Imre Blank, Philippe Pollien, Chahan Yeretzian

Imre Blank, Philippe Pollien, Chahan Yeretzian
Nestlé Research Centre
Lausanne, Switzerland
PTR-MS Igls / IBk January 2003 1

Outline of the presentation
Basic principles of the Maillard reaction
Major reaction phases, intermediates, its role in food and medicine
PTR-MS data for reaction mechanisms
Maillard model systems based on proline
Main approaches in aroma research
Sensory directed chemical analysis
Formation kinetics of key odorants by PTR-MS
Maillard model systems based on proline
… but will not go into technical details of PTR-MS
PTR-MS Igls / IBk January 2003 2

Importance of the Maillard reaction
00-now
95-99
Time period (year)
90-94
85-89
80-84
75-79
70-74
65-69
Food
Flavours
Medicine
MS
0 200 400 600 800 1000 1200 1400
Publication number
PTR-MS Igls / IBk January 2003 3

Aldose
+ amino compound
-H 2
O
A
N-substituted glycosylamine
B
Amadori rearrangement
C
1-Amino-1-deoxy-2-ketose
(Amadori compound)
D
Schiff base
of HMF
or furfural
+ H 2
O
+ amino
compound
-3 H 2
O
- amino
compound
HMF
or furfural
G
F
Aldimines
E
-CO 2
Aldehyde
+ amino acid
Strecker
degradation
+ amino
compound
Aldoles and
N-free
polymers
+ amino
compound
C
-2 H 2
O
Reductones
-2H +2H
dehydroreductones
G G G
F
Fission
products
(acetol,
pyruvaldehyde,
diacetyl etc.)
F
+ amino
compound
G
Aldimines or
ketimines
G
(Hodge, 1953)
Aroma
Taste
Colour
Antioxidants
Texture
Nutritional value
Contaminants
Toxic compounds
MELANOIDINS (brown nitrogenous polymers and copolymers)
PTR-MS Igls / IBk January 2003 4

Major fluxes in the Maillard reaction
HO
HO
OH
O
OH
R
N
H
N-Glycosyl
COOH
OH
HO
OH
R
+
HO
H 2
N COOH
O OH
H 2
O
Aldose sugar
Amino acid
HO
HO
OH
OH
OH
R
N
H
COOH
N H 2
A
R
COOH
HO
HO
OH
3-DO
O
O
O H 2
HO
HCOOH
O
HMF
CHO
A : 1,2-enolisation
(pH < 7)
OH
Flavour
precursor

Formation of volatile compounds in prolinebased
Maillard reaction systems
Reaction conditions
Precursors
Glucose/L-proline (1:1), Fru-Pro (0.2 mol)
Buffer Phosphate (50 mL, 0.2 M, pH 5-8)
Temperature
Time
Analytical methods
Boiling (reflux)
Up to 7 h
PTR-MS: Direct headspace analysis
GC-MS: Tenax trapping, solvent extraction
PTR-MS Igls / IBk January 2003 6

Tenax
Experimental set-up
for Maillard PTR-MS
HS Conc [ppbv]
2000
1500
1000
500
0
mass 45 mass 47 mass 51
mass 57 mass 59 mass 71
mass 75 mass 87 mass 145
mass 43 mass 61
0 50 100 150 200 250 300
Time[min]
8000
7000
6000
5000
4000
3000
2000
1000
0
HS Conc. [ppbv], masses 43 and 61
(T= 90°C)
On-line monitoring of volatiles
generated upon heating
PTR-MS Igls / IBk January 2003 7

HO
HO
HO
HO
HO
HO
HO
OH
O
OH
OH
OH
OH
OH
R
N
H
OH
R
N
H
O
N
H
R
COOH
COOH
COOH
Amadori compound
HO
OH
OH
OH
R
N
H
COOH
N H 2
N H 2
OH
HO
OH
+
HO
O OH
H 2
O
A
R
B
R
COOH
COOH
HO
HO
Hexose sugar
HO
HO
OH
3-DO
O
OH
O
O
Melanoidins
Flavour
1-DO
O
[H]
O H 2
O H 2
HO
N H 2
O
HO
R
O
O
O
COOH
Amino acid
HCOOH
HMF
O
HDMF
OH
CH 3 OOH
DDMP
CHO
OH
HS Conc[ppbv]
HS conc. [ppbv]
180
160
140
120
100
80
60
40
20
0
H
?
O
OH
m/z = 47 Formic acid + X
0 50 100 150 200 250 300
Glu/Pro
Amadori
8000
7000
6000
5000
4000
3000
2000
1000
0
O
OH
Time[min]
m/z = 61 Acetic acid
0 50 100 150 200 250 300 350
Reaction time [min]
PTR-MS Igls / IBk January 2003 8

Coupling of GC with EI-MS and PTR-MS
PTR-MS Igls / IBk January 2003 9

HS Conc[ppbv]
180
160
140
120
100
80
60
40
20
0
m/z = 47 Formic acid + X
0 50 100 150 200 250 300
Time[min]
HS conc. [ppbv]
8000
7000
6000
5000
4000
3000
2000
1000
0
m/z = 61 Acetic acid
0 50 100 150 200 250 300 350
Reaction time [min]
RT First trap Second trap
(min) Peak hight % Peak hight %
4.46 7400 75 3847 34
12.5 2419 25 2234 19
33.45 0 0 5399 47
0 0 0 0
Total 9819 100 11480 100
RT First trap Second trap
(min) Peak hight % Peak hight %
10.5 6165 4 0 0
31.2 136646 96 7470083 100
0 0 0 0
0 0 0 0
Total 142811 100 7470083 100
1st trap
2nd trap
: 10-15 min
: 1h25-1h30
PTR-MS Igls / IBk January 2003 10

HO
HO
HO
HO
HO
HO
HO
OH
O
OH
R
N
H
COOH
Amadori compound
HO
OH
OH
OH
OH
OH
OH
OH
R
N
H
O
N
H
R
OH
R
N
H
COOH
COOH
COOH
N H 2
N H 2
OH
HO
OH
+
HO
O OH
H 2
O
A
R
B
R
COOH
COOH
HO
HO
Hexose sugar
HO
HO
OH
3-DO
O
OH
O
O
Melanoidins
Flavour
1-DO
O
[H]
O H 2
O H 2
HO
N H 2
HO
O
HO
R
O
O
COOH
Amino acid
HCOOH
O
O
O
HMF
HDMF
OH
CH 3 OOH
DDMP
OH
CHO
HS conc. [ppbv]
HS conc. [ppbv]
12
9
6
3
0
HO
O
m/z = 127 HMF
CHO
0 50 100 150 200 250 300 350
Reaction time [min]
Glu/Pro
Amadori
(pH 7)
60
50
40
30
20
10
0
m/z= 145 3,5-Dihydroxy-2,3-dihydro-6-methyl-4H -
pyran-4-one
0 50 100 150 200 250 300 350
Reaction time [min]
HO
O
O
OH
PTR-MS Igls / IBk January 2003 11

HO
HO
HO
HO
HO
HO
HO
OH
O
OH
OH
OH
OH
OH
R
N
H
OH
R
N
H
O
N
H
R
COOH
COOH
COOH
Amadori compound
HO
OH
OH
OH
R
N
H
COOH
N H 2
N H 2
OH
HO
OH
+
HO
O OH
H 2
O
A
R
B
R
COOH
COOH
HO
HO
Hexose sugar
HO
HO
OH
3-DO
Sugar
O
OH
O
O
fragmentation
1-DO
O
[H]
O H 2
O H 2
HO
N H 2
HO
O
HO
R
HCOOH
O
O
O
COOH
Amino acid
O
O
HMF
HDMF
OH
CH 3 OOH
DDMP
OH
CHO
HS conc. [ppbv]
HS conc. [ppbv]
600
500
400
300
200
100
0
O
m/z = 57 1-Hydroxy-2-propanone
OH
0 50 100 150 200 250 300 350
Glu/Pro
Amadori
60
50
40
30
20
10
0
Reaction time [min]
m/z = 89 3-Hydroxy-2-butanone
0 50 100 150 200 250 300 350
Reaction time [min]
(pH 7)
O
OH
PTR-MS Igls / IBk January 2003 12

Principle approaches in aroma research
OAV =
C x
T x
C x : Concentration
T x : Threshold value
2
SH
SH
O
SH
OH
1
OH
SH
SH
O
O
OH
Concentration (µg/L)
1000
800
600
400
200
0
Sauvignon
blanc
1 2 3 4 5
25
20
15
10
PTR-MS Igls / IBk January 2003 13
5
0
Odor Activity Value

Formation of odorants in proline-based
Maillard reaction systems
Reaction conditions
Precursors
Glucose/L-proline (1:1), Fru-Pro (0.2 mol)
Buffer Phosphate (50 mL, 0.2 M, pH 5-8)
Temperature
Time
Analytical methods
Boiling (reflux)
Up to 7 h
GC-O, GC-MS: Solvent extraction
PTR-MS: Direct headspace analysis
PTR-MS Igls / IBk January 2003 14

Volatile compounds formed in proline-based
Maillard model systems 1
Fru-Pro: Caramel-like, roasty Glc/Pro: Roasty, popcorn-like
O
O
OH
O
OH
O
OH
O
OH
O
O
HO
O
OH
CH HO 3 COOH HCOOH
O
OH
HO
O
OH
OH
O
OH
O
OH
O
O
O
O
N
O
N
H
O
N
N
O
O
N
H
O
PTR-MS Igls / IBk January 2003 15

Formation of key odorants in Maillard systems
based on proline 2
Concentration (ug/mmol)
Concentration (ug/mmol)
25000
20000
15000
10000
50
40
30
20
10
0
5000
0
OH
0 1 2 3 4 5
Time (h)
N
H
0 1 2 3 4 5
Time (h)
O
N
O
O
K
Concentration (ug/mmol)
Amadori
Glu/Pro
(pH 7)
Concentration (ug/mmol)
300
250
200
150
100
5
4
3
2
1
0
50
0
0 1 2 3 4 5
Time (h)
PTR-MS Igls / IBk January 2003 16
O
O
OH
0 1 2 3 4 5
Time (h)
N
O

Monitoring of key odorants by PTR-MS
HS conc. [ppbv]
HS conc. [ppbv]
8000
7000
6000
5000
4000
3000
2000
1000
0
16
14
12
10
8
6
4
2
0
O
O
O
OH
OH
m/z = 61 Acetic acid
0 50 100 150 200 250 300 350
Reaction time [min]
m/z = 129 Furaneol
0 50 100 150 200 250 300 350
Reaction time [min]
HS conc. [ppbv]
Glu/Pro
Amadori
(pH 7)
HS conc. [ppbv]
1000
900
800
700
600
500
400
300
200
100
0
30
25
20
15
10
5
0
m/z= 87 Diacetyl
0 50 100 150 200 250 300 350
Reaction time [min]
m/z = 126 2-Acetyltetrahydropyridine
0 50 100 150 200 250 300 350
Reaction time [min]
N
O
O
O
PTR-MS Igls / IBk January 2003 17

Glucose/proline (90°C)
Tenax trapping (1h20 to 1h40min)
GC EI-MS/PTR-MS coupling
O
PTR-MS Igls / IBk January 2003 18
Acetaldehyde
3-Hydroxy-2-butanone
1-Hydroxy-2-propanone
Acetic acid
2-Acetyltetrahydropyridine
O
O
N
Acetone
Diacetyl

Key odorants below the detection limit of PTR-MS
(sample 50-fold diluted)
3.5
m/z = 112 2-Acetyl-1-pyrroline
12
m/z = 127 3,4-Dimethylcyclopentenolone
HS conc. [ppbv]
3
2.5
2
1.5
1
0.5
0
Reaction time [min]
0 50 100 150 200 250 300 350
HS conc. [ppbv]
10
8
6
4
2
0
Reaction time [min]
0 50 100 150 200 250 300 350
Glu/Pro
Amadori
(pH 7)
N
O
O
OH
PTR-MS Igls / IBk January 2003 19

H O
H O
O
OH
O
OH
OH
OH
+
N
N
H
O
COOH
O
HS conc. [ppbv]
600
500
400
300
200
100
0
m/z = 57 1-Hydroxy-2-propanone
(pH 7)
Glc/Pro
Amadori
0 50 100 150 200 250 300 350
Reaction time [min]
N
N
H
O
N
OH
H
N
N
H O
O
(Schieberle et al., 2000)
O
O
[O]
H 2
O
CO 2
O
HS conc. [ppbv]
30
25
20
15
10
5
0
m/z=126 2-Acetyl-1,4,5,6-tetrahydropyridine
(pH 7)
0 50 100 150 200 250 300 350
Reaction time [min]
Glc/Pro
Amadori
PTR-MS Igls / IBk January 2003 20

Good correlation of PTR-MS and GC-IDA data
HS conc. [ppbv]
30
25
20
15
10
5
0
m/z = 126, Acatyltetrahydropyridine
0 50 100 150 200 250 300 350
Reaction time [min]
Glu/Pro
Amadori
HS conc. [ppbv]
PTR-MS
16
14
12
10
8
6
4
2
0
Glu/Pro
Amadori
m/z = 129, Furaneol
0 50 100 150 200 250 300 350
Reaction time [min]
Concentration (ug/mmol)
50
40
30
20
10
0
0 1 2 3 4 5 6
Time (h)
K
Glu/Pro
Amadori
Concentration (ug/mmol)
GC-IDA
300
250
200
150
100
50
0
0 1 2 3 4 5 6
Time (h)
K
Glu/Pro
Amadori
PTR-MS Igls / IBk January 2003 21

Conclusions
Key odorants can be monitored by PTR-MS
Furaneol, acetic acid, diacetyl, 2-AP, ATHP, methional, etc.
Good correlation between PTR-MS and GC-IDA data
Comparable kinetic curves
PTR-MS gives complementary information to chromatography-based tools
rapid, time-resolved, dynamic changes of volatiles, mechanistic insight
Broad application in thermal-induced reactions
Maillard reaction, lipid oxidation, flavour formation, off-flavours, ...
Some remaining issues:
Sensitivity, fragmentation, quantification, selectivity, ...
PTR-MS Igls / IBk January 2003 22

Acknowledgements
Stéphanie DEVAUD
Walter MATTHEY-DORET
Fabien ROBERT
Santo ALI
Christian LINDINGER
PTR-MS Igls / IBk January 2003 23