Flavor Precursors for Enzymes

1
Flavor Precursors for Enzymes
2
Precursors of Flavor Compound Formation by
Enzymes and Microorganisms

• Free fatty acid in milk
• Diacetyl in butter
• Garlic or Onion
• Tomato flavor
• Aspargusic acid
• Fruit flavor by Neurospora

3
Free Fatty Acid Formation from Triglycerides by
Lipase
Optimum temperature 1540 oC Lipids
Lipase free fatty acid
4
Diacetyl Formation in Butter from Lactose
Lactose
Oxalacetic acid
Citric acid
S. Lactis
COOH
COOH
- Acetate
CO
CH2
CH3
CH2
HO-C-COOH
H-COH
COOH
CH2
COOH
COOH
Lactic acid
OH
OH
(odorless )
H
C
C
C
C
H
3
O
H
3
H
H
-CO
2
2,3-butylane glycol
CH3
CO
COOH
H
Pyruvic acid
Aldol condensation
C
H
O
O
OH
3
-CO
2
H
C
O
H
C
C
-CO
Acetoin
H
C
C
C
C
H
3
2
3
3
COOH
H
CH3
O
Acetyl lactic acid
CO
O
O
H
H
C
C
C
C
H
3
3
Acetaldehyde
Diacetyl
5
Fresh Banana Flavor Formation by Enzyme
Fresh banana processing processed banana
lost flavor Banana
peel extraction enzyme
(flavorase) Processed banana Flavorase
fresh banana flavor
34 hrs If pyruvate, acetate, amino acid,
unsaturated fatty acid are added, then the time
for flavor production will be shortened to 30
min.
6
Onion and Garlic Flavor Formation by Enzyme
Enzymatic reaction of cysteine derivative
O
R-S-CH2-CH-COOH
NH2

R CH3- (Allin)
Propenyl alcohol or Allyl alcohol and oxidation
R CH3-CHCH- (Propenyl allin ? Onion more
predominant)
R CH2CH-CH2- (Allyl allin ? Garlic)
Alliinase
H2O
NH3

CH3-CH2-CHSO
Thiopropenal oxide
Propenyl sulfenic acid
Pyruvic acid
Lachrymator in onion
7
Formation of 2-Methyl-2pentenal
CH3-CH2-CHSO
Thiopropanal oxide (m/z 90)
Onion enzyme
Lachrymator in onion
Propenyl cysteine sulfoxide (Propenyl allin)
- S
CH3-CH2-CHO
CH3-CHCH-OH
(m/z 58)
(m/z 58)
Aldol condensation and Dehydration (H2O)
2-Methyl-2-pentenal (m/z 98)
8
Fresh Onion and Garlic Flavor Formation
O
Sulfenic acid
2 R-S-H
- H2O
O
Thiosulfinate (fresh flavor of onion and garlic)
R-S-S-R
O
H3C-CHCH-S-S-CHCH-CH3
Fresh onion odor
2-Propenylsulfinate
O
Fresh pleasant garlic-like odor
H2CCH-CH2-S-S-CH2-CHCH2
1-Propenyl or Allyl sulfinate
9
Flavor Properties of Thiosulfinate and
Thisulfonate
O
Thiosulfinate
Fresh Flavor
R
S
S
R
O
Thiosulfonate
Aged Flavor
S
R
S
R
O
O
R
S
O
Bitter Flavor (Off Flavor)
10
Reactions of Propenyl Cysteine Sulfoxide
Propenyl allin (Propenyl cycteine
sulfoxide) Onion flavor
O
H3C-CHCH-S-CH2-CH-NH2
COOH
O
H2O
S
CH2
H2C
O
- H2O
CH-COOH
H3C-HC
H3C-CH-CH2-S-CH2-CH-NH2
N
OH
COOH
H
Cycloalliin
(No flavor contribution)
11
Biogenesis of Flavor Compounds in Tomato
Important volatile flavor compounds in tomato
3-cis-hexenol Green
note isovalervaldehyde hexanol
Green or grassy odor
hexanal 2-trans-hexenal 2-cis-hexenal 2-isobutylt
hiazole Strong green leaf odor 3-methyl-1-butano
l
Fruit flavor
12
Tomato or Fruit Flavor Formation from Leucine
C
O
O
H
C
O
O
H
C
H
O
H

C
H
O
2
NADH H
C
H
H
N
C
O
C
H
C
O
ADH
2
C
H
2
2
2
C
H
C
H
C
H
H
C
2
2
C
H
H
C
3
NAD

3
C
H
H
C
C
H
H
C
C
H
3
3
C
H
3
3
C
H
C
H
3
3
L-
Leucine
3-Methyl-butanal
3-Methyl-1-
butanol
2-Keto-4-methyl-
pentanoic acid
Tomato of fruit flavor
Tomato flavor
ADH - alcohol NAD oxidoreductase
alcohol dehydrogenase
13
Enzyme Functions on Flavor Flavor Formation
add to 14C label leucine
Crude extract of fresh tomato
Formation of 14C label 3-methyl-1-butanol
add to
Boiled extract of fresh tomato No
formation of 14C label 3-methyl-1-butanol Indica
tes the enzymatic nature of this reaction
14
Precursor of 2-Isobutylthiazole
May be L-leucine
3-Methyl-1-butanal
formed from leucine reacts with
cysteinamine (HS-CH2-CH2-NH2) to produce
2-Isobutylthiazole
N
C
H
3
Tomato flavor
C
H
S
C
H
2
C
H
3
15
Fatty Acid as Flavor Precursor
O
16 15 13 12
10 9
CH3-CH2-CHCH-CH2-CHCH-CH2-CHCH-(CH2)7-C-OH
cis cis cis

Linolenic acid
lipoxygenase
O

2
O
16 15 13 12 11
10 9
CH3-CH2-CHCH-CH2-CH-CHCH-CHCH-(CH2)7-C-OH
trans cis
OOH
13-hydroperoxide
O
CH3-CH2-CHCH-CH2-C-H
3-cis-hexenal
16
Fatty Acid as Flavor Precursor
O
O
CH3-CH2-CHCH-CH2-C-H
CH3-CH2-CH2-CHCH-C-H
AOR
3-cis-hexenal
2-trans-hexenal
CH3-CH2-CHCH-CH2-CH2OH
AOR
O
3-cis-hexenol
CH3-CH2-CH2-CH2-CH2-C-H
CH3-CH2-CH2-CHCH-CH2OH
hexanal
trans-2-hexenol
CH3-CH2-CHCH-CH2-CH2OH
n-hexenol
AOR alcohol oxidoreductase
17
Asparagusic Acid in Asparagus
Asparagusic acid 1,2-dithiolane-4-carboxylic
acid Asparagusic acid , its methyl and
ethyl esters and several other sulfur compounds
were synthesized in the intact plant cells of
asparagus..
S
S
COOH
18
Asparagusic Acid in Asparagus
NH2
O
COOH
COOH
COOH
Valine
2-Methyl propanoic acid
CH3
Desaturation
S
S
S
COOH
H2S
COOH
COOH
CH3
CH3
CO
CO
S
S
H
SH
S
S
H
S
H
S
H
Desaturation
H2S
COOH
COOH
COOH
COOH
COOH
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Flavor formation by NeurosporaProduction of
Fruity Aroma by Various Strains of Neurospora
Neurospora Species Aroma Neurospora
sitophila ATTC46892 Fruity Neurospora No
1 Fruity Neurospora No. 2 Fruity Neurosp
ora No. 3 Fruity Neurospora No
4 Fruity Neurospora No 5 Fruity Neurospo
ra No 6 Fruity Neurospora No
7 Fruity Neurospora tetrasperma NRRA2164 No
aroma Neurospora crassa NRRA 2223 No
aroma Neurospora sitophila NRRA 2884 No
aroma Neurospora intermedia NRRA 5506 No
aroma
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Fruit Flavor Formation by Neurospora
Neurospora sitophila ATTC46892, Neurospora
No.1,2,3,4,5,6, and 7 were isolated from
beiju. Tweenty strains of Neurospora sp.isolated
from the state of Sao Paulo did not produce
fruity aroma
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Dynamic headspace/gas chromatography of volatiles
of Neurospora

• Neurospora sitophila ATCC46892
• Neurospora sp. No 6
• Neurospora sp. No. 5
• Neurospora sp. No.1
• Neurospora sp. NRRL 2884

• Ethyl acetate
• Ethanol
• 3-methyl-1-butanol
• Ethyl hexanoate
• 1-octen-3-ol

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Volatile Compounds (ppm) produced by Neurospora
sp. Isolated from beiju
Ethyl
Ethanol
3-Methyl-
Ethyl
1-Octen-
Acetate
1-butanol
hexanoate
3-ol
Neurospora
4.8
128
318
59
40
sitophila
Neurospora
9.0
111
ND
ND
ND
Sp. 1
Neurospora
0.9
111
117
10
50
Sp. 5
Neurospora
2.8
99
208
20
ND
Sp. 6
ND Not detected
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