethyl thiazole Dimethyl trisulfide 3-(Methylthio) propanal 2-(Methylthio) ethanol Ethyl 3-(methylthio) propanoate 2-Ethoxy thiazole 3-(Methylthio) propanol Subtotal

0.90 0.88 0.09 14.37 0.03 5.37 11.19 311.72 344.55

(0.8) (<0.1) (6.2) (31.4) NE (0.6) (0.2) (0.4) (39.7)

2.29 0.73 0.29 22.69 ND 0.52 66.05 1013.84 1106.41

(2.2) (<0.1) (19.2) (49.5) ND (0.1) (1.3) (1.2) (73.5)

0.64 0.55 0.03 5.07 ND 0.01 6.65 86.73 99.68

(0.6) (<0.1) (1.7) (11.1) ND (<0.1) (0.1) (0.1) (13.6)

0.96 0.48 0.03 4.21 ND 0.01 4.51 12.74 22.95

(0.9) (<0.1) (2.1) (9.2) ND (<0.1) (0.1) (<0.1) (12.3)

1.75 0.54 0.99 23.51 ND 0.01 0.12 6.73 33.72

(1.7) (<0.1) (66.0) (51.2) ND (<0.1) (<0.1) (<0.1) (118.0)

1.95 0.47 0.93 28.26 ND 0.01 0.12 1.45 33.73

(1.8) (<0.1) (61.8) (61.7) ND (<0.1) (<0.1) (<0.1) (125.4)

1.12 0.63 0.94 27.10 ND 0.01 0.23 0.58 30.61

(1.1) (<0.1) (62.5) (59.3) ND (<0.1) (<0.1) (<0.1) (122.9)

1.26 0.54 0.84 22.70 ND 0.30 0.27 0.20 26.22

(1.2) (<0.1) (56.2) (49.6) ND (<0.1) (<0.1) (<0.1) (107.0)

0.11 0.27 0.04 2.49 ND 0.01 0.58 1.90 5.39

(0.1) (<0.1) (2.9) (5.4) ND (<0.1) (<0.1) (<0.1) (8.4)

0.12 0.27 0.02 2.35 ND 0.01 7.49 1.36 11.61

(0.1) (<0.1) (1.2) (5.1) ND (<0.1) (0.1) (<0.1) (6.6)

Nitrogen-containing compounds Trimethylamine Methyl pyrazine 2,6-Dimethyl pyrazine 2,4-Dimethyl pyrazine

ND ND 0.21 0.04

ND ND (<0.1) (<0.1)

ND ND 0.21 0.04

ND ND (<0.1) (<0.1)

ND ND 0.21 0.04

ND ND (<0.1) (<0.1)

ND ND 0.21 0.04

ND ND (<0.1) (<0.1)

2.18 0.54 0.97 6.20

(42.7) (<0.1) (<0.1) (<0.1)

2.96 0.77 28.95 0.71

(58.1) (<0.1) (0.2) (<0.1)

2.14 1.62 9.28 4.60

(42.0) (<0.1) (0.1) (<0.1)

0.15 1.11 6.11 1.86

(3.0) (<0.1) (<0.1) (<0.1)

ND 0.17 8.96 0.04

ND (<0.1) (0.1) (<0.1)

ND 0.33 21.92 2.39

ND (<0.1) (0.1) (<0.1)

1 51 61 62

1033

(continued on next page)

A. Giri et al. / Food Research International 43 (2010) 1027–1040

Esters Ethyl acetate Ethyl isobutyrate 2-Methylpropyl acetate Ethyl butanoate Ethyl-2-methylbutanoate Ethyl-3-methylbutanoate Butyl acetate Isobutyl isobutanoate Isoamyl acetate Ethyl pentanoate Ethyl hexanoate 3-Methylbutyl butanoate 2-Methylbutyl 2methylbutanoate Isoamyl isovalerate Ethyl heptanoate Ethyl octanoate Ethyl decanoate Subtotal

Peak no.

Volatile compounds

Fish miso with koji

63 64 81 82 83 89 92 94 129

Ethyl pyrazine 2,3-Dimethyl pyrazine 2-Ethyl 3-methyl pyrazine 2,3,5-Trimethyl pyrazine 3-Ethyl- 2,3-dimethyl pyrazine 1,3-Dimethyl 1H-pyrazole Tetramethyl pyrazine 3,5-Diethyl 2-methyl pyrazine 2-Acetyl pyrrole Subtotal

0.07 0.10 ND ND ND 0.31 ND ND 113.22 113.95

130 131 132 133 134 135

85 102 111 117 118 a b

Aromatic compounds Ethyl benzene p-Xylene o-Xylene Cymene Propyl benzene p-Cymene Phenyl ethyne Benzaldehyde Benzene acetaldehyde Acetophenone Ethyl benzoate 4-Ethyl benzaldehyde Ethylphenyl acetate 2-Phenylethyl acetate p-Guaiacol Benzyl alcohol Phenylethyl alcohol Benzene acetaldehyde aethylidiene Phenol 4-Vinyl guaiacol 3-Ethoxy benzaldehyde 4-Ethyl guaiacol 2-Methyl phenol 4-Ethyl phenol Subtotal Acids Acetic acid 2-Methyl propanoic acid Butanoic acid 2-Methyl butanoic acid 3-Methyl butanoic acid Subtotal

NE, not estimated ND, not detected

Soy miso (light)

Soy miso (dark)

Nampla (premium)

Nampla (standard)

Nouc-mam

Oyster sauce

(<0.1) (<0.1) ND ND ND NE ND ND (<0.1) (<0.1)

0.07 0.10 ND ND ND 0.19 ND ND 37.92 38.53

(<0.1) (<0.1) ND ND ND NE ND ND (<0.1) (<0.1)

0.07 0.10 ND ND ND 0.00 ND ND 0.59 1.01

0.16 0.06 16.89 0.10 0.04 ND ND 6.40 3.62 ND 0.50 0.03 40.77 46.75 0.59 1.75 477.34 0.08

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND ND (<0.1) NE ND (<0.1) (<0.1) (0.3) (0.2) NE (<0.1) (0.8) NE

0.07 0.12 8.84 0.09 0.22 ND ND 0.71 22.64 ND 0.31 0.03 19.03 10.90 2.13 0.71 155.26 0.20

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND ND (<0.1) NE ND (<0.1) (<0.1) (0.1) (<0.1) NE (<0.1) (0.3) NE

3.01 0.33 0.03 149.97 5.04 6.07 759.52

(<0.1) (<0.1) NE (1.7) (<0.1) (<0.1) (3.1)

1.56 1.57 0.08 525.46 11.44 15.99 777.36

ND 36.31 ND 26.82 14.16 77.28

ND (<0.1) ND (39.7) (<0.1) (39.7)

ND 102.35 ND 4.21 33.74 140.30

Soy sauce (light)

Soy sauce (dark)

(<0.1) (<0.1) ND ND ND NE ND ND (<0.1) (<0.1)

0.07 0.10 ND ND ND 0.00 ND ND 2.65 3.07

(<0.1) (<0.1) ND ND ND NE ND ND (<0.1) (<0.1)

1.23 19.75 ND 0.34 0.08 0.09 0.66 0.05 0.16 32.23

(<0.1) (<0.1) ND (<0.1) NE NE (<0.1) NE (<0.1) (42.7)

0.87 1.17 0.02 1.09 0.02 0.00 0.69 0.09 0.16 37.50

(<0.1) (<0.1) (0.1) (<0.1) NE NE (<0.1) NE (<0.1) (58.3)

14.37 9.65 0.07 1.14 0.07 0.00 1.05 0.15 0.16 44.29

(<0.1) (<0.1) (0.2) (<0.1) NE NE (<0.1) NE (<0.1) (42.3)

0.58 5.65 0.01 0.30 0.05 0.00 0.15 0.04 0.16 16.18

(<0.1) (<0.1) (<0.1) (<0.1) NE NE (<0.1) NE (<0.1) (3.1)

0.07 1.36 ND 0.68 0.00 0.00 0.11 ND 13.19 24.59

(<0.1) (<0.1) ND (<0.1) NE NE (<0.1) ND (<0.1) (0.1)

2.50 21.70 ND 1.08 ND 0.05 0.08 ND 113.88 163.94

(<0.1) (<0.1) ND (<0.1) ND NE (<0.1) ND (<0.1) (0.2)

0.16 0.07 0.15 0.04 0.01 ND ND 1.38 2.65 ND 0.24 0.03 2.47 2.87 0.00 0.06 101.59 2.05

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND ND (<0.1) NE ND (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (0.2) NE

0.09 0.06 0.12 0.07 0.02 ND ND 1.25 6.37 ND 0.26 0.03 2.81 1.25 0.00 0.06 47.70 2.45

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND ND (<0.1) NE ND (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (0.1) NE

0.08 0.07 4.09 0.03 0.02 ND 0.02 19.91 ND 0.53 0.11 0.22 9.42 0.12 0.38 0.13 3.15 0.02

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND NE (<0.1) ND (<0.1) (<0.1) (<0.1) (0.1) (<0.1) NE (<0.1) (<0.1) NE

0.09 0.06 8.16 0.01 0.00 ND ND 8.05 ND 0.95 0.14 0.09 26.05 0.23 0.30 0.08 2.15 0.01

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) ND ND (<0.1) ND (<0.1) (<0.1) (<0.1) (0.2) (<0.1) NE (<0.1) (<0.1) NE

0.10 0.09 0.91 0.01 0.01 7.47 ND 0.16 ND 0.49 0.10 0.47 1.15 0.12 0.00 0.06 0.91 ND

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (1.5) ND (0.0) ND (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) ND

0.07 0.06 0.26 0.01 0.00 5.56 ND 15.24 ND ND 0.14 0.07 2.89 0.16 0.01 0.07 1.42 ND

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (1.1) ND (<0.1) ND ND (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) ND

0.11 0.08 0.14 0.01 0.01 3.06 0.01 0.90 ND ND 0.25 0.13 0.56 0.24 0.48 0.07 90.84 0.10

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) ND ND (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (0.2) NE

0.08 0.06 0.10 0.03 0.03 4.16 0.04 5.02 ND ND 0.23 0.10 0.64 0.51 0.80 0.41 17.58 0.18

(<0.1) (<0.1) (<0.1) (<0.1) (<0.1) (0.8) NE (<0.1) ND ND (<0.1) (<0.1) (<0.1) (<0.1) NE (<0.1) (<0.1) NE

(<0.1) (0.1) NE (4.9) (<0.1) (<0.1) (6.5)

0.19 0.17 0.03 18.64 0.59 0.12 133.54

(<0.1) (<0.1) NE (0.2) (<0.1) (<0.1) (0.4)

0.26 0.11 0.01 11.14 6.31 0.12 80.50

(<0.1) (<0.1) (<0.1) (0.1) (<0.1) (<0.1) (0.3)

10.65 0.06 ND 2.74 0.08 0.12 51.94

(<0.1) (<0.1) ND (<0.1) (<0.1) (<0.1) (0.2)

5.72 0.06 0.02 1.88 0.33 0.75 55.13

(<0.1) (<0.1) NE (<0.1) (<0.1) (<0.1) (0.2)

1.76 0.06 ND 0.16 0.08 0.12 14.20

(<0.1) (<0.1) ND (<0.1) (<0.1) (<0.1) (1.5)

0.15 0.07 ND 0.16 0.08 0.28 26.69

(<0.1) (<0.1) ND (<0.1) (<0.1) (<0.1) (1.2)

0.60 0.13 0.00 40.33 1.27 7.62 146.94

(<0.1) (<0.1) NE (0.5) (<0.1) (<0.1) (1.3)

2.06 0.18 0.01 13.54 1.13 0.97 47.87

(<0.1) (<0.1) NE (0.2) (<0.1) (<0.1) (1.1)

ND (<0.1) ND (6.2) (<0.1) (6.3)

ND 1.37 ND 2.48 17.13 20.98

ND (<0.1) ND (3.7) (<0.1) (3.7)

ND 2.06 ND 1.18 24.73 27.97

ND (<0.1) ND (1.7) (<0.1) (1.8)

252.34 196.30 22.33 52.09 636.99 1160.05

(45.5) (<0.1) (116.9) (77.2) (0.4) (240.1)

17.36 31.05 10.05 15.27 159.34 233.08

(3.1) (<0.1) (52.6) (22.6) (0.1) (78.5)

0.83 10.94 8.61 20.22 74.64 115.25

(0.2) (<0.1) (45.1) (30.0) (<0.1) (75.2)

1.03 21.82 11.45 14.26 73.07 121.63

(0.2) (<0.1) (60.0) (21.1) (<0.1) (81.3)

1.25 6.25 1.17 3.29 34.39 46.35

(0.2) (<0.1) (6.1) (4.9) (<0.1) (11.3)

2.38 173.91 2.34 39.42 173.83 391.88

(0.4) (<0.1) (12.3) (58.4) (0.1) (71.2)

A. Giri et al. / Food Research International 43 (2010) 1027–1040

31 35 39 41 43 55 74 98 109 112 115 121 123 124 125 126 127 128

Fish miso without koji

1034

Table 3 (continued)

1035

A. Giri et al. / Food Research International 43 (2010) 1027–1040 Table 4 Relative proportion (%) of OAVs of the primary odor-active compounds in different miso and sauce samples.

a

Peak no.

Volatile compounds

Respective organoleptic groupa

Fish miso with koji

Fish miso without koji

Soy miso (light)

Soy miso (dark)

Nampla (premium)

Nampla (standard)

Noucmam

Oyster sauce

Soy sauce (light)

Soy sauce (dark)

1 76 88 117 85 111 45 108 16 22 6 13 23 5 8 9

Trimethylamine Dimethyl trisulfide 3-(Methylthio) propanal 2-Methyl butanoic acid Acetic acid Butanoic acid 3 -Methyl-1-butanol 2,4-Nonadienal(EZ) 2,3-Butanedione Ethyl-2-methylbutanoate Ethyl acetate Ethyl isobutyrate Ethyl-3 -methy lbutanoate 2-Methylpropanal 2-Methylbutanal 3-Methylbutanal

Ammoniacal Fishy Meaty Cheesy Cheesy Cheesy Rancid Rancid Caramel Caramel Fruity Fruity Fruity Nutty Nutty Nutty

0.00 1.84 9.31 11.78 0.00 0.00 0.80 1.01 28.04 11.21 7.99 10.22 8.72 1.45 3.80 3.83

0.00 7.54 19.42 2.44 0.00 0.00 5.74 2.04 10.98 1.69 0.67 1.45 0.93 8.70 13.21 25.21

0.00 1.80 11.87 3.93 0.00 0.00 13.36 2.29 28.59 3.92 5.48 10.97 8.11 3.45 3.07 3.16

0.00 3.32 14.44 2.73 0.00 0.00 10.91 3.32 14.94 8.95 3.48 8.59 10.57 12.30 2.57 3.90

9.91 15.30 11.87 17.88 10.55 27.10 0.12 0.57 1.06 1.34 0.00 0.08 0.20 1.80 1.65 0.56

18.90 20.11 20.07 7.36 1.02 17.11 0.03 2.46 1.84 2.84 0.00 0.07 0.17 3.67 0.15 4.20

15.64 23.26 22.04 11.14 0.06 16.77 0.04 3.50 1.11 0.40 0.00 0.27 0.24 3.23 1.53 0.76

1.39 25.98 22.95 9.77 0.09 27.75 0.04 3.22 4.87 0.43 1.31 0.66 0.25 0.55 0.24 0.48

0.00 2.16 4.05 3.63 0.17 4.57 2.19 0.91 13.99 5.25 0.81 1.61 2.64 22.02 8.43 27.56

0.00 0.79 3.29 37.48 0.28 7.86 0.24 0.40 7.72 0.67 0.12 2.88 2.61 34.59 0.48 0.60

Volatile compounds were assigned to the respective organoleptic groups based on olfactometric characterization.

panels were considered as odor description of the respective compounds. 2.9. Calculation of odor activity values The odor activity value (OAV) is the ratio of concentration in the food of interest to sensory odor threshold for a particular compound (Nursten & Reineccius, 1996). It is suggested that OAVs provide further information on which compound(s) plays an important role in flavor (Guth & Grosch, 1994) since both concentration in the food and sensory threshold are taken into account. For each compound, the OAVs were determined by dividing the concentration of the odorant in the samples by the mean values of its estimated orthonasal threshold. 2.10. Sensory evaluation Flavor profile characterization for ten different types of fermented miso and sauce samples were performed by 17 sensory panelists from Department of Food Science and Technology, Tokyo University of Marine Science and Technology, Japan. The training was conducted for four different types including fish miso, soy miso, fish sauce and soy sauce samples over two consecutive sessions of 3 h each in the sensory room in isolated booths at room temperature and under mild light. Miso and sauce samples, stabilized at 40 °C were introduced to panel without any dilution. The order of the sample presentation was randomized. The attributes those best expressed both the fermented miso and sauce aroma characteristics were selected by panel. From them a total of eight consensual odor descriptors including ‘‘fishy”, ‘‘meaty”, ‘‘cheesy”, ‘‘rancid”, ‘‘fruity”, ‘‘nutty”, ‘‘ammoniacal” and ‘‘caramel” were established. The respective odor intensities were rated on the following scale using one interval steps: 0, very weak; 1, weak; 2, moderate; 3, strong; 4, very strong. Based on the panel generated consensual descriptors the sensory analysis was carried out for ten different miso and sauce products where each of 10 g of sample was introduced to panel. Between two sample analyses an interval of at least 15 min was maintained to avoid any sensory fatigue of the panelists. 2.11. Comparison test for reconstructed model aroma A comparison test was performed to evaluate the closeness between the odor of different miso and sauce products to their respective reconstructed model aroma. The model aromas were reconstructed with the 16 compounds listed in Table 4. All of the

compounds were diluted with methanol to a 1% concentration and then further diluted with deodorized water and mixed in the amounts based on the quantification results shown in Table 3. For the comparison test the panelists were same as those were used in sensory analysis mentioned in Section 2.10, however, the comparison test was carried out with different task to compare the original sample to their respective reconstructed model aroma based on same sensory attributes used in Section 2.10 with 5-point intensity scale ranged from -2 (much less intense) to +2 (much high intense) where 0 was considered as identical to the original sample. 2.12. Statistical analysis Statistical analysis was carried out by statistical analysis system software NCSS (NCSS 2007 vs. 07.1.10, Kaysville, Utah) for principal components analysis (PCA). PCA was carried out using OAVs of the individual volatile compounds as variables. Repeated measures analysis of variance (ANOVA) was used to analyze the sensory data to predict any panelists/sample/session effects and also to ensure that the product descriptors to be used discriminated between the samples. Significance of difference was defined at P < 0.05 for sensory analysis (n = 17). 3. Results and discussion 3.1. Quantification and characterization of headspace volatiles of fish miso, soy miso, fish sauce and soy sauce products A total of 135 head space volatile compounds isolated by Tenax TA trap were detected using GC-FID and were identified by mass spectra, retention indices, and odor descriptions in comparison with those of authentic standards. One hundred and twenty-three compounds were consistently quantified by standard curves of the authentic standards. The accuracy of standard curves were high as indicated by linear regression coefficient (r), which ranged from 0.7436 to 0.9988 and relative standard deviation (RSD) values which ranged from 0.896% to 3.654%. Rest twelve compounds were tentatively identified using either mass spectra and/or retention index and their quantification was carried out based on using the internal standard. Calculated retention indices, odor descriptions, estimated thresholds in water and standard curves with validation ranges are presented in Table 2. Volatile classes for different commercial miso and sauce samples were presented in Table 3. Aldehydes, due to their low threshold values, are important aroma compounds in different food

1036

A. Giri et al. / Food Research International 43 (2010) 1027–1040

stuffs. The odor description obtained in the present study clearly indicated that the volatile aldehydes can contribute to desirable aroma as well as rancid odor and flavor to the products. Straight and branched chain aldehydes generally provide herbaceous, grassy and pungent aromas, while unsaturated aldehydes are linked with vegetal and fishy notes. The aldehyde in fermented meat paste products are generally derived from the Strecker degradation of amino acids (Dwivedi, 1975). In some cases they can also be originated from the decomposition of hydroperoxides and peroxyl radicals supposed to be initial product of oxidized fat (Sink, 1973). For all the miso samples, the volatile alcohol class contributed the major volatiles. Ethanol, although present at higher levels, did not reach its perception threshold in any types of miso. Sugar fermentation by mold such as Saccharomyces cerevisiae, Saccharomyces ellipsoidens, and mixed culture of Aspergillus sp. yields alcohols, however, a smaller proportion of alcohol may be formed by decomposition of secondary hydroperoxide of fatty acids (Girand & Durance, 2000). Branched chain alcohols are produced during yeast fermentation from two different sources: from carbohydrates by the EMP pathway and from amino acids via the Ehrlich pathway. 3-Methyl1-butanol can be attributed to the oxidative deamination of the free amino acids precursors leucine by the Ehrlich mechanism. The acetates of higher alcohols and the ethyl esters of fatty acids are, perhaps, the most desirable compounds in food products to enhance the aroma of the finished products. The esters are produced during the alcoholic fermentation of the fish meat or soy and koji, and for this reason it is possible to conclude that the yeasts together with the fermentation condition have a desirable role in the production. In freshly fermented miso, ethyl ester produced a fruity nuance. The esters of medium chain fatty acids are of the most interest in this respect. Higher amount of yeast strain and oxidative state of fermentation can accelerate the formation of this group of volatile compounds. One of the major groups of carbonyl compounds, those formations appeared to be associated with microbial activity, was ketones. The volatile ketones were most likely the products of lipid and/or amino acid degradation and several were reported among the volatile compounds in a fermenting room of Natto, a fermented soy paste in Japan (Tanaka & Shoji, 1993). Autoxidation of unsaturated fatty acids via hydroperoxides generation has also been reported as a possible mechanism for the formation of methyl ketones (Thomas, Dimick, & McNeil, 1971). Selke, Rohwedder, and Dutton (1975) reported the formation of a homologous series of methyl ketones as a result of b-oxidation of the carbon chain from the carbonyl end followed by decarboxylation. Among the ketones detected in miso and sauce samples, diacetyl (2,3-butanedione) is quite important sensorily than others because of its very low threshold value. 2,3-Butanedione generally contributes caramel and butter scotch nuance of the product. However, the formation, subsequent re-assimilation by the yeast followed by degradation of diacetyl to acetoin and further to 2,3-butanediol may significantly decrease the sweet aroma of the product because the reduced compounds have much higher thresholds and lesser impact on the miso and sauce aroma. Among the hetrocyclics, furans are well reported in different fermented products. Furans can be found in dehydrated or fermented condensates of carbohydrate or formed by the Amadori rearrangement pathways (Whistler & Daniel, 1985). An oxidation product of fatty acids, 2-pentyl furan, was reported to being impart reversion, beany, grassy and licorice like flavor in soybean oil (Taylor & Mottram, 1990). The presence of several furans and its derivatives such as furfural, which could be resulted from glucose pyrolysis and the Maillard reaction, was confirmed on the fermented paste products (Lee, Kang, & Min, 2003). The sulfur compounds, including dimethyl disulfide with flavor notes of cooked cabbage and dimethyl trisulfide with meaty and

cooked onion flavor notes existed in several miso and sauce samples. Because of their low sensory threshold values, those sulfurcontaining compounds are important fraction of aroma in numerous food products. The sulfur-containing compounds have previously been reported in different fermented bean and fish paste products (Landaud, Helinck, & Bonnarme, 2007) and can be originated either from raw materials or during the fermentation process from the free, peptidic and proteinic sulfur-containing amino acids as well as glutathione pool in the fish tissue (Herbert & Shewan, 1976). The sulfurs in heterocyclic sulfur-containing compounds, such as thiazoles of roasted meaty odor note, may be derived from amino acids, including cysteine, cystine and methionine or from vitamin B1 (Girand & Durance, 2000). In the present study, 3-(methylthio) propanal (methional), a Strecker aldehyde of methionine, was detected in most of the miso and sauce samples. The production of methional was reported as a process mediated by amino transferase and a-ketoacid decarboxylase activities (Amarita, Fernandez-Espla, Requena, & Pelaez, 2001). 3(Methylthio) propanol (methionol), existed in several miso samples, has been reported to be produced by the 4-metylthio-2oxobutyric acid, a transamination product of methionine, which yields methional, followed by a reduction step. Detection of thioesters, namely 3-(methylthio) propanoic acid ethyl ester in different miso samples indicated the oxidative status of fermentation, that leads to the oxidative conversion of methional/methionol to respective acids and subsequently esterification by enzymatic or chemical reactions (Landaud et al., 2007). One of the major volatile classes with most odorous constituents is nitrogen-containing compounds. Among the nitrogen-containing compounds trimethylamine (TMA) contributed the major proportion and might impart fishy flavors to those fish sauce products. Although found in small nitrogen-containing heterocyclic pyrazines namely trimethyl pyrazine and tetramethyl pyrazine might impart roasted and nutty flavor to the several miso products. They are normally formed through the Maillard reaction (Fox & Wallace, 1997) during fermentation. However, certain microorganisms, for instance, Corynebacterium glutamicum are also able to biosynthesize pyrazines from amino acids (Fox, Lucey, & Cogan, 1990). Several aromatic compounds were detected in different miso and soy samples and might contribute quality aroma to different miso and sauce products. The aromatic compounds in several fermented food products are generally produced as a result of catabolism of aromatic amino acids, begins with a transamination step which produces indole pyruvate, phenyl pyruvate and p-hydroxy phenyl pyruvate formed from tryptophan, phenyl alanine and tyrosine, respectively (Marilley & Casey, 2004). Five volatile acids, including acetic acid, 2-methyl propanoic acid, butanoic acid, 2-methyl butanoic acid and 3-methyl butanoic acid existed in different commercial sauce products. However, none of the miso products contained acetic acid and butanoic acids. A distinct separation was observed between miso and sauce samples based on their relative proportion of acids. Volatile acid class represents the major constituents for the sauce samples except for soy sauce (light). On the other hand, all the miso products exhibited significantly lower proportion of acids. Volatile acids can be produced as a result of either lipolysis or from amino acid metabolism in terms of valine deamination (Montel, Masson, & Talon, 1998). 3.2. Assessment of the relative importance of key odor-active compounds To assess the importance of the major odor-active compounds to the overall aroma, 16 key compounds with an odor-active values of equal to or greater than intermediate (i.e. P 9) were considered to be potentially significant contributors. The relative proportion (%) of OAVs and respective organoleptic groups are presented in

1037

A. Giri et al. / Food Research International 43 (2010) 1027–1040

Table 4. TMA, usually described as ‘‘rotten fish, ammoniacal”, had an odor threshold of 0.051 lg/L and existed in different fish sauces. Among the fish sauce samples, Nampla (standard) resulted higher OAV and relative proportion (%) than Nampla (premium), Noucmam and oyster sauce. TMA was decided as a single most compound responsible for ‘‘ammoniacal” organoleptic perception of fish sauce products considering its odor note. The OAV and relative proportion (%) of dimethyl trisulfide with a odor threshold of 0.015 lg/L, described as ‘‘fishy, sulfury, cooked onion”, were significantly higher in Nampla (premium), Nampla (standard), Noucmam and oyster sauce than miso and soy sauce samples. A reduction of this off-odor impact in fish miso with koji than fish miso without koji indicating a role of koji to suppress the formation of such compounds. Considering its odor note, dimethyl trisulfide was classified under ‘‘fishy” organoleptic perception. The odor of 3-(methylthio) propanal (methional), described as ‘‘baked potato, meaty and onion”, had an odor threshold of 0.458 lg/L, and was found important aroma active compound for all the types of miso and sauce products. However, its odor impact was found maximum for fish sauces particularly in oyster sauce. It also exhibited greater impact on fish miso without koji compared to fish miso with koji. Considering the meaty odor perception methional was classified under ‘‘meaty” organoleptic perception. Three volatile acids, including 2-methyl butanoic acid, acetic acid and butanoic acid with odor thresholds ranging between 0.191 and 5.541 lg/L were described as ‘‘cheese, vinegar and rancid butter”. The odor impact of 2-methyl butanoic acid was higher in dark-color varieties of soy sauce, Nampla (premium) and fish miso with koji. Whereas the aroma impact of acetic acid was significantly higher in Nampla (premium) than other sauce products. In addition acetic acid was not detected in any of the miso products. The odor impact of butanoic acid was significantly higher in fish sauce products particularly in Nampla (premium) and oyster sauce than those of soy sauce products. Butanoic acid was also not detected in any miso samples. These acids were grouped together for representing organoleptic class of ‘‘cheese” considering their odor note. 3-Methyl-1-butanol with ‘‘rancid, pungent” odor note and 2,4nonadienal (EZ) with ‘‘fried fat, rancid” odor note, both representing the ‘‘rancid” organoleptic perception, had an odor threshold of 4.013 lg/L and 0.125 lg/L respectively. 3-Methyl-1-butanol characterized fish miso without koji and 2,4-nonadienal characterized Nouc-mam and oyster sauce with high rancid odor. 2,3-Butanedione and ethyl-2-methylbutanoate having ‘‘creamy and caramel” note represented the ‘‘caramel” organoleptic class and had a odor threshold of 0.059 lg/L and 0.159 lg/L, respectively. The aroma impact of 2,3-butanedione was higher in miso samples, moderate in soy sauces and quite lower in fish sauce samples. A significant difference was observed between fish miso with koji and fish miso without koji. The odor impact of ethyl-2-methylbutanoate was higher in fish miso with koji and dark varieties of soy miso. Ethyl acetate, ethyl isobutyrate and ethyl-3-methylbutanoate represented ‘‘fruity” organoleptic class and had an odor threshold values of 5.015, 0.154 and 0.152 lg/L, respectively. Aroma impacts

Ammoniacal 60 Nutty*

Fishy

*

Nutty

20

*

Fruity

Caramel

*

*

Cheesy

Rancid Fish miso without koji

Fruity

0

Caramel

Fishy

Ammoniacal 60 Fishy

40

Meaty

Cheesy

Fruity

0

Caramel

Nutty

*

20

Meaty

*

Soy miso (dark)

Nampla (premium)

*

40

Ammoniacal 60 Fishy

Nutty

*

20

Cheesy

Fruity

Meaty

Cheesy

* Rancid

Nampla (standatd)

Nouc-mam

Fishy

40 20

0

Caramel

Rancid

Rancid Soy miso (light)

To compare and assess the olfactometric perceptions obtained by GC-O, the 16 major odor-active compounds were grouped into their respective organoleptic classes and scored in terms of their relative proportion (%) of OAVs (Fig. 1). Comparative studies revealed that olfactometric proportion (%) were significantly lower for meaty and nutty nuance in fish miso prepared with koji than without koji. In addition fruity and caramel aroma developed higher in fish miso prepared with koji. Significant differences for nutty odor between soy miso light and dark varieties were observed. However, both the products exhibited higher caramel and fruity aroma notes. Nampla (premium) and Nampla (standard) were both olfactometrically characterized by strong cheesy, meaty, fishy and ammoniacal odor note. Nouc-mam and oyster sauce, also exhibited high cheesy, meaty, fishy and ammoniacal note by olfactometric analysis, where ammoniacal odor were significantly higher in Nouc-mam and cheesy odor note were higher in oyster sauce. Soy sauce was olfactometrically characterized by high nutty and cheesy aroma notes with caramel nuance. Light-color varieties of soy sauce showed significantly higher nutty odor whereas significantly higher cheesy note was detected for dark-color varieties of soy sauce. Organoleptic perception of different miso and sauce products evaluated by panels were presented in Fig. 2. Significantly higher scores were observed for fishy, meaty, rancid, and nutty odor for fish miso prepared without koji and cheesy, caramel, and fruity aroma for fish miso prepared with koji. It can be concluded that addition of koji reduced the fishy, meaty, rancid and nutty aroma of the fish miso. For soy miso products organoleptic analysis could not differentiate significantly. Both the products exhibited higher caramel and fruity aroma notes by organoleptic analysis, also suggesting similarities to the results obtained from olfactometric analysis. Nampla (premium) and Nampla (standard) were characterized by strong cheesy, meaty, fishy and ammoniacal odor note. However, no significant differences were observed by organoleptic analysis. In contrast, organoleptic analysis revealed a significantly higher ammoniacal and nutty odor note for another fish sauce Nouc-mam. Organoleptic analyses strongly support the olfactometric findings for soy sauce products. In addition rancid odor

*

Nutty

20

*Meaty

0

Fish miso with koji

40

3.3. Olfactometric and organoleptic characterization and comparison of aroma profile

Ammoniacal 60

Ammoniacal 60

40

of those compounds were significantly higher in miso samples except fish miso prepared without koji. 2-Methylpropanal, 2-methylbutanal and 3-methylbutanal as described as ‘‘nutty, malty, almond” represented ‘‘nutty” organoleptic class and had odor threshold values of 1.501, 1.005 and 0.159 lg/L, respectively. The odor impact of 2-methylpropanal was higher in dark variety of soy sauce. The odor impact of both 2-methylbutanal and 3-methylbutanal were higher for fish miso without koji and light variety of soy sauce. Significantly lower ‘‘nutty” aroma impact for fish miso with koji than fish miso without koji, suggest that application of koji for fish miso can reduce the ‘‘nutty” odor.

Fruity

0

Caramel

Meaty

*

Cheesy

Rancid Oyster sauce

Soy sauce (light)

Fig. 1. Spider webs of Olfactometric proportions (%) for different miso and sauce samples ( indicates the significant differences).

Soy sauce (dark)

1038

A. Giri et al. / Food Research International 43 (2010) 1027–1040 Ammoniacal 4

*

Nutty

*

Fruity

* Fishy

3

Ammoniacal 4

Ammoniacal 4 3

Nutty

3

Nutty

Fishy

Ammoniacal 4 Fishy

Nutty

Ammoniacal 4

*

*

3

*

Fishy

3

Nutty

2

2

2

2

2

1

1

1

1

1

*Meaty

0

Caramel

Cheesy

*

*

*

Fruity

0

Caramel

Cheesy

Rancid Fish miso with koji

Meaty

Fruity

0

Caramel

0

Fruity

Cheesy

Soy miso (light)

Soy miso (dark)

Meaty

Caramel

Cheesy

Rancid

Rancid

Fish miso without koji

Meaty

Fruity

0

Nampla (standard)

Meaty

Caramel

Rancid

Nampla (premium)

Fishy

Cheesy

* Rancid

Nouc-mam

Oyster sauce

Soy sauce (light)

Soy sauce (dark)

Fig. 2. Spider webs of organoleptic scores for different miso and sauce samples ( indicates the significant differences).

Ammoniacal

Ammoniacal Nutty

Fishy

Fruity

Meaty

Caramel

Cheesy

Nutty

Fruity

Meaty

Caramel

Rancid Fish miso with koji (R)

Fish miso with koji (O)

Fish miso without koji (R)

Meaty

Caramel

Cheesy Rancid

Nampla (standard) (R)

Nutty

Fish miso without koji (O)

Caramel

Cheesy

Nouc-mam (R )

Caramel

Cheesy

Nouc-mam (O)

Fruity

Meaty

Cheesy

Oyster sauce (R )

Caramel

Soy miso (dark) (0)

Nutty

Fishy

Fruity

Meaty

Caramel

Fishy

Fruity

Meaty

Caramel

Cheesy

Cheesy Rancid

Rancid Soy sauce (light) (R )

Nampla (premium) (O)

Ammoniacal

Nutty

Oyster sauce (O)

Cheesy

Nampla (premium) (R )

Ammoniacal Fishy

Caramel

Meaty

Rancid

Soy miso (dark) (R)

Soy miso (light) (O)

Fishy

Fruity

Rancid

Soy miso (light) (R)

Nutty

Nutty

Meaty

Rancid

Rancid Nampla (standard) (O)

Cheesy

Fishy

Fruity

Ammoniacal

Meaty

Caramel

Meaty

Ammoniacal

Nutty

Rancid

Fishy

Fruity

Fishy

Fruity

Ammoniacal Fishy

Fruity

Cheesy

Ammoniacal

Nutty

Rancid

Ammoniacal Nutty

Ammoniacal

Fishy

Soy sauce (light) (O)

Soy sauce (dark) (R )

Soy sauce (dark) (O)

Fig. 3. Spider webs of organoleptic scores for comparison test of reconstructed model aroma for different miso and sauce samples (R indicates the reconstructed model aroma and O indicates the original sample).

To evaluate the present findings, a comparison test was performed for different miso and sauce products to their respective reconstructed model aroma. The model aromas were reconstructed with the 16 compounds listed in Table 4. The result of the similarity test for reconstructed model aroma were presented in Fig. 3. The organoleptic profiles of the reconstructed model aromas were similar to those of the miso and sauce products. However, little differences for some sauce products were observed without statistical significance. According to these results, it was confirmed that the 16 compounds selected as potent odorants appear to contribute to the odors of the miso and sauce products. 3.5. Multivariate analysis To simplify the interpretation of the relationship between the fermentation process of fish miso, fish sauce and soy sauce, principle component analysis (PCA) was carried out on the volatile components of 10 different fermented products. It is evident that PCA of OAVs using all the volatile compounds gave a distinct separation of the various fermented miso and sauce samples (data not shown). However, in the assessment of the flavor

5

100

4

80 All compounds (eigenvalue) Major 16 compounds (eigenvalue)

3

60

All compounds (cumulative percentage) Major 16 compounds (cumulative percentage)

2

1

0

40

20

1st

2nd

3rd

4th

5th

6th

0

Eigenvalue cumulative proportion (%)

3.4. Comparison test for reconstructed model aroma

of the large number of products, it would be advantageous if the number of the odor-active compounds could be reduced while maintaining adequate discrimination potential. A comparative study of eigenvalue and eigenvalue cumulative proportions (%) of first six principle components obtained from PCA of the OAVs of all the compounds and major 16 odor-active compounds are presented in Fig. 4. The results suggested that although the numbers of volatile compounds were reduced from 135 to 16, it still maintains adequate discrimination potential. A PCA plot using the factor loadings of OAVs of major 16 aroma active compounds in Table 4 is presented in Fig. 5a and b. Reducing

Eigenvalue

was observed significantly higher for dark varieties of soy sauce by organoleptic score. Comparison between olfactometric and organoleptic evaluation, however, revealed that some sensory attributes particularly the ‘‘ammoniacal” note for fish miso exhibit higher scores compared to olfactometric analysis. The results suggested that there might be other compounds including ammonia contributing to those notes which were not evident due to the limitations of the GC technique applied for volatile isolation.

Principal components Fig. 4. Eigenvalue and eigenvalue cumulative proportion of first six principal components from PCA using OAVs of all volatile compounds and of OAVs of the primary 16 odor-active compounds in different miso and sauce samples.

A. Giri et al. / Food Research International 43 (2010) 1027–1040

1039

Fig. 5. PCA plot using factor loadings of OAVs of primary 16 odor-active compounds in different miso and sauce samples on PC 1 and PC 2 (a) and on PC 1 and PC 3 (b).

Fig. 6. PCA plot using factor scores of OAVs of primary 16 odor-active compounds in different miso and sauce samples on PC 1 and PC 2 (a) and on PC 1 and PC 3 (b).

the number of the aroma compounds from 135 to 16 gave a good separation accounting for 63.99% of the total variance [PC1 (40.12%) + PC2 (23.87%)] (Fig. 5a) and was adequate for segregating the representative fermented products based on aroma. A distinct separation was achieved between all fish sauce samples as those were positioned on the positive side to the rest. The Fig. 5a also indicated the miso samples both prepared from fish and soy on the negative side of PC1 suggesting contrast in aroma profile from those of fish sauce samples. This is probably based on the fact that those fish sauces contain high levels of volatile acids and trimethylamine which were not found in the miso samples. However, a separation was achieved between fish miso with koji and without koji in the PCA plot between PC1 (40.12%) and PC3 (14.84%) (Fig. 5b), where fish miso with koji was positioned on the negative side along with soy miso (light and dark) and fish miso without koji on the positive side of PC3 with soy sauce (light). The compounds contributing to PC3, thus can characterize the effect of koji on the aroma profile of the fish miso. The factor scores of OAVs of major 16 aroma active compounds between PC1 and PC2 presented in Fig. 6a revealed that all the volatile acids, trimethylamine and 3-(methylthio) propanal were loaded in the positive side of PC1 and suggesting that those com-

pounds were responsible for characterizing the fish sauce aroma from other miso products. The PCA plot of the factor scores between PC1 and PC3 (Fig. 6b) revealed that most of the volatile esters and 2,3-butanedione were positioned in the negative side of both PC1 and PC3 suggesting the importance of those compounds to contribute fruity to caramel nuance in miso products apart from fish miso prepared without koji.

4. Conclusions A variety of compounds appear to have contributed the total flavor of different commercial miso and sauce products, however, the present findings can characterize and differentiate the products from each other based on olfactometric evaluation. The present results indicate that 16 major odor-active compounds should allow accurate characterization of miso and sauce products. Judging the relative abundances, it can be conclude that fermented fish and soy miso products are the result of alcoholic fermentation rather than acid fermentation. In contrary, different commercial sauce products confirmed acid fermentation as a technique for their production. In addition, the relative proportion of OAVs also revealed

1040

A. Giri et al. / Food Research International 43 (2010) 1027–1040

that overall aroma of most of the sauce products were characterized by volatile acids, which is in contrary to the miso products. Olfactometric and organoleptic findings clearly differentiated miso products with sweet, fruity aroma notes, whereas fish sauce products were characterized by ammoniacal, fishy, nutty and cheesy odor note. Soy sauce products, however, were dominated by nutty and cheese aroma. Use of koji for fish miso production was found effective to enhance sweet aroma to the product with a reduction of nutty, meaty and rancid nuance. References Amarita, F., Fernandez-Espla, D., Requena, T., & Pelaez, C. (2001). Conversion of methionine to methional by Lactococcus lactis. FEMS Microbiology Letters, 204, 189–195. Apriyantono, A., Husain, H., Lie, L., Jodoamidjojo, M., & Puspitasari-Nienaber, N. L. (1999). Flavor characteristics of Indonesian soy sauce (Kecap manis). In F. Shahidi & C. T. Ho (Eds.), Flavor chemistry of ethnic foods (pp. 15–31). Plenum Press: New York. ASTM. (1992). Standard practice for determination of odor and taste thresholds by a forced-choice method of limits. E-679-91. In Annual Book of Standards (Vol. 15.07, pp. 35–39). Philadelphia, PA: American Society for Testing and Materials. Dougan, J., & Howard, G. E. (1975). Some flavoring constituents of fermented fish sauces. Journal of the Science of Food and Agriculture, 26, 887. Dwivedi, B. K. (1975). Meat flavor. CRC Critical Reviews Food Technology, 5, 487–535. Fox, P. F., & Wallace, J. M. (1997). Formation of flavour compounds. Advances in Applied Microbiology, 45, 17–85. Fox, P. F., Lucey, J. A., & Cogan, T. M. (1990). Glycolysis and related reactions during cheese manufacture and ripening. Critical Reviews in Food Science and Nutrition, 29, 237–253. Fukami, K., Funatsu, Y., Kawasaki, K., & Watabe, S. (2004). Improvement of fish sauce odor by treatment with bacteria isolated from the fish-sauce mash (moromi) made from frigate mackerel. Journal of Food Science, 69, 45–49. Girand, B., & Durance, T. (2000). Headspace volatiles of sockeye and pink salmon as affected by retort process. Journal of Food Science, 65, 34–39. Giri, A., Osako, K., & Ohshima, T. (2009). Effect of raw materials on the extractive components and taste aspects of fermented. Fish paste – sakana miso. Fisheries Science, 75, 785–796. Guth, H., & Grosch, W. (1994). Identification of the character impact odorants of stewed beef juice by instrumental analyses and sensory studies. Journal of Agricultural and Food Chemistry, 42, 2862–2866. Herbert, R. A., & Shewan, J. M. (1976). Roles played by bacterial and autolytic enzymes in the production of volatile sulphides in spoiling North Sea cod (Gadus morhua). Journal of the Science of Food and Agriculture, 27, 89–94. Itoh, K., & Ebine, H. (1970). Miso no Kagaku to Gijutsu, 198, 19 (in Japanese).

Iwabuchi, S., & Shibasaki, K. (1973). Nippon Shokuhin Kogyo Gakkaishi, 20, 48 (in Japanese). Kim, H. J., Lee, E. J., Shin, O. S., Ji, W. D., Choi, M. R., & Kim, J. K. (1996). Volatile components in the soy sauce manufactured by Bacillus species and fused yeast. Journal of Microbiology and Biotechnology, 6(3), 194–201. Kobayashi, A., & Sugawara, E. (1999). Flavor components of shoyu and miso Japanese fermented soybean seasonings. In F. Shahidi & C. T. Ho (Eds.), Flavor chemistry of ethnic foods (pp. 5–14). New York: Plenum Press. Landaud, S., Helinck, S., & Bonnarme, P. (2007). Formation of volatile sulfur compounds and metabolism of methionine and other sulfur compounds in fermented food. Applied Microbiology and Biotechnology, 10, 253. Lee, J. H., Kang, J. H., & Min, D. B. (2003). Optimization of solid-phase microextraction for the analysis of the head space volatile compounds in kimchi, a traditional Korean fermented vegetable product. Journal of Food Science, 68(3), 844–848. Marilley, L., & Casey, M. G. (2004). Flavours of cheese products: metabolic pathways, analytical tools and identification of producing strains. International Journal of Food Microbiology, 90, 139–159. McIver, R. C., Brooks, R. I., & Reineccius, G. A. (1982). Flavor of fermented fish sauce. Journal of Agricultural and Food Chemistry, 30, 1017. Montel, M. C., Masson, F., & Talon, R. (1998). Bacterial role in flavour development. Meat Science, 49, S111–S123. Nursten, H., & Reineccius, G. (1996). Workshop on current and future problems in flavour research. In A. J. Taylor & D. S. Mottram (Eds.), Flavour science (pp. 459–461). Cambridge, UK: The Royal Society of Chemistry. Saisithi, P., Kasermsarn, B., & Dollar, A. M. (1966). Microbiology and chemistry of fermented fish sauce. Journal of Food Science, 31, 105. Selke, E., Rohwedder, W. K., & Dutton, H. J. (1975). Volatile components from tristearin heated in air. Journal of the American Oil Chemists Society, 52, 232–235. Seo, J. S., Chang, H. G., Ji, W. D., Lee, E. J., Choi, M. R., Kim, H. J., et al. (1996). Aroma components of traditional Korean soy sauce and soybean paste fermented with the same meju. Journal of Microbiology and Biotechnology, 6(4), 278–285. Sink, J. D. (1973). Lipid soluble components of meat, odor and their biochemical origin. Journal of the American Oil Chemists Society, 50, 470–474. Tanaka, T., & Shoji, Z. (1993). Analysis of volatile compounds in the nattofermenting room by gas chromatography-mass spectrometry. Nippon Shokuhin Kogyo Gakkaishi, 40, 656–660. Taylor, A. J., & Mottram, D. S. (1990). Composition and odour of volatiles from autoxidized methyl arachidonate. Journal of the Science of Food and Agriculture, 50, 407–417. Thomas, C. P., Dimick, D. S., & McNeil, J. H. (1971). Sources of flavor in poultry skin. Food Technology, 25(4), 109–115. Van-Chom, T. (1958). The volatile fatty acids of noucnam. In Proceedings, 9th Pacific science congress; Pacific Sc. Assoc. (Vol. 5, p. 135). Bangkok, Thailand. Whistler, R. L., & Daniel, J. R. (1985). Carbohydrates. In O. Fennema (Ed.), Food Chemistry (pp. 69–137). New York, NY: Marcel Dekker Inc.. Yasuhira, H., Itoga, K., & Mochizuki, T. (1974). Reports of the Shinshu-Miso Research Institute, 15, 22.