Retro-aldol and redox reactions of Amadori compounds: Mechanistic studies with variously labeled D-[13C]glucose

Article abstract of DOI:10.1021/jf9502921

Oxidation-reduction reactions necessary to justify many of the products observed in Maillard model systems are usually attributed to molecular oxygen and the so-called reductons. The proline specific 1-(1′-pyrrolidinyl)-2-propanone and 1-(1′-pyrrolidinyl)-2-butanone are such compounds that require reduction steps to justify their formation. Experimental evidence using glucose separately labeled at ¹³C1, ¹³C2, ¹³C3, ¹³C4, ¹³C5, and ¹³C6 indicates that 1-(1′-pyrrolidinyl)-2-propanone is formed by two related pathways, initiated by a retro-aldol cleavage of proline Amadori compound at C3-C4, and 1-(1′-pyrrolidinyl)-2-butanone is formed by three pathways, one initiated by a retro-aldol reaction at C2-C3 of the 1-(prolino)-1-deoxy-4-hexosulose (an isomer of Amadori product formed by carbonyl migration) and two others by similar retro-aldol reactions at C4-C5 from both 3-deoxyglucosone and 1-(prolino)-1,4-dideoxy-2,3-hexodiulose. All of the proposed mechanisms require reduction steps for the formation of the target compounds. Model studies have indicated that reductions in Maillard systems can be effected by three pathways: through hydride transfer from formic acid; through cyclic dimerization of α-hydroxy carbonyl compounds followed by electrocyclic ring opening to produce oxidation/reduction products; and by disproportionation of enediols with α-dicarbonyl compounds through double proton transfer.

Full text of DOI:10.1021/jf9502921

672
J. Agric. Food Chem. 1996, 44, 672−681
Retr o-Ald ol a n d Red ox Rea ction s of Am a d or i Com p ou n d s:
Mech a n istic Stu d ies w ith Va r iou sly La beled D-[13C]Glu cose
Alexis Huyghues-Despointes and Varoujan A. Yaylayan*
Department of Food Science and Agricultural Chemistry, McGill University,
21,111 Lakeshore, Ste. Anne de Bellevue, Quebec, Canada H9X 3V9
Oxidation-reduction reactions necessary to justify many of the products observed in Maillard model
systems are usually attributed to molecular oxygen and the so-called reductons. The proline specific
1-(1′-pyrrolidinyl)-2-propanone and 1-(1′-pyrrolidinyl)-2-butanone are such compounds that require
reduction steps to justify their formation. Experimental evidence using glucose separately labeled
at 13C1, 13C2, 13C3, 13C4, 13C5, and 13C6 indicates that 1-(1′-pyrrolidinyl)-2-propanone is formed by
two related pathways, initiated by a retro-aldol cleavage of proline Amadori compound at C3-C4,
and 1-(1′-pyrrolidinyl)-2-butanone is formed by three pathways, one initiated by a retro-aldol reaction
at C2-C3 of the 1-(prolino)-1-deoxy-4-hexosulose (an isomer of Amadori product formed by carbonyl
migration) and two others by similar retro-aldol reactions at C4-C5 from both 3-deoxyglucosone
and 1-(prolino)-1,4-dideoxy-2,3-hexodiulose. All of the proposed mechanisms require reduction steps
for the formation of the target compounds. Model studies have indicated that reductions in Maillard
systems can be effected by three pathways: through hydride transfer from formic acid; through
cyclic dimerization of R-hydroxy carbonyl compounds followed by electrocyclic ring opening to produce
oxidation/reduction products; and by disproportionation of enediols with R-dicarbonyl compounds
through double proton transfer.
Keyw or d s: Amadori; decomposition mechanisms; oxidation; reduction; Maillard reaction;
13C-labeled glucoses; proline; morpholine
INTRODUCTION
basic media. Generation of these fragments in sugar
amino acid or amine mixtures has been studied exten-
sively. Hayashi and Namiki (1980) have shown that
sugar alkylamine mixtures can generate glyoxal and
pyruvaldehyde as their diimine derivatives at a pre-
Amadori stage if the mixtures are heated for 4 min. The
formation of these C2 and C3 sugar fragments increased
with pH, indicating a base-catalyzed retro-aldol reac-
tion. Morita and Takagi (1986) demonstrated, by trap-
ping R-dicarbonyl compounds with o-phenylenediamine
(OPD), that D-glucose when incubated at 100 °C (pH 10)
for 1 h can generate six R-dicarbonyl derivatives con-
taining C6, C5, C4, and C3 fragments and almost 60% of
the dicarbonyl generated was pyruvaldehyde, indicating
the preference of D-glucose to undergo retro-aldol cleav-
age at the C3-C4 position, possibly from a 3-deoxyglu-
cosone intermediate which was estimated to be 14% of
the total R-dicarbonyls detected by OPD. Recently,
Huber and Ledl (1990) were able to isolate and char-
acterize 1-deoxy- and 3-deoxyglucosones and 1-amino-
1,4-dideoxy-2,3-hexodiulose from heated Amadori prod-
ucts in water at 100 °C by trapping these reactive
intermediates with OPD as their quinoxaline deriva-
tives.
The use of 13C-enriched sugars for the elucidation of
reaction mechanisms concerning the Maillard reaction
has been well documented in the literature (Tressl et
al., 1993). However, in these studies, sugars enriched
at positions other than C1 are seldom used. To study
the mechanism of formation of short chain compounds
arising from the Maillard reaction, the use of variously
enriched sugars (at positions other than C1) is neces-
sary. Weenen et al. (1994) used D-[1-13C]glucose, D-[2-
13C]glucose, and D-[1-13C]fructose in the presence of
amino acid to study the decomposition reactions and the
formation of pyrazines, which was proposed to proceed
through dimerization of C3 and C4 units of R-aminocar-
Amadori products (1), deoxyglucosones, and other
sugar derivatives formed under Maillard reaction condi-
tions can undergo retro-aldol reactions, especially under
basic conditions, to produce more reactive C2, C3, C4 and
C5 sugar fragments containing R-hydroxy carbonyl and
R-dicarbonyl moieties that can react more efficiently
with amines than C6 sugars. R-Dicarbonyls are gener-
ally much more reactive toward nucleophiles than
simple carbonyls. Amadori rearrangement products
(ARP) under basic conditions can generate acetic acid,
glyceraldehyde, pyruvaldehyde, and other lower sugars
in addition to free amino acid. The process of generating
more reactive C2, C3, and C4 carbonyl species from
Amadori products becomes important in vivo where less
reactive amino sites such as nucleotide bases can now
be modified by such species, especially in diabetic
patients, to produce DNA-linked advanced glycation
products (Papoulis et al., 1995). Similarly, if glycated
proteins can generate more reactive sugar fragments
(Zyzak et al., 1995), especially if the glycation sites are
close to the basic amino acids, then, due to the proximity
of these species to the target protein, the smaller
fragments can further accelerate their post-translational
modification. In addition, C3 sugar fragments have been
implicated in generation of free radicals, browning, and
weak chemiluminescence in model systems. The reac-
tion of pyruvaldehyde with methylamine, for example,
generates free radicals and chemiluminescence (Namiki
et al., 1993). Smaller sugar fragments can be formed
during pre- or post-Amadori stages under Maillard
reaction conditions or from sugar solutions alone in
* Author to whom correspondence should be ad-
dressed [telephone (514) 398-7918; fax (514) 398-7977;
e-mail czd7@MusicA.McGill.CA].
0021-8561/96/1444-0672$12.00/0
© 1996 American Chemical Society

Retro-Aldol and Redox Reactions of Amadori Compounds
J. Agric. Food Chem., Vol. 44, No. 3, 1996 673
Ta ble 1. Ma ss Sp ectr om etr ic Da ta
1-(1′-p yr r olid in yl)-2-p r op a n on e (2)
from pyrolysis of Amadori proline: 127 (2), 85 (06), 84 (100), 82 (2), 56 (07), 55 (20), 54 (3), 43 (07), 42 (43), 41 (07)
from acetol/ pyrrolidine reaction mixture: 127 (2), 85 (07), 84 (100), 82 (2), 56 (07), 55 (19), 54 (3), 43 (07), 42 (42), 41 (07)
from pyruvaldehyde/proline/formic acid reaction mixture: 127 (1), 85 (06), 84 (100), 82 (1), 56 (06), 55 (19), 54 (2), 43 (07), 42 (43), 41 (06)
from pyrolysis of 1-(prolino)-1-deoxy-D-glyceraldehyde: 127 (2), 85 (07), 84 (100), 82 (2), 56 (07), 55 (18), 54 (3), 43 (07), 42 (41), 41 (08)
from glycerladehyde/pyrrolidine reaction mixture: 127 (2), 85 (06), 84 (100), 82 (2), 56 (06), 55 (18), 54 (3), 43 (06), 42 (39), 41 (06)
from dihydroxyacetone/pyrrolidine reaction mixture: 127 (2), 85 (06), 84 (100), 82 (2), 56 (06), 55 (18), 54 (3), 43 (06), 42 (39), 41 (06)
literature data:a 127 (2), 85 (06), 84 (100), 82 (2), 56 (06), 55 (18), 54 (3), 43 (07), 42 (31), 41 (04)
2-h yd r oxy-1-(1′-p yr r olid iyl)-1-bu ten -3-on e (3)
from Amadori proline: 155 (72), 138 (10), 137 (12), 126 (8), 112 (96), 84 (100), 83 (27), 70 (74), 57 (14), 56 (30), 55 (41), 43 (66), 42 (50)
literature data:a 155 (67), 138 (09), 137 (11), 126 (7), 112 (97), 84 (100), 83 (26), 70 (64), 57 (11), 56 (24), 55 (36), 43 (52), 42 (55)
2-h yd r oxy-1-(N-m or p h olin o)-1-bu ten -3-on e
from Amadori morpholine: 172 (2), 171 (21), 127 (8), 126 (2), 112 (12), 100 (100), 98 (7), 86 (11), 85 (8), 84 (5), 83 (4), 70 (35), 57 (5), 56
(17), 55 (18), 43 (20), 42 (24), 41 (17)
3-(1′-p yr r olid in yl)-2-bu ta n on e (4)
from 3-hydroxy-2-butanone/proline reaction mixture: 141 (0.4), 99 (8), 98 (100), 70 (4), 69 (7), 68 (2), 56 (27), 55 (9), 54 (4), 42 (7), 41 (8)
from pyrolysis of proline Amadori: 141 (0.2), 99 (8), 98 (100), 70 (4), 69 (8), 68 (2), 56 (30), 55 (9), 54 (4), 42 (7), 41 (9)
1-(N-m or p h olin o)-2-p r op a n on e (6)
from pyrolysis of Amadori morpholine: 143 (2), 115 (4), 101 (6), 100 (100), 86 (2), 85 (2), 70 (11), 57 (6), 56 (33), 55 (2), 54 (2),
43 (11), 42 (22), 41 (1)
from morpholine/acetol reaction mixture: 143 (2), 115 (1), 101 (7), 100 (100), 86 (1), 85 (2), 70 (12), 57 (3), 56 (32), 55 (2),
54 (2), 43 (11), 42 (20), 41 (8)
d i-TMS of 1-(p r olin o)-1-d eoxy-D-glycer a ld eh yd e (7)
from pyrolysis of Amadori proline: 332 (1), 331 (3), 316 (6), 288 (2), 258 (1), 216 (3), 215 (10), 214 (56), 202 (5), 201 (16), 200 (100),
189 (2), 186 (8), 174 (1), 173 (2), 172 (13), 170 (1), 156 (1), 147 (1), 142 (11), 130 (3), 129 (17), 103 (18), 96 (8), 83 (7), 82 (8), 75 (11), 74 (4),
73 (40), 70 (4), 68 (2), 61 (2), 59 (4), 58 (2), 55 (14), 54 (3), 45 (7), 43 (3), 42 (4)
1-(N-m or p h olin o)-1-d eoxy-D-glycer a ld eh yd e
from pyrolysis of Amadori morpholine: 159 (0.5), 129 (0.2), 114 (2), 101 (6), 100 (100), 87 (2), 86 (2), 85 (2), 72 (3), 70 (8), 57 (6), 56 (22),
55 (3), 44 (5), 43 (13), 42 (16)
1, 2-(1′,1′-d ip yr r olid in yl)-1-p r op en e (10)
from pyrolysis of glyceraldehyde/proline mixture: 181 (14), 180 (100), 179 (2), 165 (3), 152 (8), 151 (16), 150 (3), 139 (9), 138 (11), 137
(47), 136 (8), 135 (2), 134 (3), 125 (9), 124 (20), 123 (70), 122 (23), 121 (3), 120 (3), 112 (5), 111 (61), 110 (68), 109 (47), 108 (20), 107 (2),
106 (2), 98 (11), 97 (29), 96 (56), 95 (15), 94 (8), 93 (2), 85 (3), 84 (40), 83 (70), 82 (27), 81 (20), 80 (10), 79 (3), 71 (4), 70 (38), 69 (25), 68
(40), 67 (9), 66 (3), 56 (17), 55 (33), 54 (19), 53 (7), 52 (2), 44 (3), 43 (9), 42 (42), 41 (43)
from acetol/2× pyrrolidin reaction mixture: 181 (13), 180 (100), 179 (7), 165 (4), 152 (7), 151 (15), 150 (4), 139 (8), 138 (9), 137 (42), 136
(7), 135 (2), 134 (2), 125 (8), 124 (18), 123 (63), 122 (22), 121 (2), 120 (3), 112 (5), 111 (61), 110 (64), 109 (46), 108 (20), 107 (2), 106 (2), 98
(10), 97 (27), 96 (54), 95 (15), 94 (9), 93 (2), 85 (3), 84 (43), 83 (72), 82 (26), 81 (20), 80 (10), 79 (3), 71 (5), 70 (39), 69 (25), 68 (40), 67 (9),
66 (3), 56 (17), 55 (35), 54 (21), 53 (8), 52 (3), 44 (3), 43 (10), 42 (47), 41 (48)
1,2-(N,N-d im or p h olin o)-1-p r op en e
from Amadori morpholine: 213 (13), 212 (100), 167 (3), 155 (5), 154 (35), 153 (32), 140 (7), 139 (36), 138 (7), 129 (7), 128 (6), 127 (45), 126
(26), 125 (13), 124 (7), 123 (8), 114 (5), 113 (5), 112 (27), 111 (4), 110 (11), 109 (10), 108 (4), 100 (35), 98 (9), 97 (18), 96 (66), 95 (32), 94 (7),
86 (14), 85 (11), 84 (19), 83 (16), 82 (23), 81 (6), 80 (6), 70 (18), 69 (25), 68 (22), 67 (9), 58 (3), 57 (9), 56 (28), 55 (30), 54 (19), 45 (9), 44 (8),
43 (19), 42 (41), 41 (36)
from acetol/2× morpholine reaction mixture: 213 (13), 212 (100), 167 (2), 155 (5), 154 (36), 153 (33), 140 (8), 139 (38), 138 (6), 129 (7),
128 (6), 127 (48), 126 (27), 125 (13), 124 (6), 123 (8), 114 (7), 113 (5), 112 (29), 111 (4), 110 (11), 109 (10), 108 (4), 100 (38), 98 (11), 97 (20), 96
(70), 95 (33), 94 (8), 86 (15), 85 (11), 84 (21), 83 (17), 82 (26), 81 (6), 80 (6), 70 (19), 69 (27), 68 (24), 67 (9), 58 (4), 57 (10), 56 (32), 55 (33),
54 (19), 45 (10), 44 (12), 43 (20), 42 (47), 41 (40)
a
Tressl et al. (1993).
MATERIALS AND METHODS
bonyl compounds arising from the interaction of R-di-
carbonyls with amino acids through Strecker degrada-
All reagents and chemicals were purchased from Aldrich
Chemical Co. (Milwaukee, WI). [1-13C]-D-Glucose, [2-13C]-D-
glucose, [6-13C]-D-glucose, and [2-13C]-D-ribose were also pur-
chased from Aldrich. [3-13C]-D-Glucose, [4-13C]-D-glucose, and
[5-13C]-D-glucose were purchased from ICON Services Inc.
(Summit, NJ ). Melting points were determined on a Fischer
melting point apparatus and are uncorrected. 13C NMR
spectra were recorded at 125.8 MHz using dioxane as external
standard (δ 67.4) on a Bruker AMX500 instrument. Infrared
spectra were recorded in CaF2 IR cells on a Nicolet 8210
Fourier-transform spectrometer equipped with a deuterated
triglycine sulfate (DTGS) detector. The synthesis of Amadori-
proline was performed according to published procedures
(Vernin et al., 1992).
Syn th esis of 1-(P r olin o)-1-d eoxy-D-glycer a ld eh yd e (7).
D-Glyceraldehyde (2.0 g, 0.02 mol) and l-proline (9.2 g, 0.04
mol) were mixed in methanol (40.0 mL), stirred at room
temperature for 3 h, and stored at 4 °C for 15 h. The resulting
precipitate was filtered and crystallized from water/methanol
(1:1 v/v): yield 17.5% (0.67 g, 0.0035 mol); mp 111-112 °C;
νmax 270, 210 nm; FTIR (methanol) 1743 (CdO), 1622 cm-1
(COO-); 13C NMR (D2O) δ 206.84 (CdO), 176.33 (COO-),
72.87 (C-2′ proline), 68.51 (C-5′ proline), 62.76 (CH2OH), 58.77
(-CH2N), 31.22 (C-3′ proline), 25.69 (C-4′ proline); EIMS, di-
TMS derivative, m/ z (relative intensity) 332 (1), 331 (3), 316
tion (Weenen et al., 1994). These authors proposed a
mechanism consistent with the 13C distribution, accord-
ing to which the sugars first underwent â-elimination
to form various deoxyosones which then, through retro-
aldol cleavages at C3-C4, C2-C3, or C5-C4, formed
pyruvaldehyde and 1-hydroxy-2,3-butanedione. To un-
derstand and confirm the mechanism of formation of
C3 and C4 units in the Maillard reaction and to
circumvent their dimerization to form pyrazines, proline
was used as the model amino acid, which is known to
form volatile short chain pyrrolidine derivatives related
to pyruvaldehyde and 1-hydroxy-2,3-butanedione. Fur-
thermore, D-glucoses 13C-enriched at C1, C2, C3, C4, C5,
and C6 positions were systematically reacted with
proline to confirm the origin of C3 and C4 units and to
monitor the occurrence, if any, of chain lengthening
through aldol condensations that could be detected by
observing the scrambling of labels. However, it should
be recognized that more than one mechanism can be
compatible with the observed distribution patterns of
the labeled products.

674 J. Agric. Food Chem., Vol. 44, No. 3, 1996
Huyghues-Despointes and Yaylayan
Sch em e 1. P a th w a ys of Retr o-Ald ol Rea ction s of P r olin e Am a d or i Com p ou n d a n d P er cen t Distr ibu tion of
P r od u cts in Ea ch P a th w a ya
a
[H] ) reductions; RA [x,y] ) retro-aldol cleavage at Cx-Cy. Carbon numbering indicates original D-glucose carbon locations.
(6), 288 (2), 258 (1), 216 (3), 215 (10), 214 (56), 202 (5), 201
(16), 200 (100), 189 (2), 186 (8), 174 (1), 173 (2), 172 (13), 170
(1), 156 (1), 147 (1), 142 (11), 130 (3), 129 (17), 103 (18), 96
(8), 83 (7), 82 (8), 75 (11), 74 (4), 73 (40), 70 (4), 68 (2), 61 (2),
59 (4), 58 (2), 55 (14), 54 (3), 45 (7), 43 (3), 42 (4).
RESULTS AND DISCUSSION
A. Retr o-Ald ol Rea ction s a n d F or m a tion of C2,
C3, a n d C4 F r a gm en t s. 1-(1′-Pyrrolidinyl)-2-pro-
panone (2), 2-hydroxy-1-(1′-pyrrolidiyl)-1-buten-3-one
(3), N-acetylpyrrolidine, and 1-(1′-pyrrolidinyl)-2-bu-
tanone (4) are C2, C3, and C4 sugar adducts of pyrroli-
dine that can be detected in l-proline/D-glucose heated
mixtures (Tressl et al., 1993) and also in the pyrolysis
products of l-proline/D-glucose and proline Amadori
compound (1) (Huyghues-Despointes et al., 1994), in
relatively good yields. Understanding the mechanism
of their formation can elucidate the post- and pre-
Amadori fragmentation pathways of sugar moieties into
smaller fragments and their chemical reactivity. Tressl
et al. (1993) using D-[13C1]glucose and proline demon-
strated by GC/MS that 60% of the 1-(1′-pyrrolidinyl)-
2-propanone formed was unlabeled and 40% was labeled
at the C3 position of the propanone moiety, indicating
at least two different pathways of formation. Huy-
ghues-Despointes et al. (1994) studied the effect of
pyrolysis temperature on the yield of 1-(1′-pyrrolidinyl)-
2-propanone (2) from the proline Amadori product and
found that the temperature vs yield relation had two
maxima, one at 200 °C and another at 300 °C, which
confirms the presence of more than one mechanism of
formation, each having different energy requirements.
In addition, comparison of the relative yields of 1-(1′-
pyrrolidinyl)-2-propanone from the Amadori product at
the same temperature (250 °C) but under different
P yr olysis/GC/MS An a lysis. A Hewlett-Packard GC/mass
selective detector (5890 GC/5971B MSD) interfaced to a CDS
Pyroprobe 2000 unit was used for the Py/GC/MS analysis. Two
modes of sample introduction were used: (a) a 1 mg equivalent
of sample dissolved in methanol/water (70:30) was applied on
the platinum filament with a total heating time (THT) of 10 s
or (b) 1-5 mg solid samples (13C-labeled or unlabeled) were
introduced inside the quartz tube (0.3 mm thickness) and
plugged with quartz wool and inserted inside the coil probe
with a THT of 20 s. The GC column flow rate was 0.8 mL/
min for a split ratio of 92:1 and a septum purge of 3 mL/min.
The Pyroprobe interface was set at the temperature at which
the sample was to be pyrolyzed, and the Pyroprobe was set at
the desired temperature at a rate of 50 °C/ms. Capillary direct
MS interface temperature was 180 °C; ion source temperature
was 280 °C. The ionization voltage was 70 eV, and the electron
multiplier was 1494 V. The mass range analyzed was 20-
350 amu. The column was a fused silica DB-5 column (30 m
length × 0.25 mm i.d.; 0.25 µm film thickness; Supelco, Inc.).
The column initial temperature was -5 °C for 3 min and was
increased to 50 °C at a rate of 30 °C/min; immediately the
temperature was further increased to 270 °C at a rate of 8
°C/min and kept at 270 °C for 5 min. Products that were not
found in the mass spectral libraries were tentatively identified
by comparison with literature mass spectral data and/or by
comparison of the mass spectra produced from their heated
precursors as listed in Table 1.

Retro-Aldol and Redox Reactions of Amadori Compounds
J. Agric. Food Chem., Vol. 44, No. 3, 1996 675
tion of 13C labels in 1-(1′-pyrrolidinyl)-2-propanone (2),
2-hydroxy-1-(1′-pyrrolidiyl)-1-buten-3-one (3), N-acetyl-
pyrrolidine, and 1-(1′-pyrrolidinyl)-2-butanone (4) gen-
erated from the pyrolysis of proline with D-glucoses
separately 13C-labeled at C1, C2, C3, C4, C5, and C6.
Careful analysis of the mass spectra of 2, 3, 4, and
N-acetylpyrrolidine from all 13C-labeling experiments
has indicated that C3 fragments are produced by a retro-
aldol reaction at C3-C4 (designated RA [3,4]) of Ama-
dori compound (pathway A, Scheme 1) and C2 and C4
fragments are produced simultaneously by retro-aldol
cleavage at C4-C5 (RA [4,5]) of both 1-prolino-1,4-
dideoxy-2,3-hexodiulose (4D) and 1-deoxyglucosone (1DG)
(pathways B and C, Scheme 1). However, the major
retro-aldol cleavage to produce C2 and C4 fragments is
initiated at C2-C3 (RA [2,3]) from the Amadori com-
pound, after a carbonyl migration (pathway D, Scheme
1). In principle, similar retro-aldol cleavage (RA [2,3])
at a pre-Amadori stage can occur from the free D-glucose
and/or glycosylamine (pathway E, Scheme 1). Higher
relative yields of C4 products obtained from the pyrolysis
of pure Amadori compounds relative to D-glucose/L-
proline mixtures indicates the importance of pathway
D relative to pathway E.
Ta ble 2. P er cen t Ch r om a togr a p h ic P ea k Ar ea s of
Com p ou n d s Id en tified by P y/GC/MS fr om Mor p h olin e
Am a d or i Com p ou n d P yr olyzed for 20 s a t 250 °C
tR (min)
compound
area %
7.1
8.9
2,3-butanedione
acetol
0.5
0.7
9.8
11.7
15.1
acetic acid
morpholine
21.6
32.6
0.2
2,5-dimethyl-4-hydroxy-2,3-dihydro-
3-furanone
16.4
16.8
1-(N-morpholino)-2-propanonea (5)
2,3-dihydro-3,5-dihydroxy-6-methyl-
4(H)-pyran-4-one
10.1
1.9
19.3
22.9
1-(N-morpholino)-1-deoxy-D-glyceraldehydeb
2-hydroxy-1-(N-morpholino)-1-buten-
3-oneb (6)
0.3
10.1
23.8
1,2-(N,N-dimorpholino)-1-propenea
1.9
total
79.8
a
Suggested structures based on mass spectrometric fragmenta-
tion patterns and analysis of reaction mixture of morpholine with
acetol which produces products having the same fragmenation as
the corresponding peaks in the morpholine Amadori compound.
b Suggested structures based on mass spectrometric fragmentation
patterns only.
pyrolysis conditions (one in a quartz tube which favors
bimolecular interactions and the other on a ribbon
probe) have indicated that the formation of 1-(1′-
pyrrolidinyl)-2-propanone (2) mainly occurs through
bimolecular interactions of fragments cleaved from
Amadori product (Huyghues-Despointes et al., 1994).
Detailed studies to elucidate the different pathways
and mechanism of formation of C2, C3, and C4 sugar
fragments and their derivatives in model systems
require the use of D-glucose labeled at different carbons
to accurately assign the position in the final products.
Quartz tube Py/GC/MS is especially suited to perform
small scale reactions without the need to isolate or
extract the reaction mixture that leads to loss of
valuable isotopically labeled products. In the present
study, Py/GC/MS was used to investigate the distribu-
Mechanism of Formation of C3 FragmentssPathway
A. Under the basic conditions of Amadori products
derived from secondary amines, retro-aldol reactions
become important relative to Amadori products derived
from primary amines. 1-(1′-Pyrrolidinyl)-2-propanone
(2) and 2-hydroxy-1-(1′-pyrrolidinyl)-1-buten-3-one (3)
could be considered products of such reactions in proline
Amadori compound 1. Corresponding morpholino (sec-
ondary amine) derivatives 5 and 6 (see Tables 1 and 2)
are similarly observed in the pyrolysis products of
morpholine Amadori compound as major peaks. In
addition, 5 was formed from a reaction mixture of acetol
and morpholine as the only major product. Retro-aldol
reactions leading to these products and carbon-nitrogen
cleavage reaction to form proline and 2,3-dihydro-3,5-
Ta ble 3. Ma ss Sp ectr om etr ic Da ta of Selected P r od u cts of th e Rea ction of 13C-En r ich ed D-Glu cose w ith P r olin e
N-Acetylpyrrolidine
D-glucose
114 (4.4), 113 (54.3), 112 (3.2), 86 (1.2), 85 (19.6), 72 (1.8), 71 (8.0), 70 (77.0), 69 (2.0), 68 (5.1), 56 (6.7), 55 (7.6),
44 (3.6), 43 (100), 42 (13.7), 41 (12.9)
D-[1-13C]glucose
D-[2-13C]glucose
D-[3-13C]glucose
D-[4-13C]glucose
D-[5-13C]glucose
D-[6-13C]glucose
114 (53.5), 113 (19.3), 112 (0.4), 86 (18.4), 85 (6.4), 72 (1.1), 71 (9.9), 70 (99.1), 69 (2.5), 68 (6.2), 56 (8.6), 55 (8.8),
44 (33.0), 43 (100), 42 (14.1), 41 (16.3)
114 (55.1), 113 (20.2), 112 (3.1), 86 (19.4), 85 (6.1), 72 (1.2), 71 (10.5), 70 (97.8), 69 (2.1), 68 (6.0), 56 (4.1), 55 (8.7),
44 (31.7), 43 (100), 42 (14.6), 41 (16.0)
114 (6.2), 113 (51.0), 112 (3.1), 86 (1.9), 85 (18.6), 72 (1.7), 71 (7.9), 70 (75.4), 69 (1.8), 68 (5.0), 56 (6.6), 55 (7.4),
44 (4.5), 43 (100), 42 (13.2), 41 (13.1)
114 (5.8), 113 (47.3), 112 (2.2), 86 (1.7), 85 (17.8), 72 (1.2), 71 (7.5), 70 (74.4), 69 (1.5), 68 (4.7), 56 (6.2), 55 (7.3),
44 (4.1), 43 (100), 42 (13.3), 41 (13.3)
114 (12.0), 113 (42.3), 112 (2.8), 86 (4.1), 85 (15.5), 72 (1.5), 71 (7.7), 70 (75.4), 69 (1.8), 68 (5.1), 56 (6.0), 55 (7.8),
44 (9.1), 43 (100), 42 (13.6), 41 (13.5)
114 (14.3), 113 (50.0), 112 (3.4), 86 (4.6), 85 (17.5), 72 (1.6), 71 (8.1), 70 (81.2), 69 (1.6), 68 (5.3), 56 (6.3), 55 (7.2),
44 (9.6), 43 (100), 42 (13.4), 41 (12.9)
1-(1′-Pyrrolidinyl)-2-propanone (2)
D-glucose
128 (0.2), 127 (2.0), 85 (06.4), 84 (100), 82 (2.2), 56 (6.0), 55 (17.8), 54 (2.4), 44 (0.4), 43 (6.3), 42 (38.2), 41 (5.5)
128 (1.3), 127 (1.4), 85 (13.6), 84 (100), 82 (2.2), 56 (7.6), 55 (19.6), 54 (2.6), 44 (2.6), 43 (7.8), 42 (42.1), 41 (6.0)
128 (1.4), 127 (1.3), 85 (06.4), 84 (100), 82 (2.1), 56 (6.0), 55 (16.8), 54 (2.3), 44 (2.8), 43 (4.0), 42 (34.8), 41 (5.0)
128 (2.6), 127 (2.2), 85 (100), 84 (99.2), 82 (2.2), 56 (14.8), 55 (28.2), 54 (2.9), 44 (2.1), 43 (43.3), 42 (44.1), 41 (9.0)
128 (0.1), 127 (1.7), 85 (22.2), 84 (100), 82 (2.1), 56 (7.9), 55 (19.9), 54 (2.7), 44 (2.0), 43 (11.7), 42 (40.7), 41 (6.6)
128 (1.1), 127 (1.3), 85 (09.1), 84 (100), 82 (2.2), 56 (6.6), 55 (18.4), 54 (2.4), 44 (2.3), 43 (5.9), 42 (39.6), 41 (6.1)
128 (1.4), 127 (2.0), 85 (43.7), 84 (100), 82 (2.2), 56 (9.5), 55 (21.4), 54 (2.5), 44 (1.6), 43 (19.3), 42 (39.6), 41 (6.9)
D-[1-13C]glucose
D-[2-13C]glucose
D-[3-13C]glucose
D-[4-13C]glucose
D-[5-13C]glucose
D-[6-13C]glucose
1-(1′-Pyrrolidinyl)-2-butanone (4)
D-glucose
142 (0.0), 141 (1.6), 85 (5.7), 84 (100), 82 (1.5), 56 (3.5), 55 (10.7), 54 (1.5), 44 (0.2), 43 (1.5), 42 (21.0), 41 (3.6)
142 (0.5), 141 (1.3), 85 (7.2), 84 (100), 82 (1.4), 56 (3.8), 55 (11.3), 54 (1.7), 44 (0.2), 43 (1.8), 42 (22.4), 41 (3.9)
142 (0.5), 141 (1.2), 85 (7.3), 84 (100), 82 (1.4), 56 (3.7), 55 (10.9), 54 (1.7), 44 (0.1), 43 (1.8), 42 (21.8), 41 (3.8)
142 (2.6), 141 (0.7), 85 (92.6), 84 (100), 82 (1.5), 56 (9.7), 55 (17.5), 54 (2.3), 44 (1.5), 43 (21.8), 42 (27.6), 41 (6.4)
142 (1.8), 141 (0.4), 85 (33.3), 84 (100), 82 (1.3), 56 (5.5), 55 (12.8), 54 (1.9), 44 (0.8), 43 (7.8), 42 (24.6), 41 (4.6)
142 (1.1), 141 (0.7), 85 (7.0), 84 (100), 82 (1.4), 56 (4.0), 55 (11.4), 54 (2.1), 44 (0.5), 43 (2.0), 42 (24.5), 41 (4.0)
142 (1.7), 141 (1.0), 85 (45.2), 84 (100), 82 (1.4), 56 (5.7), 55 (12.7), 54 (1.7), 44 (0.5), 43 (9.4), 42 (22.5), 41 (4.1)
D-[1-13C]glucose
D-[2-13C]glucose
D-[3-13C]glucose
D-[4-13C]glucose
D-[5-13C]glucose
D-[6-13C]glucose

676 J. Agric. Food Chem., Vol. 44, No. 3, 1996
Huyghues-Despointes and Yaylayan
Ta ble 4. Distr ibu tion of La bels in 2
Sch em e 2. Retr o-Ald ol F r a gm en ta tion a t C3-C4 of
dihydroxy-6-methyl-4(H)-pyran-4-one seem to be the
principal degradation steps in Amadori compounds
derived from secondary amines under ribbon pyrolysis
condition, which, as demonstrated earlier (Huyghues-
Despointes et al., 1994), releases the initial degradation
products of an analyte.
P r olin e Am a d or i P r od u ct (P a th w a y A)a
To verify the pathways of formation of C3 fragments,
model studies with D-[13C]glucose labeled at different
carbons were carried out with proline using Py/GC/MS
as described earlier (Huyghues-Despointes et al., 1994).
The distribution of the 13C labels in the C3 fragmenta-
tion indicator compound 2 was analyzed. The results
are summarized in Tables 3 and 4. According to Table
4, 55% of 2 is formed from the first three carbon atoms
of D-glucose and 45% from the last three carbon atoms.
This distribution could be rationalized by the formation
of 1-(prolino)-1-deoxy-D-glyceraldehyde and glyceralde-
hyde from proline Amadori product by a retro-aldol
reaction at C3-C4 (pathway A, Scheme 1) as described
in detail below.
Formation of 1-(Prolino)-1-deoxy-D-glyceraldehyde and
Glyceraldehyde from Proline Amadori Product. Ama-
dori products can undergo a retro-aldol reaction by
cleavage at the C3-C4 bond as shown in Scheme 2 to
produce 1-(prolino)-1-deoxy-D-glyceraldehyde (7) and a
free glyceraldehyde molecule. The latter, being in
equilibrium with the dihydroxacetone form, can react
with free proline, from either end of the molecule, to
produce an isotopomeric mixture of compound 7 with
labeled D-glucoses at the last three carbon atoms by
Amadori rearrangement mechanism. It seems that
carbonyl migration occurs quite efficiently in glyceral-
dehyde at 250 °C through an enediol mechanism.
1-(Prolino)-1-deoxy-D-glyceraldehyde (7) was not de-
tected in the pyrolysis products of the proline Amadori
a
RA [x,y] ) retro-aldol cleavage at Cx-Cy.
compound but was detected as its trimethylsilyl deriva-
tive in trace amounts after treatment of the reaction
mixture with trimethylsilyl chloride. However, the
corresponding 1-(morpholino)-1-deoxy-D-glyceraldehyde
was detected in the pyrolysate of morpholine Amadori
compound without silylation. The structure was ten-
tatively assigned on the basis of the mass spectrometric
fragmentation pattern in comparison with the proline
analog (Tables 1 and 2). Other products observed in
the pyrogram of morpholine Amadori compound are
listed in Table 2.
1-(Prolino)-1-deoxy-D-glyceraldehyde (7) was synthe-
sized to examine its stability and potential of conversion
into 1-(1′-pyrrolidinyl)-2-propanone under pyrolysis con-
ditions. Figure 1 shows the pyrogram of compound 7
generated at 250 °C. The main product formed under
pyrolysis condition was 1-(1′-pyrrolidinyl)-2-propanone,
indicating the facile formation of 2 from 7. However,

Retro-Aldol and Redox Reactions of Amadori Compounds
J. Agric. Food Chem., Vol. 44, No. 3, 1996 677
F igu r e 1. Pyrogram of 1-(prolino)-1-deoxy-D-glyceraldehyde (7).
Sch em e 3. â-Elim in a tion Rea ction of 1-(P r olin o)-
initiated from the 2,3-enediol form (7′) as shown in
Scheme 3 to produce a free proline and a pyruvaldehyde
molecule. To assess the potential of pyruvaldehyde/
proline to produce 1-(1′-pyrrolidinyl)-2-propanone, the
mixture was analyzed by Py/GC/MS. The major prod-
ucts identified were 1-(1′-pyrrolidinyl)-2-propanone, ac-
etol, 1,2-propanediol, and acetic acid among others,
including several aldol condensation products. Gi and
Baltes (1994) also detected 1,2-propanediol and acetol
in the heated mixtures of histamine and pyruvaldehyde.
Pyruvaldehyde, therefore, can produce 1-(1′-pyrrolidi-
nyl)-2-propanone in the presence of free proline or
pyrrolidine. Two possible mechanisms can be consid-
ered for this transformation: either the pyruvaldehyde
reacts with pyrrolidine and forms the imminium ion (8,
Scheme 4), which is then reduced to 2, or pyruvaldehyde
is reduced first into 2-hydroxypropionaldehyde, which
can exist in equilibrium with acetol, and then undergoes
Amadori rearrangement to 2 as shown in Scheme 4.
Acetol has been detected in heated mixtures of pyru-
valdehyde/amino acids and in heated Amadori product
mixtures. In addition, it can react with proline (or
pyrrolidine) to produce the Heyn’s product 9 as shown
in Scheme 4. To test this hypothesis, acetol was reacted
with pyrrolidine at 100 °C for 10 min and the ether
extract was analyzed by GC/MS. When equimolar
amounts of reactants were used, surprisingly, the major
product was 2, not 9, and only trace amounts of Heyn’s
product (9) was detected in addition to 1,2-dipyrrolidi-
nyl-1-propene (10). However, when acetol was treated
with a 2-fold excess of pyrrolidine, the area of the peak
assigned to 10 increased 6 times relative to the acetol/
pyrrolidine equimolar mixture. Compound 10 can be
detected in the pyrolysis mixtures of pyruvaldehyde/
proline, glycerladehyde/proline, proline Amadori, and
acetol/pyrrolidine and in 1-(prolino)-1-deoxy-D-glycer-
aldehyde. A related compound (11) was isolated by
Tressl et al. (1993) in a heated D-glucose/L-proline
mixture, which in principle could be formed from
1-deoxyglucosone through a similar reaction sequence
followed by cyclization.
1-d eoxy-D-glycer a ld eh yd e (P a th w a y A)a
a
Carbon numbering indicates original D-glucose carbon
locations.
Sch em e 4. F or m a tion P a th w a ys of 1-(1′-P yr r olid in yl)-
2-p r op a n on e (2) fr om P yr r olid in e a n d P yr u va ld eh yd e
(P a th w a y A)a
a
[H] ) reductions.
the pyrolysis of a mixture of proline and glyceraldehyde
produces several major peaks, among them 1-(1′-pyr-
rolidinyl)-2-propanone. Although 7 is stable enough to
be formed from Amadori products, it degrades quickly
to form 1-(1′-pyrrolidinyl)-2-propanone as described
below.
Mechanism of Conversion of 1-(Prolino)-1-deoxy-D-
glyceraldehyde into 1-(1′-Pyrrolidinyl)-2-propanone.
1-(Prolino)-1-deoxy-D-glyceraldehyde (7), similar to other
Amadori products, can undergo â-elimination reaction
Mechanism of Formation of C2 and C4 Fragments.
Detection of 2-hydroxy-1-(1′-pyrrolidiyl)-1-buten-3-one
(3), 1-(1′-pyrrolidinyl)-2-butanone (4), and N-acetylpyr-
rolidine in the reaction mixture of L-proline/D-glucose
or in the pyrolysis products of proline Amadori com-
pound indicates the occurrence of fragmentation of

678 J. Agric. Food Chem., Vol. 44, No. 3, 1996
Huyghues-Despointes and Yaylayan
Ta ble 5. Distr ibu tion of La bels in 4
D-glucose and/or Amadori compounds to produce C2 and
C4 fragments. Analysis of the data from labeling
experiments has indicated that both 2-hydroxy-1-(1′-
pyrrolidinyl)-1-buten-3-one (3) and 1-(1′-pyrrolidinyl)-
2-butanone (4) have similar distributions of labels;
however, due to the purity of the peak associated with
2-hydroxy-1-(1′-pyrrolidiyl)-1-buten-3-one, percentage
distributions of labels are shown only for 1-(1′-pyrro-
lidinyl)-2-butanone (Tables 3 and 5) and N-acetylpyr-
rolidine (Tables 3 and 6). Careful analysis of the data
from all of the labeling experiments indicates at least
three pathways of formation of 1-(1′-pyrrolidinyl)-2-
butanone, designated B, C, and D (Scheme 1). Tables
3, 5, and 6 summarize the distribution of labels in
N-acetylpyrrolidine and 4. According to the labeling
experiments, 70% of 3, 4, and N-acetylpyrrolidine is
produced through retro-aldol cleavage at C2-C3 (path-
way D) and 30% through similar retro-aldol cleavage
at C4-C5 (pathways B and C) (Scheme 1). In pathways
B, C, and D acetic acid (which eventually reacts with
pyrrolidine to form N-acetylpyrrolidine) is assumed to
be formed from oxidation of the C2 fragment glycolade-
hyde, due to the similar percentage of distribution of
expected labels in N-acetylpyrrolidine and compounds
3 and 4.
produce 2-hydroxy-1-(1′-pyrrolidinyl)-1-buten-3-one and
glycoldehyde, and both products can undergo thermal
oxidation-reduction to produce 1-(1′-pyrrolidinyl)-2-
butanone and acetic acid, respectively (see Schemes 1
and 5 and section B for the mechanism of oxidation-
reduction). This pathway, however, represents only
around 5% of the total yield of C2/C4 products. This
pathway leads to the formation of acetic acid containing
C5 and C6 of the D-glucose and to 1-(1′-pyrrolidinyl)-2-
butanone containing C1, C2, C3, and C4 of D-glucose;
C1 of D-glucose is attached to pyrrolidine, as shown in
Schemes 1 and 5.
Formation of C2/ C4 Fragments from Pathway C.
1-Deoxyglucosones (1DG) can be formed through â-
elimination of Amadori compounds under basic condi-
tions, which also promotes retro-aldol reactions from the
acyclic forms of 1DG. Such retro-aldol reaction at C4-
C5 (RA[4,5]), similar to pathway B, can generate a
glycoladehyde (eventually acetic acid) and 1-hydroxy-
2,3-butandione (12). The latter can react with pyrro-
lidine through Amadori rearrangement to produce
2-hydroxy-1-(1′-pyrrolidinyl)-1-buten-3-one and ulti-
mately 1-(1′-pyrrolidinyl)-2-butanone as described above.
Pathway C represents around 25% of the total yield of
C2/C4 products and leads to the formation of acetic acid
consisting of C5 and C6 of the D-glucose and 1-(1′-
pyrrolidinyl)-2-butanone containing C1, C2, C3, and C4
of D-glucose, but in this case, contrary to pathway B,
C4 of D-glucose is attached to the pyrrolidine, as shown
in Scheme 1.
Formation of C2/ C4 Fragments from Pathway B. The
formation of 1-(amino acid)-1,4-dideoxy-2,3-hexodiuloses
(4D, Scheme 1) in Maillard mixtures has been estab-
lished (Huber et al., 1990). 1-(Prolino)-1,4-dideoxy-2,3-
hexodiulose formed from the Amadori compound can
undergo a retro-aldol reaction at C4-C5 (RA [4,5]) to

Retro-Aldol and Redox Reactions of Amadori Compounds
J. Agric. Food Chem., Vol. 44, No. 3, 1996 679
Ta ble 6. Distr ibu tion of La bels in N-Acetylp yr r olid in e
Sch em e 5. F or m a tion P a th w a ys of 1-(1′-P yr r olid in yl)-
2-bu ta n on e (4) (P a th w a ys B-E)a
and by FTIR (Yaylayan and Ismail, 1995; Yaylayan et
al., 1994). A retro-aldol reaction at C2-C3 (RA[2,3]) of
1-(prolino)-1-deoxy-4-hexosulose can produce a tetrulose
moiety in equilibrium with tetrose and consisting of C3,
C4, C5, and C6 atoms of the glucose. The symmetrical
enediol form of the tetrulose can undergo a â-elimination
from either end of the molecule to produce 1-hydroxy-
2,3-butandione (12), a common intermediate with path-
way C. Alternatively, the tetrose form can undergo an
Amadori rearrangement, again from either end of the
molecule; the resulting isotopomers of the Amadori
product can â-eliminate and produce 2-hydroxy-1-(1′-
pyrrolidinyl)-1-buten-3-one, the common intermediate
of pathways B-E. Similar to glyceraldehyde, carbonyl
migration occurs quite efficiently also in tetrose moieties
at 250 °C.
B. Red ox Rea ction s of r-Hyd r oxy Ca r bon yl a n d
r-Dica r bon yl Com p ou n d s. Most of the reduction
steps needed in Maillard systems to justify the struc-
tures of observed products are attributed to the so-called
reductones, cyclic or acyclic enediols. The mechanisms
of these reduction reactions are not explored yet in
Maillard systems. Three distinct mechanisms of oxida-
tion-reductions were identified that involve common
Maillard intermediates such as R-hydroxy carbonyls,
R-dicarbonyls, and formic acid.
a
[H] ) reductions, Carbon numbering indicates original
D-glucose carbon locations.
Formation of C2/ C4 Fragments from Pathway D.
Pathway D, the major pathway for the formation of C2/
C4 fragments (70%), is based on the migration of the
carbonyl group of the Amadori product into C4 of the
D-glucose moiety, forming 1-(prolino)-1-deoxy-4-hexosu-
lose. Migration of carbonyl groups has been verified in
carbohydrates by NMR (Cervantes-Laurean et al., 1993)
Reduction of Imminium Ions by Formic Acid. Formic
acid is a common degradation product of carbohydrates,
and it is an effective and specific reagent for reductive
amination, known as the Wallach reaction (Moore,

680 J. Agric. Food Chem., Vol. 44, No. 3, 1996
Huyghues-Despointes and Yaylayan
Sch em e 7. Dim er iza tion of P yr u va ld eh yd e Hyd r a te
a n d Su bsequ en t Th er m a lly Allow ed Electr ocyclic
Rin g Op en in g
F igu r e 2. Structures of trimeric glyoxal dihydrate and
glycoaldehyde dimer.
Sch em e 6. Red u ction Mech a n ism of Im m in iu m Ion s
a n d Ca r bon yl Com p ou n d s by F or m ic Acid
Sch em e
8.
Disp r op or tion a tion
of
r-Hyd r oxy
Ca r bon yls a n d r-Dica r bon yl Com p ou n d sa
1949). If formamide or formate salts are used instead
of formic acid, the reaction is called the Leuckart
reaction (Moore, 1949). The mechanism of this reaction
has been elucidated by deuterium labeling studies
(Leonard and Sauers, 1957) and is illustrated in Scheme
6 for the reduction of 8 to 2. The mechanism involves
a hydride transfer to the unsaturated site and conver-
sion of formic acid into carbon dioxide. When benzal-
dehyde in the presence of pyrrolidine was treated with
formic acid or sodium cyanoborohydride, a specific
reductive amination reagent, both reaction mixtures
produced high yields of benzylpyrrolidine. Similarly,
when pyruvaldehyde was treated with formic acid in the
presence of pyrrolidine, compound 2 was produced as
the major product.
Cyclic Dimer Formation of R-Hydroxy Carbonyl Com-
pounds followed by Thermally Allowed Electrocyclic
Ring Opening-Formation of Acetol from Pyruvaldehyde.
R-Hydroxy carbonyls and hydrated R-keto aldehydes are
known to undergo spontaneous dimerization reactions
to produce various cyclic 1,4-dioxane derivatives. Glyc-
eraldehyde and dihydroxyacetone are known to exist in
different cyclic dimeric forms that can be crystallized
separately (Kobayashi et al., 1976); glyoxal and glycoal-
dehyde can exist in different dimeric and trimeric forms
which are available commercially (Figure 2). These 1,4-
dioxane derivatives when heated can undergo electro-
cyclic ring opening reactions to produce oxidation and
reduction products. When trimeric glyoxal diyhydrate
was pyrolyzed at 250 °C, in addition to monomeric and
dimeric units, formic acid and carbon dioxide were the
major products identified. Similarly, when pyruvalde-
hyde was heated at 100 °C for 8 min and analyzed by
GC/MS, 40 peaks were produced but the 3 major
products were 1,2-propanediol (18%), acetic acid (30%),
acetol (7%), and 2,4-dimethyl-1,3-dioxolane (7%) (form-
aldehyde protected 1,2-propanediol). Scheme 7 shows
the dimerization of the hydrated pyruvaldehyde and
subsequent electrocylic ring opening reaction to produce
the observed main products and formic acid. This
process can explain the formation of acetol from pyru-
valdehyde and its subsequent reaction with pyrrolidine
to produce compound 2. The further reduction of acetol
a
Pathway a is the concerted mechanism. Pathway b is the
biradical mechanism.
through its isomeric 2-hydroxypropionaldehyde into 1,2-
propanediol can be explained by the reducing action of
formic acid (one of the predicted but not observed
products) on aldehydes similar to imminium ions
(Scheme 6). As shown before, after reduction, formic
acid is converted into carbon dioxide and hence not
detected in the reaction mixture.
Disproportionation of R-Dicarbonyl and R-Hydroxy
Carbonyl Compounds through Enediol Intermediates
Reduction of R-Dicarbonyls to Enediols. R-Dicarbonyl
and R-hydroxy carbonyl compounds play a major role
in the Maillard reaction. The enediol forms of R-hy-
droxy carbonyl compounds are known as reductones.
One application of the chemistry of reductones that has
been studied outside the Maillard reaction context is
their use as dynamic molecular memory devices (Aviram
et al., 1982). In this context, an enediol/R-dicarbonyl
complex undergoes a double proton transfer (see Scheme
8) to convert the starting enediol into its corresponding
dicarbonyl and the starting dicarbonyl into its enediol,
thus affecting a redox reaction. These interconversions
can proceed either by a stepwise biradical mechanism
from the excited triplet state or by a concerted double
proton transfer from the ground state (Scheme 8). For
the model dihydroxyethylene/glyoxal complex the en-
ergy requirement for the biradical pathway was calcu-
lated to be 113.8 kcal mol-1 as opposed to 149.7 kcal
mol-1 for the concerted mechanism (Tachibana et al.,
1989). The significance of these calculations for the
Maillard reaction lies in the fact that it can provide a
theoretical explanation for the importance of an R-dike-
tone/enediol redox couple in generating oxidation-
reduction products and for the detection of free radicals
in Maillard systems. Reduction of 2-hydroxy-1-(1′-
pyrrolidiyl)-1-buten-3-one into its enediol form shown
in Scheme 5 could proceed through a similar mecha-

Retro-Aldol and Redox Reactions of Amadori Compounds
J. Agric. Food Chem., Vol. 44, No. 3, 1996 681
Sch em e 9. P r op osed Or igin of Ch em ilu m in escen ce
Sp ecies
droxyacetone ands D(+)- and DL-glyceraldehyde. J . Mol.
Struct. 1976, 35, 85-99.
Leonard, N. J .; Saures, R. R. Unsaturated amines XI. The
course of formic acid reduction of enamines. J . Am. Chem.
Soc. 1957, 79, 6210-6214.
Moore, M. L. Organic Reactions; Wiley: New York, 1949; p
309.
Morita, N.; Takagi, M. Browning reaction and quinoxaline
formation from carbohydrates. Dev. Food Sci. 1986, 13, 59-
65.
Namiki, M.; Oka, M.; Otsuka, M.; Miyazawa, T.; Fujimoto, K.;
Namiki, K. Weak chemiluminescence at an early stage of
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1709.
Papoulis, A.; Al-Abed, Y.; Bucala, R. Identification of N2-(1-
carboxyethyl)guanine (CEG) as a guanine advanced glyco-
sylation end product. Biochemistry 1995, 34, 648-655.
Tachibana, A.; Koizumi, M.; Tanak, E.; Minato, T. Double
proton transfer in the dihydroxyethylene-glyoxal complex.
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nism. The origin of chemiluminescent species (Namiki
et al., 1993) might be explained by the rearrangement
of carbon-centered free radicals of a R-diketone/enediol
complex shown in Scheme 8 to produce 1,2-dioxetane
structures (Scheme 9) which are known to emit photons
upon thermal decomposition (Waldemar and Katsu-
masa, 1979).
Con clu sion . Pyrolysis/GC/MS is a convenient ana-
lytical tool to perform small scale chemical reactions
with labeled starting materials. Labeling experiments
using pyrolysis/GC/MS indicate the importance of retro-
aldol reactions in model systems containing secondary
amino acids.
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Received for review May 12, 1995. Revised manuscript
received December 8, 1995. Accepted J anuary 2, 1996.X V.Y.
acknowledges funding for this research by the Natural Sci-
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