Characterization of volatile compounds from the reaction of 3-hydroxy- 2-butanone and ammonium sulfide model system

Article abstract of DOI:10.1021/jf980536s

The reactions between 3-hydroxy-2-butanone and ammoniun sulfide at 25, 50, 75, 100, 125, and 150 °C were studied. Four well-known flavor compounds, 2,4,5-trimethyloxazole, 2,4,5-trimethyl-3-oxazoline, 2,4,5-trimethylthiazole, and 2,4,5-trimethyl-3-thiazoline, were identified. Another four interesting intermediate compounds, 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazoline, 2- (1-mercaptoethyl)-2,4,5-trimethyl-3-oxazoline, 2-(1-hydroxyethyl)-2,4,5- trimethyl-3-thiazoline, and 2-(1-mercaptoethyl)-2,4,5-trimethyl-3-thiazoline, were also identified by GC-EIMS and GC-CIMS. All these intermediate compounds were formed at 25 °C. On the other hand, tetramethylpyrazine was the major product with a reaction temperature higher than 100 °C.

Full text of DOI:10.1021/jf980536s

J. Agric. Food Chem. 1999, 47, 245−248
245
Ch a r a cter iza tion of Vola tile Com p ou n d s fr om th e Rea ction of
3-Hyd r oxy-2-bu ta n on e a n d Am m on iu m Su lfid e Mod el System
J unwu Xi,† Tzou-Chi Huang,‡ and Chi-Tang Ho*,†
Department of Food Science, Cook College, Rutgers University, 65 Dudley Road, New Brunswick,
New J ersey 08901-8520, and Department of Food Science and Technology, National Pingtung University of
Science and Technology, 912 Pingtung, Taiwan
The reactions between 3-hydroxy-2-butanone and ammoniun sulfide at 25, 50, 75, 100, 125, and
150 °C were studied. Four well-known flavor compounds, 2,4,5-trimethyloxazole, 2,4,5-trimethyl-
3-oxazoline, 2,4,5-trimethylthiazole, and 2,4,5-trimethyl-3-thiazoline, were identified. Another four
interesting intermediate compounds, 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazoline, 2-(1-mercap-
toethyl)-2,4,5-trimethyl-3-oxazoline, 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-thiazoline, and 2-(1-mer-
captoethyl)-2,4,5-trimethyl-3-thiazoline, were also identified by GC-EIMS and GC-CIMS. All these
intermediate compounds were formed at 25 °C. On the other hand, tetramethylpyrazine was the
major product with a reaction temperature higher than 100 °C.
Keyw or d s: 3-Hydroxy-2-butanone; ammonium sulfide; 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazo-
line; 2-(1-mercaptoethyl)-2,4,5-trimethyl-3-oxazoline; 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-thiazoline;
2-(1-mercaptoethyl)-2,4,5-trimethyl-3-thiazoline
INTRODUCTION
perature, an interesting compound, 2-(1-hydroxyethyl)-
2,4,5-trimethyl-3-oxazoline along with 2,4,5-trimethyl-
oxazole and 2,4,5-trimethyl-3-oxazoline, was isolated
and identified (Shu and Lawrence, 1995; Fu and Ho,
1997). Our objective in the present study was to isolate
and identify the oxazoles, oxazolines, thiazoles, and
thiazolines from the reaction of 3-hydroxy-2-butanone
and the ammonium sulfide model system.
Oxazoles, oxazolines, thiazoles, and thiazolines are
four important heterocyclic aroma compounds contain-
ing nitrogen and oxygen or sulfur atoms. Maga (1981)
has reviewed the occurrence of oxazoles and oxazolines
in a variety of processed food systems. They possess
potent sensory quality at a very low concentration
described as green, sweet, and nutty aroma and have
been identified in coffee (Stofflesma et al., 1968; Vitz-
thum et al., 1975), soy sauce (Nunomura et al., 1978),
wheat (Harding et al., 1978), and cooked beef (Mussinan
et al., 1976).
The occurrence of thiazoles and thiazolines has also
been intensively reviewed (Maga, 1975; Ho and J in,
1985). They are generally described as green, nutty, and
vegetable-like aroma and have been found in heated
foods such as baked potato (Coleman et al., 1981),
roasted peanuts (Ho et al., 1983a), peanut butter (J oo
and Ho, 1997), cocoa butter (Ho et al., 1983b), and fried
chicken (Tang et al., 1983).
It has been proposed that thiazoles may be formed in
processed foods by interaction of R-dicarbonyl com-
pounds, aldehydes, ammonia, and hydrogen sulfide
(Mussinan et al., 1976). In fact, thiazoles and thiazolines
have been identified from the reaction of 2,3-pentanedi-
one, acetaldehyde, ammonia, and hydrogen sulfide
(Takken et al., 1976). Thiazoles have also been found
in various model systems involving either degradation
of glucose in the presence of fragmentation of cysteine
or cystine (Shu et al., 1885a,b) or reaction of these with
reducing sugars (Kato et al., 1973) or furaneol (Shu and
Ho, 1988). In recent studies on the reaction of 3-hy-
droxy-2-butanone with ammonium acetate at low tem-
MATERIALS AND METHODS
Ma ter ia ls. 3-Hydroxy-2-butanone (acetoin) and ammonium
sulfide (20 wt % solution in water) were purchased from
Aldrich Chemical Co. (Milwaukee, WI). Methylene chloride
was obtained from Fisher Scientific Co. (Pittsburgh, PA).
n-Ttridecane was from Alltech Assoc., Inc. (Deerfield, IL).
P r ep a r a tion of Rea ction Mixtu r e. A total of 0.88 g (0.01
mol) of 3-hydroxy-2-butanone and 6.8 mL (0.02 mol) of am-
monium sulfide were mixed into 25 mL of distilled water with
the pH value adjusted to 5.5 using 6 and 1 N HCl. The mixture
was transferred into a 0.3-L Hoke SS-DOT sample cylinder,
and the cylinder was sealed and heated at 25, 50, and 75 °C
for 2 h. One milliliter of 1000 ppm n-tridecane was added into
the reaction mixture as internal standard. Then the reaction
mixture was extracted with a total of 50 mL of methylene
chloride. The combined extract was dried over anhydrous
sodium sulfate and concentrated to a final volume of 1 mL
under a gentle stream of nitrogen gas. One microliter of extract
was injected into the GC.
P r ep a r a tion of th e Solu tion for th e Stor a ge Stu d y. For
the storage study, a series mixture was prepared with 0.01
mol of acetoin (0.88 g) and 0.02 mol of ammonium sulfide (6.8
mL) with the addition of H2O to a total volume of 25 mL. The
pH values of all of the solutions were adjusted to 5.5 with 1
and 6 N HCl solution, and then the tubes were sealed and
stored in a controlled environment incubator shaker (New
Brunswick Scientific Co., New Brunswick, NJ ) with constant
shaking at 25 °C until further use. At the end of the storage
period, internal standard n-tridecane was added and then
extracted with methylene chloride three times (20, 20, and 10
mL). The combined extract was dried over anhydrous sodium
* To whom correspondence should be addressed (fax, 732-
932-8004; e-mail, ho@aesop.rutgers.edu).
† Rutgers University.
‡ National Pingtung University of Science and Technology.
10.1021/jf980536s CCC: $18.00 © 1999 American Chemical Society
Published on Web 12/10/1998

246 J. Agric. Food Chem., Vol. 47, No. 1, 1999
Xi et al.
Ta ble 1. Vola tile Com p ou n d s Id en tified in th e Rea ction
of 3-Hyd r oxy-2-bu ta n on e/Am m on iu m Su lfid e a t 75 °C
no.
compound
1
2
2,4,5-trimethyl-3-oxazoline
2,4,5-trimethyloxazole
3
3-mercaptobutane
4
4,5-dimethylthiazole
5
2,4,5-trimethylthiazole
6
2,4,5-trimethyl-3-thiazoline
7
tetramethylpyrazine
8
9
2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazoline
2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazoline
2-(1-mercaptoethyl)-2,4,5-trimethyl-3-oxazoline
2-(1-mercaptoethyl)-2,4,5-trimethyl-3-oxazoline
2-(1-hydroxyethyl)-2,4,5-trimethyl-3-thiazoline
2-(1-hydroxyethyl)-2,4,5-trimethyl-3-thiazoline
2-(1-mercaptoethyl)-2,4,5-trimethyl-3-thiazoline
2-(1-mercaptoethyl)-2,4,5-trimethyl-3-thiazoline
F igu r e 1. EI mass spectrum of 2-(1-hydroxyethyl)-2,4,5-
trimethyl-3-oxazoline.
10
11
12
13
14
15
sulfate and concentrated to 1 mL under a gentle stream of
N2. One microliter of extract was injected into the GC.
Qu a n tita tion a n d Ch a r a cter iza tion of Vola tile Com -
p ou n d s. Gas Chromatography. GC analysis was accomplished
by using a Varian 3400 gas chromatograph. A fused silica gel
capillary column (30 m × 0.25-mm i.d., film thickness 0.25
µm, DB-1701; J &W Scientific, Folsom, CA) was used to analyze
the volatile compounds. The operating conditions were as
follows: injector and detector temperatures, 250 and 270 °C,
respectively; helium carrier flow rate, 1 mL/min; GC temper-
ature program, 40-260 °C at 3 °C/min followed by an
isothermal hold at 260 °C for 10 min.
F igu r e 2. EI mass spectrum of 2-(1-mercaptoethyl)-2,4,5-
trimethyl-3-oxazoline.
Gas Chromatography-Mass Spectrometry Analysis. EI mass
spectra were obtained using a Hewlett-Packard 5790 gas
chromatograph coupled with a Hewlett-Packard 5970A MSD
detector with electron ionization at 70 eV and an ion source
temperature of 250 °C. The operating conditions were the same
as those used in the GC analysis described above. The data
were recorded and analyzed using Hewlett-Packard MS Chem-
Station data with NIST/EPA/MSDC mass spectral database.
CI mass spectra were performed on a Finnigan ITS-40
Magnum ion trap mass spectrometer coupled with a Varian
3400 gas chromatograph, and reactant gas (methane) was
used. A fused silica gel capillary column (30 m × 0.25-mm i.d.,
film thickness 0.25 µm, DB-5; J &W Scientific, Folsom, CA) was
used. The operating conditions were as follows: injector
temperature, 260 °C; transfer line temperature, 260 °C; helium
carrier flow rate, 1 mL/min; GC temperature program, 60-
260 °C at 6 °C/min followed by an isothermal hold at 260 °C
for 12 min.
F igu r e 3. EI mass spectrum of 2-(1-hydroxyethyl)-2,4,5-
trimethyl-3-thiazoline.
Lawrence (1995) as isomers in their study in which they
were described as mild, yeasty, nutty, and bread-crust-
like aroma.
The mass spectrum of compounds 10 and 11 is
extremely similar with the previous spectra of 2-(1-
hydroxyethyl)-2,4,5-trimethyl-3-oxazoline as shown in
Figure 2 except they have a molecular weight of 173 as
determined by GC-CIMS. They are proposed to be the
syn and anti isomers of 2-(1-mercaptoethyl)-2,4,5-tri-
methyl-3-oxazoline.
Compounds 12 and 13 also have a molecular weight
of 173 as determined by GC-CIMS. However, their
GC-EIMS spectra shown in Figure 3 suggested that
they were sulfur analogues of 2-(1-hydroxyethyl)-2,4,5-
trimethyl-3-oxazoline. These two compounds were pro-
posed to be the syn and anti isomers of 2-(1-hydroxy-
ethyl)-2,4,5-trimethyl-3-thiazoline.
Compounds 14 and 15 have a molecular weight of 189
as determined by GC-CIMS. Their GC-EIMS spectra
shown in Figure 4 suggested that they were sulfur
analogues of compounds 10 and 11. They were proposed
to be the syn and anti isomers of 2-(1-mercaptoethyl)-
2,4,5-trimethyl-3-thiazoline.
RESULTS AND DISCUSSION
Vola t ile Com p ou n d s Id en t ified in t h e 3-Hy-
d r oxy-2-bu ta n on e (Acetoin )/Am m on iu m Su lfid e
Mod el System . Fifteen compounds were tentatively
identified by GC-EIMS at six different temperatures
(25, 50, 75, 100, 125, and 150 °C) as shown in Table 1.
They are 2,4,5-trimethyl-3-oxazoline (1), 2,4,5-trimethyl-
oxazole (2), 3-mercaptobutane (3), 4,5-dimethylthiazole
(4), 2,4,5-trimethylthiazole (5), 2,4,5-trimethyl-3-thia-
zoline (6), tetramethylpyrazine (7), 2-(1-hydroxyethyl)-
2,4,5-trimethyl-3-oxazoline (8, 9), 2-(1-mercaptoethyl)-
2,4,5-trimethyl-3-oxazoline (10, 11), 2-(1-hydroxyethyl)-
2,4,5-trimethyl-3-thiazoline(12,13),and2-(1-mercaptoethyl)-
2,4,5-trimethyl-3-thiazoline (14, 15).
Compounds 8 and 9 have a molecular weight of 157
as determined by GC-CIMS. They have the same GC-
EIMS spectra as shown in Figure 1. This mass spectrum
matches well with the spectral data published previ-
ously by Shu and Lawrence (1995) and Fu and Ho
(1997). They were, therefore, identified as syn and anti
isomers of 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazo-
line. These two peaks were also observed by Shu and
F or m a tion of 2-(1-Hyd r oxyeth yl)-2,4,5-tr im eth yl-
3-oxa zolin e, 2-(1-Mer ca p toeth yl)-2,4,5-tr im eth yl-3-
oxa zolin e, 2-(1-H yd r oxyet h yl)-2,4,5-t r im et h yl-3-
th iazolin e, an d 2-(1-Mer captoeth yl)-2,4,5-tr im eth yl-
3-th ia zolin e. Six temperatures (25, 50, 75, 100, 125,

Reaction of 3-Hydroxy-2-butanone and Ammonium Sulfide
J. Agric. Food Chem., Vol. 47, No. 1, 1999 247
Ta ble 2. Qu a n tita tion of Vola tile Com p ou n d s Gen er a ted fr om 3-Hyd r oxy-2-bu ta n on e/Am m on iu m Su lfid e Mod el System
quantity (mg/g of acetoin)
compound
25 °C
50 °C
75 °C
100 °C
125 °C
150 °C
2,4,5-trimethyl-3-oxazoline
2,4,5-trimethyloxazole
3-mercaptobutane
4,5-dimethylthiazole
2,4,5-trimethylthiazole
2,4,5-trimethyl-3-thiazoline
tetramethylpyrazine
2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazoline
2-(1-mecaptoethyl)-2,4,5-trimethyl-3-oxazoline
2-(1-hydroxyethyl)-2,4,5-trimethyl-3-thiazoline
2-(1-mecaptoethyl)-2,4,5-trimethyl-3-thiazoline
0.284
0.106
0.698
0.000
0.093
0.000
0.000
22.441
0.491
0.174
0.300
0.101
0.113
0.327
0.000
0.086
0.018
0.080
21.394
0.710
1.230
1.849
0.175
0.111
0.249
0.000
0.186
0.127
1.180
10.474
0.144
2.356
3.302
0.346
0.142
0.172
0.000
0.596
0.325
9.776
3.258
0.154
1.270
3.052
0.300
0.516
1.002
0.070
2.447
1.503
58.540
0.264
0.034
1.532
2.161
0.019
1.846
1.329
0.077
3.765
1.686
85.909
0.700
0.048
0.769
1.809
Sch em e 1. F or m a tion of Oxa zolin es a n d Th ia zolin es in th e Mod el Rea ction of 3-Hyd r oxy-2-bu ta n on e a n d
Am m on iu m Su lfid e System
and 150 °C) were investigated in this model system.
Quantitation of volatile compounds is summarized in
Table 2. The data show that at and below 75 °C, 2-(1-
hydroxyethyl)-2,4,5-trimethyl-3-oxazoline was the most
dominant product, whereas tetramethylpyrazine was
the major product at or higher than 100 °C. These
results agreed with the study of Fu and Ho (1997) in
which it was observed that at a temperature below 25
°C, 2-(1-hydroxyethyl)-2,4,5-trimethyl-3-oxazoline was
the major product, while at a temperature higher than
85 °C, tetramethylpyrazine was formed predominantly.
In the present study, due to the presence of hydrogen
sulfide, the formation of tetramethylpyrazine was re-
duced at high temperatures. On the other hand, ox-
azoles, thiazoles, and thiazolines were found to be more
stable despite the dramatic decrease in oxazoline pro-
duction.
tored. 2-(1-Hydroxyethyl)-2,4,5-trimethyl-3-oxazoline de-
creased dramatically during the early stage of the
storage, while tetramethylpyrazine increased over the
entire period of storage time (Figure 5). This study also
supported the mechanism proposed by Shu and Lawrence
(1995) who observed that, during their 3-week storage
study of the mixture of acetoin and diammonium
hydrogen phosphate, 2-(1-hydroxyethyl)-2,4,5-trimethyl-
3-oxazoline was readily formed during the early stage
of the reaction while tetramethylpyrazine was increased
more gradually during the final 10-21-day period. The
pathway for the formation of 2-(1-hydroxyethyl)-2,4,5-
trimethyl-3-oxazoline may be reversible, so that 2-(1-
hydroxyethyl)-2,4,5-trimethyl-3-oxazoline initially formed
is able to be converted back to R-hydroxyimine, which
in turn was tautomerized to R-aminoketone, so that the
formation of tetramethylpyrazine could be followed (Shu
and Lawrence, 1995).
Sta bility Stu d y of 3-Hyd r oxy-2-bu ta n on e/Am -
m on iu m Su lfid e Mod el System . In our 96-day stor-
age study of the mixture of 3-hydroxy-2-butanone and
ammonium sulfide. Both 2-(1-hydroxyethyl)-2,4,5-tri-
methyl-3-oxazoline and tetramethylpyrazine were moni-
P r op osed Rea ction Sch em e for Vola tile Com -
p ou n d s F or m ed in th e 3-Hyd r oxy-2-bu ta n on e/Am -
m on iu m Su lfid e Mod el System . Scheme 1 summa-
rizes the reaction for the formation of oxazolines and

248 J. Agric. Food Chem., Vol. 47, No. 1, 1999
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Received for review May 20, 1998. Revised manuscript received
October 16, 1998. Accepted October 27, 1998.
J F980536S