Supercritical Fluid Extraction and

Separation of Active Components

from Medicinal Plants
Béla Simándi
Budapest University of Technology and Economics

SOCRATES Intensive Course, Barcelona, 2004

 SCOPE of presentation
• Supercritical Fluid Extraction (SFE)
 SCOPE of presentation
• Supercritical Fluid Extraction (SFE)
• Active principles of plants
Active principles of plants

terpenoids
alkaloids
fats and waxes
 SCOPE of presentation
• Supercritical Fluid Extraction (SFE)
• Active principles of plants
• SFE of plant constituents
SFE of plant constituents
essential oils
some non-volatile terpenoids
triterpenes and steroids
carotenoids
alkaloids
plant lipids
 SCOPE of presentation
• Supercritical Fluid Extraction (SFE)
• Active principles of plants
• SFE of plant constituents
• Economic issues
• Conclusions
 Supercritical fluids
• Carbon dioxide (PC=73.8 bar, TC=31.1°C)
 Carbon dioxide
• GRAS (generally regarded as safe)
• gentle conditions (PC=73.8 bar, TC=31.1°C)
• inert, odourless, tasteless
• easily removed from products
• non-explosive
• readily available, inexpensive
• environment-friendly
 Supercritical fluids
• Carbon dioxide (PC=73.8 bar, TC=31.1°C)
• Nitrous oxide (PC=71.0 bar, TC=36.5°C)
• Propane (PC=42.5 bar, TC=96.7°C)
 Propane
• near-critical liquid (TC=96.7°C)
• flammable, explosive mixtures with air
• maximum permitted residue: 50 ppm
• significant solvent loss
• waste solvent
 Supercritical fluids
• Carbon dioxide (PC=73.8 bar, TC=31.1°C)
• Nitrous oxide (PC=71.0 bar, TC=36.5°C)
• Propane (PC=42.5 bar, TC=96.7°C)
• Water (PC=220.5 bar, TC=374.2°C)
 Supercritical fluids
• Carbon dioxide (PC=73.8 bar, TC=31.1°C)
• Nitrous oxide (PC=71.0 bar, TC=36.5°C)
• Propane (PC=42.5 bar, TC=96.7°C)
• Water (PC=220.5 bar, TC=374.2°C)

• Entrainers (ethanol, ethyl acetate, acetone)

Multipurpose extraction unit
Formation of secondary plant products
Process of primary metabolism
CO2 H2 O

Pentose cycle

Saccharides

Glycolysis

Phosphoenol-pyruvic acid

Pyruvate

Acetyl CoA

Krebs cycle

CO2
 Products of primary metabolism
Process of primary metabolism Oligo and polisaccharides
CO2 H2O

Pentose cycle Eritrose-4-PO4

Saccharides Shikimic acid
Glycolysis
Aromatic amino acids
Phosphoenol-pyruvic acid Proteines
Aliphatic amino acids
Pyruvate

Acetyl CoA Malonyl CoA

Fatty acids
Krebs cycle

CO2 IPP biological isopren

Squalene
 Secondary plant metabolism
Process of primary metabolism Products of primary metabolism Glycosides
CO2 H2O Oligo and polisaccharides Mucilages and gums
Pentose cycle
Eritrose-4-PO4 Phenolic compounds
Saccharides
Shikimic acid Lignans
Glycolysis Alkaloids
Aromatic amino acids
Phosphoenol-pyruvic acid
Aliphatic amino acids
Alkaloids
Pyruvate Peptides
Malonyl CoA
Acetyl CoA Fatty acids Fats and waxes
IPP biological isopren Tetracyclynes
Krebs cycle
Antraquinones
CO2 Squalene

Terpenes
 Terpenoids
Isoprene (C5) units

Isoprene does not participate in the

biogenesis
 Terpenoids
• C10 monoterpenes (essential oil)
• C15 sesquiterpenes (essential oil)
• C20 diterpenes (antioxidants)
• C30 triterpenes (phytosterols)
• C40 tetraterpenes (carotenes)
• (C5)n polyterpenes (caoutchouc)
 Solubility

• typical
essential oil
components
 Comparative chemical composition
of distilled oils and SFE products
Plant Compound Steam SFE
distillation
Dill limonene 55.7 42.6
(Anethum graveolens L.) d-carvone 36.5 48.6
Coriander pinene 15.3 7.7
(Coriandrum sativum L.) linalool 68.5 75.5
Celery limonene 50.5 33.4
(Apium graveolens L.) 3-butylphthalide 23.6 40.6
Parslay α-pinene 24.0 1.5
(Petroselinum crispum L.) β-pinene 21.9 3.0
myristicin 7.4 4.0
apiole 38.5 84.9
 Alteration of essential oil
components during distillation
linalyl acetate → linalool
lavandin (Lavandula intermedia Emeric)
clary sage (Salvia sclarea L.)
glycosides → thymol
thyme (Thymus vulgaris L.)
unknown precursor → α-terpineol
terpinene-4-ol
lavandin (Lavandula intermedia Emeric)
pulegone → dihydrocarvone
spearmint (Mentha spicata L.)
 Fractionation of fennel oil
(Foeniculum vulgare Mill.)
 Fractionation of fennel oil
(PE=302 bar, TE=38°C)
1st separator:75 bar, 23.5 °C

1st separator:86 bar, 23.5 °C

Effects of the separation parameters
on fractionation of fennel oil
42

38

34

0.75
T S , °C

30
1
1.5
2 26
2.5
3
4 22
5
6 18
7 68 72 76 80 84 88 92

P S, bar
 Composition of orange oil

Component Composition (%)

α-Pinene 0.45
Myrcene 1.77
d-Limonene 90.60
Octanal 0.59
Decanal 0.52
Linalool 0.37
Geranial 0.12
Phase equilibria of orange peel oil
Separation factors of orange peel
oil fractions
 Fractionation of citrus oil

Feed

Terpenes

CO2
CO2

Aroma
 Results of fractionation

Pilot plant data on deterpenation of orange oil

Aromatic content in feed oil 4.1 %
Aromatic content in 18.9 %
concentrated oil
Recovery of aromatic fraction 90 %
Solvent-to-oil ratio 100
SFE of sesquiterpene lactones
OH OH
HO O β -D-glucose-O O

OH O OH O
apigenin apigenin-7-glucoside

OH
O
O
OH OH
(-)-α-bisabolol (-)-α-bisabololoxide B
(-)-α-bisabololoxide A chamazulene

O O
O
O HO OCOCH3
O
O
cis-en-in-dicycloether trans-en-in-dicycloether matricine
Fractionated extraction of
chamomile

P T Matricine α-bisabolol Spiroether Yield

(bar) (° C) (%) (%) (%) (%)
80 40 3.5 27 14 0.12
90 40 17.0 21.0 16.7 0.4
300 40 7.5 9.2 11.7 1.65
SFE of feverfew: effects of pressure
and temperature on the yield
Recovery of parthenolide

O
O
O
parthénolide
Diterpenes investigated by SFE
O
OH
O O OH
HO
H O
O
H
O O O O
HO HO
O

baccatin-III carnosol
OH
HO
O
OH

O OH
OH

rosmanol sclareol
O
O
OH
O

O O
O
O O O
N OH O
H O
OH

taxol
 Isolation of diterpenes
from plants
Plant Component

Clary sage sclareol

(Salvia sclarea L.)
Yews paclitaxel
(Taxus brevifolia Nutt., baccatin III
Taxus cuspidata Capita)
Rosemary rosmanol
(Rosmarinus officinalis L.), 7-methyl-epirosmanol
Sage carnosol
(Salvia officinalis L.) carnosolic acid
Basic sceleton of triterpenes

H H

HO HO

α-amirin β-amirin

H
OH

HO
HO

faradiol β-sitosterol
 Isolation of triterpenes
Plant Component

Mulberry α-amyrin acetate

(Morus alba L.)
Neem nimbin
(Azadirachta indica A. Juss) salannin
azadirachtin
Marigold faradiol and its esters
(Calendula officinalis L.)
Chaste tree β-amyrin
(Vitex agnus castus L.) β-sitosterol
Dandelion β-amyrin
(Taraxacum officinale Web.) β-sitosterol
 SFE of dandelion leaves:effects of
pressure and temperature on the yields
Recovery of sterols from
dandelion leaves
Recovery of carotenoids
HO
O

bixin (annetto)
O O

OH

HO
capsanthin (paprika)

β-carotene (carrot)
OH

HO
lutein (alfalfa)

lycopene (tomato)
SFE of pigments from paprika

10,589
11,214
11,839
12,464
13,089
13,714
14,339
14,964
15,589
16,214
16,839
17,464
18,089
18,714
19,339
above
 Extraction of hop
(Humulus lupulus L.)

Water Organic solvent Steam distillation

Water extract Total resin Essential oil

Petrol soluble Petrol insoluble

‘Soft resin’ ‘Hard resin’

α acid β acid Uncharacterised

acid
Structure of major components
OH O
OH O
R
R
O OH HO
OH OH
O

α acid iso α acid

OH O
H 3C CH3
R
CH3 CH3
O OH R=
CH3
CH3
H3C
CH3
H3C
β acid
Composition of hop extract
versus time
 Alkaloids: decaffeination of coffee

• Solubility
isobars of caffeine
Decaffeination of green coffee beans
Nearly continuous SFE of coffee
 Extraction of nicotine from tobacco
Adsorbent Nicotine
CO2 H2 O by-product
regeneration
Aroma Nicotine
Tobacco removal removal
Drying
Aroma Aroma
transfer

Aroma CO2
distribution

Nicotine reduced
Conditioning tobacco
Extraction of alkaloids used
in medicine
Plant Alkaloid Recovery
paprika capsaicin good
(Capsicum annuum L.)
pepper piperine good
(Piper nigrum L.)
poppy opium reasonable
(Papaver somniferum L.) (codein,
thebaine,
papaverin)
strychnos strychnine reasonable
(Strychnos nux-vomica L.)
hemlock fruit coniine reasonable
(Conium maculatum L.)
ipecacuanhae radix emetin, poor
(Cephyaelis ipecacuanha Will.) caphaelin
 Fatty oils and waxes

• Solubility
isotherms of
soybean oil
 Effect of alcohol-entrainer on
extraction rate
g oil / 100 g dry corn germ
60

50

40

30 0 % alcohol
2.5 % alcohol
20
5 % alcohol
10 7.5 % alcohol
10 % alcohol
0
0 20 40 60 80 100 120 140 160
kg CO2 / kg dry corn germ
 List of raw materials for
γ-linolenic acid
Raw material Yield CO2 C18:3 in the extract
extraction (γ-linolenic acid)
[%] [%]
Oenothera biennis L. 22 8 - 12
Borago officinalis L. 18 18 - 22
Humulus lupulus L. 6 3-4
Cannabis sativa L. 35 3-6
Ribes rubrum L. 14 4-6
Ribes negrum L. 18 16 - 19
Ribes grosullaria L. 15 10 – 12
Rosa canina L. 12 24 - 31
Hippophae rhamnoides L. 12 26 - 30
 Partition coefficient and
selectivity of fish oil ethyl esters
Component K=y/x KA/K22
C16 0.090 7.5
C18 0.050 4.2
C20 0.027 2.3
C22 0.012 1.0
 Two-column process for
separation of EPA and DHA
Invesment costs of high pressure units
100
I=A(VTW)0.25
10

SFE SFI SFE/SFF PA

0.1
VTW
0 5 10
Production costs of SFE
Comparison of solvent extraction
with SFE
Plant capacity Production cost Notes
(t/year) (EUR/kg raw material)
SE SFE
300-400 3-8 4-10 medicinal plants,
cosmetics, spices
1000-1200 1-3.5 2-5 food additives,
specific vegetable
oils
10000-12000 0.5-1.2 0.75-1.2 coffee, hop
100000-120000 0.2-0.4 ? oilseeds
 CONCLUSIONS
• There is a growing interest in new natural
products that act very specifically without
any side-effects.
• Supercritical fluid extraction is ideal for
preparation of medicinal plant extracts.
• Carbon dioxide extracts are different from the
traditional preparations not only in chemical
composition, but in phytotherapeutical activity
as well.
• There is an urgent need for more detailed
phytochemical analysis of extracts.
 CONCLUSIONS
• Commercial supercritical fluid processes
have been still limited to few areas.
• Large capacity plants, with optimised design
and operation lead to prices that are very
often of the same order of magnitude as
those related to classical processes.
• Moreover, there are also cases were
supercritical fluids permit to make products
or operations, that cannot be realised by any
other methods.