Biosynthesis of Plant-derived flavor compounds By Dudsadee Uttapap

Biosynthesis of Plant-derived flavor

compounds

By Dudsadee Uttapap

References

1. “Flavor Chemistry and Technology”, H.B. Heath, G. Reineccius, 1986.

2. “Flavor Chemistry”, D.B. Min, http://class.fst.ohio-state.edu/fst820/default.htm

3. “Biosynthesis of plant-derived flavor compounds”, The Plant

Journal (2008) 54, 712–732

4. “Plant Biochemistry” http://www.uky.edu/~dhild/biochem/lecture.html

Biosynthesis of plant-derived flavor compounds

Flavor compoundsFlavor compounds

Flavor molecules constitute a heterogeneous group of compounds, with straight-chain, branched-chain, aromatic and heteroaromatic backbones

bearing diverse chemical groups such as hydroxyl, carbonyl, carboxyl, ester, lactone, amine, and thiol functions. More than 700 flavor chemicals h

ave been identified and catalogued

Chemical synthesis VS Chemical synthesis VS BiosynthesisBiosynthesis

Most commercial flavorants are ‘nature identical’, which means that they are the chemical equivalent of natural flavors but are chemically synthesized, mostly from petrol

eum-derived precursors

Bioproduction, including the extraction from natural sources, de novo microbial processes (fermentation), and bioconversion of natural precursors using micro-organisms or isolated enzymes

Biological functions of plant volatilesBiological functions of plant volatiles

Compounds emitted by flowers most probably serve to attract and guidCompounds emitted by flowers most probably serve to attract and guide pollinators e pollinators

volatiles might also protect the carbohydrate-rich nectar by inhibiting mivolatiles might also protect the carbohydrate-rich nectar by inhibiting microbial growth.crobial growth.

vegetative plant tissue release volatiles following herbivore damage. vegetative plant tissue release volatiles following herbivore damage. Some of these substances attract arthropods that prey upon or Some of these substances attract arthropods that prey upon or

parasitize the herbivores. parasitize the herbivores. Volatiles also act as direct repellents or toxicants for herbivores and Volatiles also act as direct repellents or toxicants for herbivores and

pathogens.pathogens.In fruits, volatile emission and accumulation facilitate seed dispersal by In fruits, volatile emission and accumulation facilitate seed dispersal by

animals and insects.animals and insects.vegetative tissues often produce and release many of the vegetative tissues often produce and release many of the

volatiles after their cells are disrupted. These volatile flavor volatiles after their cells are disrupted. These volatile flavor compounds may exhibit anti-microbial activity.compounds may exhibit anti-microbial activity.

““associated with defensive and attractive rolesassociated with defensive and attractive roles””

Aromatic compounds responsible for od or and flavor of fruits comprise;

Alcohols

Carbonyls

Acids

Esters

Lactones

Phenols

R-OH

R-CHOR-CO-R’

-R COOH

- -R COO R’

R

O CO

Estimated Estimated world world

consumption consumption of selected of selected

aroma aroma chemicals in chemicals in

flavor and flavor and fragrance fragrance

compositionscompositions

CHO

OH

OCH3

Calvin cycle

N enters roots as NO3- or NH4+. The NH4+ is incorporated into amino acids in roots and leaves and the amino acids accumulate in proteins. The main if not sole function of some proteins is to provide a store of amino acids

Amino acid synthesis

Amino acid synthesis

Glycolysis

isoprenoid biosynthesis proceeds either via the "classical" or most well studied, mevalonate pathway (cytosolic) (for the synthe

sis of sterols, sesquiterpenes, triterpenoids)

or via the non-mevalonate (1-deoxy-D-xylulose-5-phosphate, DXP) pathway for plastidic isoprenoids (carotenoids, phytol [side-chain of chlorophylls], plastoquinone, isoprene, monoterpenes a

nd diterpenes).

Biosynthesis of flavors in vegetables an d fruits

develop when tissue damage occurs

(Intact vegetable generally contains few volatiles)

Vegetable flavors

are formed during brief ripening period

Fruit flavors

Minute quantities of lipids, CHO, protein (amin o acids) are enzymatically converted to volatile fl

avors.

BIOGENESIS OF FRUIT AROMA

develops entirely during ripening period of plant

FRUIT FLAVOR COMPOUNDS

AppleApple n-hexanal, ethyl butyrate, 1-propyl n-hexanal, ethyl butyrate, 1-propyl propionate, 1-butyl acetate, trans-2-propionate, 1-butyl acetate, trans-2-hexenal, ethyl 2-methylbutyrate, 2-hexenal, ethyl 2-methylbutyrate, 2-methylbutyl acetate, 1-hexanol, hexen-methylbutyl acetate, 1-hexanol, hexen-1-ol, trans-2-hexen-1-ol, hexyl acetate, 1-ol, trans-2-hexen-1-ol, hexyl acetate, Esters; alcohols; aldehydes; ketone; Esters; alcohols; aldehydes; ketone; acids; including hexanal; ethyl 2-acids; including hexanal; ethyl 2-methyl butyrate methyl butyrate

BananaBanana alcohols; esters, including amyl acetate, alcohols; esters, including amyl acetate, isoamyl acetate, butyl butyrate, amyl b isoamyl acetate, butyl butyrate, amyl b ut yr at e ut yr at e

PeachPeach - -Ethyl acetate, dimethyl disulfide, cis 3 - -Ethyl acetate, dimethyl disulfide, cis 3 hexenyl acetate, methyl octanoate, eth hexenyl acetate, methyl octanoate, eth

- yl octanoate, 6 pentyl alpha pyrone, ga - yl octanoate, 6 pentyl alpha pyrone, ga mma decal act one mma decal act one

Lipids Polysaccharide Proteins/Enzymes Lignins

Aliphatic Terpenes Methyl-Branched AromaticAcidsAlcoholsEstersCarbonylslactones

SesquiterpenesHydrocarbonsAlcoholsCarbonylsmonoterpenes

AlcoholsAcidsEsterscarbonyls

AlcoholsAcidsEsterscarbonyls

Biosynthesis of fruit volatiles

Carbohydrate Amino

acid

Cinnamicacid

Terpene

Fattyacid -AcetylCoA

MalonylCoA Acetyl C

oA

Pyruvate

MevalonylCoA

Shikimic acid

Flavorants from carbohydrate metabolism

Furanones and pyrones

“fruit constituents”

O nly a limited number of natural vol atiles originate directly from carboh

ydrates without prior degradation of the carbon skeleton.

Furanones and pyrones

- - - -4 25Carbohydrate derived flavor molecules, including hydroxy , di- - - - - - -ff3 (2 ) ( ), 2 ,5 4 3 ( 2 )

- - - - - 4 5 32ranone (methoxyfuraneol), hydroxy methyl ( H) furanone (nor - - - - - - - 2 4 5 32furaneol), ethyl hydroxy methyl ( H) furanone (homofurane

- - - - - - - 4 2 5 32ol), hydroxy methylene methyl ( H) furanone (HM M F) and- - - - - - - 3 2 4 4hydroxy methyl H pyran on (maltol).

Glycolysis

Glucose (6C)

2 Pyruvate (3C)

Ethanol LactateTCA Cycle

CO2

+O2

-O2 -O2

Flavorants from carbohydrate metabolism

Pyruvic acid CH3COCOOH

Acetic acid CH3COOH

Acetyl CoA CH3COSCoA

+ CO2

Malonyl CoA HOOCCH2COSCoA

Malonic Acid HOOCCH2COOH

“ the most interesting is terpene biosynthesis”

Terpenoids are enzymatically synthesized from acetyl CoA and pyruvate provided by the carbohydrate pools in

plastids and the cytoplasm.

Terpenoids constitute one of the most diverse families of

natural products, with over 40 000 different structures of

terpenoids

Many of the terpenoids produced are non-volatile and are involved in important plant processes such as membrane

structure (sterols), photosynthesis (chlorophyll side chains, carotenoids), redox chemistry (quinones ) and growth

regulation (gibberellins, abscisic acid, brassinosteroids)

Flavorants from carbohydrate metabolism

Important plant-derived volatile terpenoids.

Biosynthesis of Terpenes

“ -isoprene is derived from acetyl CoA”

Classification of Terpenes

Apocarotenoid formationApocarotenoid formation

Carotenoid substrates are oxidatively cleaved to yield the apocaroten oid derivatives (right).

Some of the volatile organic compounds in wine come from the grape's skin, or exocarp, while others come from the grape's flesh, or mesocarp. Organic acids give wine its tartness, and sugars give it sweetness. Terpenes provide floral or fruity flavors. Norisoprenoids impart a honeylike character. Thiols are the sulfur-based

compounds behind complex wine aromas such as guava, passionfruit or grapefruit — but when thiols go wrong, they can make a wine taste "funky."

products; acids, alcohols, diketones, ketones, esters of these compounds.

Lipids

metabolic pathway for lipid biosynthesis plays a significant role in flavor formation.

Alpha-, -Beta oxidation

Oxidation via lipoxygenase

Lipoxygenase activity is believed to be the major source of volatiles in plants.

Oxidation via Lipoxygenase

Major products : volatile C6 and C9 aldehydes and alcohols

Substrate : unsaturated fatty acid ( linoleic and linolenic acids).

Lipoxygenase enzymes (dioxygenase) catalyze reactions between O2 and polyunsaturated fatty acids

- Linolenic acid derived flavor molecules.

AAT, al cohol acyl CoAt ransf erase ; ADH, al cohol dehydrogenase; AE R, alkenal oxidoreductase; AOC, allene oxide cyclase; AOS, allene oxi

de synthase; HPL, hydroperoxide lyase; JMT, jasmonate methyltransferase; LOX, lipoxy

- - - -genase; OPR, 12 oxo phytodienoic acid reductase; 3Z,2E EI, 3Z,2E e nal isomerase.

Fatty acid precursors (Tomato)

- and -oxidation of fatty acids

- Palmitoyl CoA (1 6 :0 )

- Myristoyl CoA (1 4 :0 )

+ Acetyl-CoA

the specific pathways in plants are not well understood

-Formation of pear flavors via beta oxidation

Lactones

Amino Acid Metabolism

Amino acid metabolism yields short chain aliphatic and aromatic alcohols, acids, carbonyls and esters

They are the primary source of branched chain aliphatic flavor compounds

their pathways have been barely analyzed in plants.

amino acid precursors(Tomato)

- (a) Catabolism of branched chain amino acids leading to methyl branched f lavor compounds, and (b) postulated biosynthesis of sotolon. Formation of

aldehyde (a) from amino acids requires the removal of both carboxyl and a mino groups. The sequence of these removals is not fully known and could b

e the opposite to that shown or aldehyde could be formed in one step by ald ehyde synthase

- Biosynthesis of amino acid derived flavor compounds

Starting amino acids: Tyrosine and phenylalanine products: phenolic/spicy in character

Shikimic acid formation

Vegetable Flavors

Vegetable flavors

- flavor again arises from major metabolic processes e.g. Lipids, CHO & amino acids.

The role or importance of S compounds to v egetable flavor is quite significant.

the precursors, enzymes and end flavors are quit e different from fruits.

Nonvolatile PrecursorsLinoleic, Linolenic acid Thioglucosinolates Cysteine-sulfoxides Methyl-methionine

Precursor-splitting EnzymesLypoxygenase Thioglucosidases C-S-lyases None (Heating)

CarbonylsAlcoholsOxo-acids

IsothiocyanatesNitrilesS C OThiocyanates

PolysulidesAlkyl-thosulfinates

CH3-S-CH3

Carbohydrate

Fatty acid Amino acid

Formation of flavor in vegetables

Vegetable Flavor Categories

Genus Allium

Enzymes produce volatiles from derivatives of cysteine (sulfoxides)

Genus Brassica

Enzymes produce volatiles from glucosinolates

Alliaceous vegetables

garlic ( Allium sativum L.)

onion ( Allium cepa L.)

chive ( Allium schoenoprasum L.)

leek ( Allium porrum L.)

Characteristic flavors

not exist in the bulb before processing

are produced when the cellular tissues are ruptured by cutting or chewing

flavor is produced very rapidly by the action of an enzyme on the odorless precursors wh ich coexist in the cells

Onion and Garlic Flavor

Enzymatic reaction of cysteine derivative

Glucosinolate precursors are important to the flavor of both the Brassica and Cruciferae fami

ly

Cruciferae family includes radish, horseradish,mustard.

GLUCOSINOLATES

thiocyanate, nitrile, or isothiocyanate

& glucose

Hydrolysis of the glucosinolate

glucosinolate

thioglucosidase

Natural carbon pools for the Natural carbon pools for the productionproduction

of flavor compounds, and the of flavor compounds, and the pathwayspathways

Mevalonic acid Acetate Mevalonic acid

Isoprene

Shikimic acid Pyruvate + Erythrose phosphate

“ the most interesting is terpene biosynthesis”

most of essential oils get flavor from terpenoids (10 carbon)

- Limonene a monoterpene hydrocarbon - is the major terpene in many or most citrus products. Orange > 95% of the essenti al oil is limonene,

lemon ~ 65% limonene, yet is of little flavor significance.

- Citral oxygenated monoterpene - seldom comprises > 2% - of the essential oil of lemon largely carries the lemon flavo

r.

Flavorants from carbohydrate metabolism

1 Methane 1 Methane 11 Undecane 11 Undecane 2 1 Henicosane 2 1 Henicosane 31 Hentriacont 31 Hentriacontaneane

2 Ethane 2 Ethane 12 Dodecane 12 Dodecane 22 Docosane 22 Docosane 32 Dotriaconta 32 Dotriacontaffff

3 Propane 3 Propane 13 Tridecane 13 Tridecane 23 Tricosane 23 Tricosane 33 Tritriaconta 33 Tritriacontaffff

ffffff4 ffffff4 14 Tetradecan 14 Tetradecanff 24 Tetracosane 24 Tetracosane 40 Tetracontan 40 Tetracontan

ff

fffffff5 fffffff5 15 Pentadecan 15 Pentadecanff

25 Pentacosan 25 Pentacosanff

50 Pentaconta 50 Pentacontaffff

6 Hexane 6 Hexane 16 Hexadecan 16 Hexadecanff 26 Hexacosane 26 Hexacosane 60 Hexacontan 60 Hexacontan

ff

7 Heptane 7 Heptane 17 Heptadecan 17 Heptadecanff

27 Heptacosan 27 Heptacosanff

70 Heptaconta 70 Heptacontaffff

8 Octane 8 Octane 18 Octadecane 18 Octadecane 28 Octacosane 28 Octacosane 80 Octacontan 80 Octacontanff

f fffff9 f fffff9 19 Nonadecan 19 Nonadecanff 29 Nonacosane 29 Nonacosane 90 Nonacontan 90 Nonacontan

ff

10 Decane 10 Decane 20 Icosane 20 Icosane    30 Triacontane 30 Triacontane 100 Hectane 100 Hectane

Isoamyl acetate, a strong fruity odor described as similar to banana or pea

r - - 2 M ethyl butyl acetate has a strong a

pple scent