SKILL 3RD SEMESTER
ROLE OF MEDICINAL AND AROMATIC PLANTS IN HEALTH CARE SYSTEM
Dr. Shah Rafiq
Topics:
Methods of isolation of major active constituents from medicinal plants (cold and Soxhlet
extraction), preparation of active-content-enriched extracts.
Latest methods of extraction of essential oil.
Methods of Isolation of Major Active Constituents from Medicinal Plants (Cold Extraction and
Soxhlet Extraction)
Medicinal plants contain many kinds of chemical compounds. Some are primary metabolites (sugars,
proteins, lipids), and others are secondary metabolites, which are responsible for pharmacological
activity.
Major bioactive constituents
Alkaloids (e.g., morphine, atropine, quinine, reserpine)
Glycosides (e.g., digoxin, sennosides, aloin)
Flavonoids and phenolics (e.g., quercetin, rutin, tannins)
Terpenoids and essential oils (e.g., menthol, eucalyptol)
Saponins, steroids, resins and coumarins
Extraction is the process of separating the medicinally active constituents of plant tissue from the
inactive or inert material (cell wall matter, cellulose and so on) using a suitable solvent and a suitable
method. The product is called the extract, and the leftover plant material is the marc. The solvent
used is also called the menstruum.
Objectives of extraction
1.
To obtain therapeutically active components in concentrated form.
2.
To remove inert or unwanted material.
3.
To prepare the crude drug for standardisation and formulation (tinctures, fluid extracts, dry
extracts).
4.
To prepare the sample for isolation and structure elucidation of pure compounds.
General Scheme for Isolation of Bioactive Compounds
1.
2.
3.
4.
5.
6.
7.
8.
Collection and authentication of plant material
Drying and size reduction
Selection of solvent
Extraction (cold or hot method)
Filtration and concentration of the extract
Fractionation (liquid-liquid partitioning, precipitation)
Purification (column chromatography, preparative TLC, HPLC, crystallisation)
Characterisation (TLC, UV, IR, NMR, MS)
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Pre-extraction Preparation of Plant Material
a)
Collection and authentication
The correct species, plant part, season and stage of growth must be used, because the content of
constituents varies with these factors.
A botanist should authenticate the plant, and a voucher specimen should be kept.
b)
Drying
Drying stops enzymatic degradation and microbial growth.
Shade drying is used for thermolabile compounds and volatile oils.
Oven drying at 40 to 50 °C or sun drying may be used for stable constituents.
Fresh material is sometimes used directly for volatile constituents, but its water content dilutes
the solvent.
c)
Size reduction (grinding or powdering)
Grinding increases the surface area and ruptures cell walls, which improves solvent
penetration.
The powder should be moderately coarse to fine (commonly No. 20 to No. 60 sieve).
Very fine powder can clog percolators and thimbles and make filtration difficult. Coarse
powder gives poor extraction efficiency.
d)
Defatting (when required)
Seeds and other oily materials are first extracted with a non-polar solvent such as petroleum
ether or hexane, so that fats do not interfere with later extraction.
Selection of Solvent
The governing rule is "like dissolves like": polar compounds dissolve in polar solvents and non-polar
compounds in non-polar solvents.
Characteristics of an ideal solvent
Selective for the target compounds
Chemically inert (does not react with the extract)
Low boiling point, so it is easily removed
Non-toxic, non-flammable and inexpensive
Does not cause complex formation or degradation
Solvents in increasing order of polarity
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Solvent
Polarity
Petroleum ether /
Hexane
Very low
Chloroform
Low
Alkaloid bases, terpenoids, some glycosides
Diethyl ether
Low
Volatile oils, alkaloids, terpenoids
Ethyl acetate
Medium
Flavonoid aglycones, phenolics
Acetone
Medium-high
Flavonoids, phenolics, tannins
Unit II
Typically extracts
Fats, waxes, oils, sterols, carotenoids
Dr. Shah Rafiq
Solvent
Ethanol
Methanol
Water
Polarity
Typically extracts
High
Alkaloids, glycosides, flavonoids, tannins,
saponins
High
Broad range, including polar glycosides
Sugars, glycosides, tannins, saponins, gums,
mucilage
Note: Ethanol and water-ethanol mixtures are the most widely used in industry because they are
relatively safe. Methanol extracts a wide range of compounds but is toxic.
Very high
Classification of Extraction Methods
a.
Cold (room temperature) methods: maceration (simple, multiple and with agitation) and
percolation.
b.
Hot methods: decoction, infusion and digestion, Soxhlet (continuous hot) extraction, reflux
extraction, steam distillation.
c.
Modern methods: ultrasound-assisted extraction (UAE), microwave-assisted extraction
(MAE), supercritical fluid extraction (SFE, with CO2), pressurised liquid extraction.
Cold Extraction
Cold extraction is carried out at room temperature without applying heat. It is best suited to
thermolabile constituents and is therefore called
a "gentle" method.
Maceration
Principle: The coarse or powdered drug is kept
in contact with the solvent (menstruum) in a
closed vessel for a set period, with occasional
shaking or stirring. The solvent penetrates the
cell wall and dissolves the soluble constituents,
and the solution diffuses out until equilibrium is
reached between the concentration inside and
outside the cells.
Apparatus: A wide-mouthed, closed container (glass jar or stoppered bottle) made of inert material.
Procedure (simple maceration)
1.
Place the weighed, coarsely powdered plant material in a closed container.
2.
Add the solvent (commonly about 3 to 5 times the weight of the drug, or enough to cover it
completely).
3.
Close the vessel and keep it at room temperature for 3 to 7 days with frequent shaking or
stirring.
4.
Strain or filter the liquid. Press the marc to recover the remaining liquid.
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5.
Combine the liquids, clarify by filtration or by standing and decanting, and concentrate.
Types of maceration
Simple maceration: The procedure above, used for most drugs and for tinctures.
Multiple maceration (double or triple): The solvent is divided into portions. The drug is
macerated with the first portion and strained, and the marc is macerated again with the second
portion. This gives more complete extraction than one large volume.
Maceration with agitation (dynamic maceration): Occasional or mechanical stirring, for
example with a shaker, increases the rate of diffusion and shortens the time.
Advantages
Simple, inexpensive and needs minimal equipment
Suitable for thermolabile constituents
Little chance of thermal decomposition
Disadvantages
Time-consuming (days)
Extraction is often incomplete, because equilibrium is reached and some drug is left in the
marc
Uses a large volume of solvent
Solvent may be lost by evaporation if the container is not tightly closed
Examples of use: Tincture of ginger, tincture of orange, tincture of tolu, and extraction of colored
pigments and thermolabile flavonoids.
Percolation
Principle: A continuous downward flow
of fresh solvent through a bed of drug in a
conical vessel (percolator). Because fresh
solvent is always in contact with the drug,
the concentration gradient stays high, giving
more complete extraction than maceration.
Apparatus: A percolator, a narrow conical
or cylindrical vessel with a stopcock at the bottom.
Procedure
1.
Moistening (imbibition): The powder is moistened with enough solvent to make it evenly
damp and left for about 4 hours in a closed container so that it swells and the solvent
penetrates.
2.
Packing: The moistened drug is packed uniformly into the percolator over a cotton or filter
pad, neither too loose nor too tight.
3.
Maceration (soaking): Solvent is added until it stands above the drug bed. The stopcock is
closed and the drug is macerated for about 24 hours.
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4.
Percolation: The stopcock is opened and the extract is drawn off slowly, usually at a rate of
about 1 mL per minute per 1000 g of drug. Fresh solvent is added continuously so that the drug
is always covered by a layer of solvent.
5.
Collection continues until the drug is exhausted, meaning the percolate no longer gives a test
for the constituents.
Reserved percolation: The first portion of the strong percolate is set aside as "reserved". The later,
weaker percolate is concentrated at low temperature and added to the reserved portion. This
avoids heating the most concentrated extract.
Advantages
More complete extraction than maceration
Suitable for potent drugs and for preparing fluid extracts and tinctures
Disadvantages
Needs more skill and more solvent
The extract may be dilute
Not suitable for very fine powders (channelling or blocking of the bed)
Soxhlet (Continuous Hot) Extraction
Principle
Soxhlet extraction is a continuous extraction with
recycled solvent. The solvent is heated, its vapour
condenses, and the condensed liquid repeatedly
percolates through the solid sample held in a thimble.
When the thimble chamber is full, a siphon empties it
back into the flask. Each cycle uses fresh, distilled
solvent, so extraction proceeds as though a large
volume of solvent were used, while only a small volume
is actually needed. The extracted compounds are nonvolatile, so they accumulate in the flask as pure solvent
is redistilled.
Apparatus
A Soxhlet apparatus has three main parts:
1.
2.
3.
Round-bottom flask at the bottom, containing the solvent and boiling chips or a stir bar,
sitting on a heating mantle or water bath.
Soxhlet extractor (extraction chamber) in the middle, containing the thimble with the
sample, a vapour (bypass) side arm and a siphon tube.
Condenser (reflux condenser, usually a water-cooled Liebig or Allihn type) on top. The thimble
is made of thick filter paper or cellulose. It must be porous enough for the solvent to pass but
must retain the plant powder.
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Procedure (Step by Step)
1.
Weigh the dried, coarsely powdered plant material (for example, 20 to 50 g).
2.
Place the powder in the thimble and plug the top with cotton wool so that no powder floats out.
3.
Place the thimble in the extraction chamber, ensuring the drug is below the top of the siphon
arm.
4.
Fill the round-bottom flask to about one-half to two-thirds with solvent and add boiling chips.
5.
Assemble the apparatus and start the flow of cooling water through the condenser.
6.
Heat the flask at a controlled rate. The solvent boils, its vapour rises through the side arm to
the condenser, condenses, and drips onto the drug in the thimble.
7.
The chamber fills with warm solvent, which dissolves the compounds. When the level reaches
the top of the siphon, the whole volume is siphoned back into the flask. This completes one
cycle.
8.
Repeat for a fixed number of cycles or hours (commonly 6 to 24 hours, at about 4 to 6
siphonings per hour) until the drug is exhausted. The solvent in the extraction chamber then
becomes colourless and gives a negative test for the constituents.
9.
Stop heating, allow to cool and dismantle the apparatus.
10. Concentrate the extract from the flask, usually by rotary evaporator under reduced pressure,
then dry it in a desiccator or vacuum oven.
Factors Affecting Soxhlet Extraction
Particle size: smaller particles give better extraction, but very fine powder can block the
thimble
Solvent type and polarity: must match the target compounds
Extraction time and number of cycles
Temperature: governed by the boiling point of the solvent
Sample-to-solvent ratio
Moisture content of the sample: high moisture reduces efficiency with non-polar solvents
Rate of heating: too fast a rate causes flooding and poor contact
Advantages
The solvent is recycled, so a small volume gives an efficient, exhaustive extraction
Very high extraction efficiency, more complete than maceration
The drug is always in contact with fresh, warm solvent
Requires little supervision once set up
Very suitable for non-polar and moderately polar compounds such as lipids, essential oils
and alkaloids
Simple and widely used as a standard reference method
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Disadvantages
Thermal degradation of heat-sensitive compounds, because the extract stays at the boiling
point of the solvent for a long time
Long extraction times (hours)
Large amounts of solvent are consumed overall, and evaporating them takes energy
Not suitable for thermolabile constituents (some glycosides, volatile oils and certain
flavonoids)
The recycled solvent gives no agitation, so it is less suitable for compounds that are poorly
soluble in the solvent
Unsuitable for solvents with high boiling points, or for solvent mixtures whose composition
changes on distillation
Difficult to scale up for large industrial batches
Flammable solvents pose a fire hazard, so heating mantles or water baths should be used rather
than open flames
Common Uses
Determination of fat or oil content (petroleum ether)
Extraction of alkaloids (for example from Cinchona, Datura, Nux vomica)
Defatting of seeds before further extraction
Extraction of steroids, terpenoids and lipophilic phytoconstituents
Comparison of Cold Extraction and Soxhlet Extraction
Feature
Cold extraction (maceration
/ percolation)
Soxhlet extraction
Temperature
Room temperature
Boiling point of the solvent
Solvent handling
Static, or single pass
(percolation)
Recycled continuously
Time
Days (maceration); hours to a
day (percolation)
Hours
Solvent volume
Large
Small (recycled)
Efficiency
Lower (maceration) to
moderate (percolation)
High, near exhaustive
Suitability
Thermolabile compounds
Thermostable compounds
Risk of decomposition
Low
Possible
Equipment
Simple and cheap
Glassware, heat source, water supply
Scale
Easy to scale
Difficult to scale up
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Post-extraction Steps
1.
2.
3.
4.
5.
6.
Filtration: Filter the extract through filter paper or muslin cloth to remove particles.
Concentration: Remove the solvent using a rotary vacuum evaporator at low temperature
and reduced pressure (about 40 to 50 °C), which protects heat-sensitive compounds.
Drying: Dry in a desiccator, vacuum oven, or by lyophilisation (freeze-drying) to obtain a dry
extract.
Fractionation: Separate the crude extract into groups of compounds by liquid-liquid partition
between immiscible solvents (hexane, chloroform, ethyl acetate, butanol, water), acid-base
treatment (for alkaloids), or precipitation with solvents or reagents.
Purification: Column chromatography (silica gel or alumina), preparative TLC, HPLC and
recrystallisation.
Identification: TLC, UV, IR, NMR and mass spectrometry.
Example: Isolation of Alkaloids by Soxhlet Extraction
1.
2.
3.
4.
5.
Powder the dried plant material and moisten it with alkali (ammonia or lime). This liberates
free alkaloid bases from their salts.
Extract with an organic solvent (chloroform or ether) in a Soxhlet apparatus.
Shake the organic extract with dilute acid (dilute HCl or H2SO4). The alkaloids move into the
aqueous layer as water-soluble salts, and neutral impurities remain in the organic layer.
Make the aqueous layer alkaline. The free alkaloid bases precipitate or can be extracted back
into an organic solvent.
Concentrate and purify by chromatography or crystallisation.
II. Preparation of Active Content-Enriched Extracts
Medicinal plants contain hundreds of chemical constituents, but usually only a small group of them is
responsible for the therapeutic effect. A crude herbal extract therefore carries a large proportion of
inert or unwanted material, such as chlorophyll, waxes, sugars, proteins, and cellulose fragments,
alongside the constituents of interest. An active content-enriched extract is an extract that has been
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Dr. Shah Rafiq
deliberately processed so that the percentage of the bioactive constituent, or of a class of constituents,
is markedly higher than in the starting plant material or in a simple crude extract.
Key definitions
Term
Meaning
Crude extract
Concentrated preparation obtained by treating plant material with a
solvent and removing the solvent; contains the full spectrum of
extractable compounds.
Enriched extract
Extract in which the content of the active compound or class (for
example, total flavonoids, total saponins, curcuminoids) has been raised
by additional processing steps.
Standardized extract
Extract adjusted to a defined content of a marker or active constituent,
within stated limits, by blending or by process control. An enriched
extract is often also standardised.
Fraction
A portion of an extract separated based on polarity, size, acidity or
affinity (for example, the ethyl acetate fraction).
Isolated compound
A single, purified chemical entity (for example, berberine, quercetin);
purity is usually above 95%.
Active constituent
Compound(s) responsible for the therapeutic activity.
Marker
Chemically defined constituent used for quality control, whether or not
it is therapeutically active.
Drug-extract ratio
(DER)
Quantity of plant material used to make a unit quantity of extract; for
example, 10:1 means 10 kg of herb gives 1 kg of extract.
Why Enrich Extracts?
Consistent therapeutic dose: natural variability in plant content is reduced, so each dose
delivers a predictable amount of active.
Smaller dose volume: a higher active concentration allows a capsule or tablet of practical size
instead of large amounts of powder.
Improved efficacy and bioavailability: removal of interfering matrix components such as
tannins or polysaccharides can improve absorption and pharmacological response.
Improved safety: unwanted or toxic compounds (for example pyrrolizidine alkaloids,
aristolochic acid, excess furanocoumarins) can be depleted deliberately.
Better stability and formulation behaviour: less hygroscopic and less sticky material;
improved flow, compressibility and shelf life.
Regulatory and commercial requirements: nutraceutical and phytopharmaceutical products
commonly require a declared minimum marker content.
Economic advantage: lower transport, storage and processing costs per unit of active.
General Workflow: Preparation of an enriched extract follows a logical sequence. Each stage
affects yield and final marker content, so errors early on cannot be corrected later.
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1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
Selection and authentication of the plant material (botanical identity, correct part,
chemotype).
Collection, post-harvest handling and drying under conditions that protect the actives.
Size reduction to an optimum particle size.
Pre-treatment where necessary (defatting, enzymatic inactivation, hydrolysis, moistening).
Primary extraction with a suitable solvent and method.
Filtration, clarification and concentration of the miscella (liquid extract).
Enrichment or fractionation (partitioning, adsorption, precipitation, chromatography,
membranes and so on).
Final concentration and drying (vacuum drying, spray drying, freeze drying).
Standardisation with carriers or by blending batches to a fixed marker content.
Quality control, packaging and stability testing.
Enrichment Strategies
Primary extraction gives a mixture that is only partially enriched. Enrichment exploits differences in polarity,
acid-base behaviour, molecular size, solubility, adsorption affinity and volatility between the target
compounds and the accompanying matrix. Usually, several methods are combined in sequence.
Liquid-liquid partitioning (solvent fractionation)
The concentrated extract is suspended in water (or aqueous alcohol) and shaken successively with immiscible
organic solvents of increasing polarity, for instance n-hexane, then dichloromethane or chloroform, then ethyl
acetate, then n-butanol. Each layer collects compounds according to their partition coefficient (K =
concentration in organic phase/concentration in aqueous phase).
Fraction
Typically enriches
n-Hexane/petroleum ether
Lipids, waxes, chlorophyll, sterols, non-polar terpenoids
Dichloromethane/chloroform
Alkaloid free bases, less polar terpenoids, some flavonoid aglycones
Ethyl acetate
Flavonoid aglycones and less polar glycosides, phenolic acids, coumarins
n-Butanol
Saponins, polar flavonoid glycosides, iridoid glycosides
Residual aqueous
Sugars, amino acids, polysaccharides, tannins, very polar glycosides, salts
Practical
points:
emulsions can form,
especially with saponins
and proteins; they are
broken by adding salt,
centrifuging or standing.
Repeating the extraction
three times with smaller
volumes recovers more
solute than a single
large-volume extraction.
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Acid-base partitioning for alkaloids
This is the classic and most effective enrichment method for basic nitrogen compounds.
1.
Extract the plant material with acidified water or aqueous alcohol (for example, 1–5% hydrochloric,
sulfuric or tartaric acid). Alkaloids form salts and dissolve.
2.
Wash the acidic solution with a non-polar solvent (hexane or ether) to remove neutral and acidic lipophilic
impurities, which are discarded.
3.
Basify the aqueous phase with ammonia or sodium carbonate to about pH 9–10. Alkaloid salts convert to
free bases, which are now insoluble in water.
4.
Extract the basic solution with chloroform or dichloromethane (or ethyl acetate). Alkaloid bases move
into the organic layer.
5.
Wash, dry over anhydrous sodium sulfate and evaporate to give a total alkaloid fraction, often at 5–10
times or higher enrichment.
A similar principle applies to acidic compounds (phenolic acids, some lignans): extract at alkaline pH into water,
wash with an organic solvent, acidify, and re-extract into an organic solvent.
Adsorption and macroporous resin chromatography
Macroporous adsorption resins (non-polar polystyrene-divinylbenzene types such as the D101 and Amberlite
XAD series, or weakly polar and polar types) are the most widely used industrial method for enriching
flavonoids, saponins, phenolics, anthocyanins and glycosides from aqueous or hydroalcoholic extracts.
1.
2.
3.
4.
Resin pre-treatment: wash with ethanol and then water to remove monomers and porogens.
Loading: pass the crude aqueous extract through the resin column at a controlled flow (typically a few
bed volumes per hour). Target compounds adsorb through hydrophobic and hydrogen-bond
interactions; sugars, salts and very polar impurities pass through.
Washing: water (2–5 bed volumes) removes polar impurities.
Elution: stepwise gradient of ethanol in water (for example, 30%, 50%, 70%, 95%). The fraction with
the target compound is collected.
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5.
Regeneration: flush with higher ethanol concentration, and if necessary, dilute acid or alkali, then water.
Selection depends on the resin's surface area, pore diameter, polarity and the molecular size and polarity
of the target. Enrichment factors of 5- to 20-fold in total flavonoids or saponins with recoveries above
80% are commonly reported. The technique is scalable, uses relatively cheap ethanol and can be run
continuously.
Precipitation and crystallisation
Alcohol precipitation:
adding ethanol to an
aqueous concentrate to
70–80%
precipitates
polysaccharides, proteins
and pectins, which are
filtered off. This purifies
the supernatant that
contains
smaller
molecules; conversely,
polysaccharides
themselves
can
be
enriched
from
the
precipitate.
Water precipitation: pouring an ethanol extract into water precipitates resins and lipophilic
constituents; useful for removing resin or enriching such compounds.
Isoelectric or pH precipitation: compounds precipitate at their isoelectric point or as insoluble salts.
Salting-out: ammonium sulfate or sodium chloride reduces solubility of organic compounds.
Crystallization and recrystallization: achieved by cooling, solvent evaporation or anti-solvent
addition; this is the final step to obtain high-purity enriched products such as curcuminoids or rutin.
Traditional lead acetate precipitation of tannins is now avoided because of toxicity; gelatin,
polyvinylpolypyrrolidone (PVPP) or resin adsorption are used to remove tannins.
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Membrane separation
Pressure-driven membranes separate molecules by size and are attractive because they operate at ambient
temperature without phase change.
Process
Approximate cutoff
Application
Microfiltration
0.1–10 µm
Removes suspended solids and microorganisms
Ultrafiltration
Roughly 1–500 kDa
Removes polysaccharides, proteins, and colloids; clarifies
extracts
Nanofiltration
Roughly 200–1000
Da
Retains small molecules such as flavonoids and saponins;
passes salts and solvent; also concentrates without heating
Reverse osmosis
Below about 200 Da
Concentration of aqueous extracts
Fractional extraction with
supercritical CO₂
By stepping pressure or by using staged
separators, non-polar compounds (volatile
oil, waxes and pigments) can be fractionated
in sequence, leaving an enriched residue or
fraction. In turmeric, for example, the
volatile oil is first removed with pure scCO₂,
and the curcuminoids are then extracted
using ethanol-modified CO₂.
III. Latest Methods of Extraction of
Essential Oils
Essential oils (EOs) are complex mixtures of volatile, lipophilic secondary metabolites produced by
aromatic plants. They give plants their characteristic odour and flavour and function in defence against
herbivores and pathogens, in attracting pollinators and in allelopathy. They are called “essential”
because they carry the essence (fragrance) of the plant, not because they are nutritionally essential.
Essential oils are obtained from flowers, leaves, bark, wood, roots, rhizomes, seeds, fruit peels and
resins. Typical examples are lavender, peppermint, eucalyptus, rose, jasmine, sandalwood, clove,
cinnamon, lemongrass, citrus peel, tea tree and palmarosa.
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Where are essential oils stored in the plant?
Oil is stored in specialised secretory structures: glandular trichomes (Lamiaceae such as mint, basil,
lavender), oil cells (Piperaceae, Lauraceae), secretory cavities or oil glands in rind (Rutaceae, i.e.,
citrus), and resin ducts (Pinaceae, Apiaceae). The location and depth of these structures directly affect
which extraction method works best. Surface trichomes rupture readily, whereas oil embedded in seeds
or woody tissue requires cell-wall disruption.
Why look for new extraction methods?
Conventional hydrodistillation and steam distillation have been used for centuries, but they have
known drawbacks: long processing times (3 to 8 hours), high energy consumption, thermal degradation
of labile compounds, hydrolysis of esters, loss of polar constituents in wastewater, and poor recovery
of high-boiling components. Solvent extraction leaves residues and may be toxic. Growing consumer
demand for natural products, tighter regulation, sustainability goals (Green Chemistry, Green
Extraction principles) and the need for higher-quality oil for pharmaceuticals, food, cosmetics and
aromatherapy have driven the development of newer techniques.
Conventional Methods in Brief
Hydrodistillation (HD)
Plant material is fully immersed in
boiling water. Oil vapours and
steam co-distil at a temperature
below the boiling point of either
component alone (Dalton’s law of
partial pressures), are condensed,
and the oil separates from the
hydrosol in a Florentine flask.
Simple, but slow and with a risk of
hydrolysis and overheating.
Steam distillation
Live steam is passed through a bed of
plant material supported above the water
level. It is the most widely used
industrial method because it avoids
direct contact of the plant with boiling
water, giving better quality than HD.
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Cold pressing
Used for citrus oils (orange, lemon,
bergamot, mandarin, lime). Rind oil
glands are ruptured mechanically and the
oil is recovered as an oil–water emulsion
followed by centrifugation. No heat is
applied, so fresh top notes are preserved,
but co-extracted waxes and pigments
need to be removed.
Solvent extraction and enfleurage
Hexane, petroleum ether or ethanol
extracts flowers to give a waxy
“concrete”, which is then washed with
ethanol to yield an “absolute”.
Enfleurage uses purified fat to absorb
fragrance from flower petals. These
methods capture delicate aromas but
leave solvent traces and are labourintensive.
Latest Methods of Essential Oil Extraction
Microwave-Assisted Extraction
Principle
Microwaves (usually 2.45 GHz)
are non-ionising electromagnetic
waves that heat materials by two
mechanisms: dipole rotation of
polar molecules (mainly water in
the plant tissue) and ionic
conduction of dissolved ions.
Because heating is volumetric and
internal, the water inside the
glandular structures is heated
rapidly, builds up internal pressure and ruptures the cells, releasing oil to the surface where it is carried
away by steam or solvent. Heating is therefore from inside to outside, in contrast to conduction heating
in conventional distillation (outside to inside).
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Process parameters
Microwave power (typically 300–1000 W for lab systems): higher power speeds extraction but
may degrade oil.
Irradiation time: yield rises then plateaus; over-irradiation causes charring and losses.
Moisture content of material: water content is essential for SFME because water absorbs
microwaves; very dry material must be rehydrated.
Particle size and loading: influences penetration depth of microwaves and heat uniformity.
Solvent dielectric properties (in solvent-based MAE): polar solvents absorb strongly, non-polar
(hexane) are microwave-transparent.
Advantages and limitations
Advantages: very short extraction time, reduced energy use (up to 70–90% less than HD in many
reports), reduced or zero solvent, higher content of oxygenated compounds, less wastewater, and good
reproducibility.
Limitations: capital cost, penetration depth limits scale-up, risk of hot spots, unsuitable for materials
with very low moisture unless rewetted, and need for safety controls (leakage, pressure). Industrialscale continuous microwave units are now available and remain an active area of development.
Examples: SFME has been successfully applied to lavender, rosemary, thyme, peppermint, orange
peel, basil and cardamom, often giving oil of comparable or better quality than HD in a fraction of the
time.
Ultrasound-Assisted Extraction (UAE)
Principle
Ultrasound consists of sound
waves above 20 kHz. When
passed through a liquid, it
produces
alternating
compression and rarefaction
cycles. During rarefaction,
microbubbles form and grow;
during compression they
collapse
violently,
a
phenomenon called acoustic
cavitation. Collapse near the plant surface generates micro-jets, shock waves, localised high
temperatures and pressures, causing cell wall disruption, pore formation and enhanced mass transfer
of oil into the medium (solvent, water or a hydrotrope). Additional mechanisms include fragmentation,
erosion, sonocapillary effect and sonoporation.
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Equipment
Ultrasonic baths (indirect sonication, 20–40 kHz, low intensity) and probe (horn) systems (direct
sonication, higher intensity, better reproducibility). Industrial flow-through sonoreactors are available.
UAE is used either as a stand-alone extraction or as a pre-treatment before hydrodistillation
(ultrasound-assisted hydrodistillation, UAHD) or in combination with microwaves (ultrasonic–
microwave-assisted extraction, UMAE).
Parameters
Frequency and intensity (W/cm²): lower frequencies (20–40 kHz) produce stronger cavitation.
Temperature: raises vapour pressure and lowers cavitation intensity; an optimum is required.
Solvent: viscosity, surface tension and vapour pressure influence bubble behaviour.
Time, liquid-to-solid ratio and particle size.
Advantages: simple, cheap, low temperature, short time, applicable to a wide variety of materials, low
solvent consumption.
Limitations: free radical generation at high intensity may degrade some compounds; probe erosion;
scale-up requires careful reactor design; non-uniform energy distribution in baths.
Supercritical Fluid Extraction (SFE), Mainly Supercritical CO2
Principle
A substance above its critical
temperature and critical pressure is
a supercritical fluid with gas-like
diffusivity and viscosity and
liquid-like density and solvating
power. For carbon dioxide, the
critical point is 31.1 °C and 73.8
bar (7.38 MPa). Small changes in
pressure and temperature markedly
change the density and hence the
solvent power of the fluid, so
extraction can be tuned to target specific compound classes. When pressure is released after extraction,
CO2 returns to gas and leaves the extract solvent-free.
Process description
1.
Liquid CO2 from a cylinder is cooled, pressurised by a high-pressure pump and heated to
supercritical conditions.
2.
Supercritical CO2 flows through the extraction vessel packed with ground plant material and
dissolves the volatile oil.
3.
The CO2 –oil mixture is depressurised through a back-pressure valve in one or more separators,
where the oil precipitates and CO2 is gasified.
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4.
CO2 is recompressed and recycled. A co-solvent (entrainer) such as ethanol (1–10%) may be
added to raise the polarity.
Fractionation
Multi-stage separators operated at
different pressures and temperatures
allow fractionation of the extract, for
example, separating waxes and
cuticular lipids (precipitated first)
from the true volatile fraction, giving
a cleaner oil than would be obtained
in a single-stage separation.
Advantages: no toxic solvent
residues, moderate temperature, no oxygen exposure, tunable selectivity, oil profile closer to the
natural aroma of the plant, CO2 is non-toxic, non-flammable, cheap and recyclable.
Limitations: high capital and operating cost, need for high-pressure equipment and trained operators,
batch operation (semi-continuous), co-extraction of waxes in some cases. It is widely used
commercially for spices, hops, herbs and high-value oils (e.g., ginger, black pepper, chamomile, rose,
vanilla).
Subcritical Water Extraction (SWE) / Pressurised Hot Water Extraction
Water heated between 100 °C and
374 °C (its critical temperature)
under enough pressure (typically
10–60 bar) to remain liquid is called
subcritical water. As temperature
rises, the dielectric constant of water
falls sharply from about 80 at 25 °C
to about 27 at 250 °C, similar to
ethanol or methanol. Hence, hot
pressurised water can dissolve
moderately polar and even nonpolar organic compounds without organic solvents. Viscosity and surface tension also fall, favouring
penetration into the matrix.
For essential oils, SWE is generally run at 100–200 °C, in static or dynamic mode, with extract
collected after cooling and separated from water (liquid–liquid partition or adsorption). It is
particularly effective for recovering oxygenated compounds, which are more valuable than
hydrocarbon terpenes, and gives cleaner oils with fewer terpene hydrocarbons than conventional HD.
It also suits extraction of polar aroma compounds from spices such as cumin, fennel, coriander,
caraway and clove.
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Advantages: water is the only solvent (cheap, safe, green), short time, selectivity by temperature.
Limitations: thermal degradation above ~200 °C, need for pressure vessels and corrosion-resistant
materials, and the need to separate dilute oil from large volumes of water.
Pressurised Liquid Extraction (PLE) / Accelerated Solvent Extraction (ASE)
A solvent (ethanol, ethyl acetate, water
or mixtures) is kept liquid above its
normal boiling point by applying
pressure (typically 50–200 bar) at 40–
200 °C. Elevated temperature increases
solubility and diffusion, while pressure
forces solvent into pores. Extraction
takes 5–20 minutes in automated cells
with much lower solvent use than
Soxhlet. PLE is more common for nonvolatile extracts but is applied to
aromatic materials to obtain aroma-rich
extracts, particularly where a wider range of constituents than steam-volatiles is required.
Pulsed Electric Field (PEF) Assisted Extraction
PEF applies very short (microsecond
to millisecond) high-voltage pulses
(typically 0.5–10 kV/cm for plant
tissue, up to 20+ kV/cm for tough
material) to plant material placed
between two electrodes in a treatment
chamber. The electric field induces
electroporation: the formation of
pores in cell membranes when the
transmembrane potential exceeds a
critical value (about 0.5–1 V). Pores
increase permeability and mass
transfer, allowing oil and other metabolites to escape more easily.
PEF is a non-thermal pre-treatment, applied before hydrodistillation, pressing or solvent extraction. It
shortens the subsequent extraction time, reduces energy, and raises yield and quality. It has been
reported to enhance recovery in orange peel, lavender, rosemary, thyme, and other materials.
Advantages: minimal heating (temperature rise below 10 °C), low energy per kg, continuous operation
possible, easy scale-up in liquid streams.
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Limitations: equipment cost, need for a conductive medium (water), less effective for dry, nonconductive material, and treatment parameters (field strength, pulse number, specific energy) must be
optimised for each plant.
Enzyme-Assisted Extraction (EAE)
Plant cell walls are composed of
cellulose, hemicellulose, pectin and
lignin, which act as a barrier to oil
release, especially in seeds, roots and
woody tissues. Enzymes such as
cellulase, pectinase, hemicellulase,
xylanase, β-glucosidase and protease
selectively hydrolyse these polymers,
weakening the wall and freeing oil and
bound aroma precursors. Enzymes can
also release glycosidically bound
volatiles, increasing aroma yield.
Procedure: powdered or macerated material is incubated in buffer at optimum pH (usually 4–6) and
temperature (40–55 °C) with enzyme (0.5–5% w/w) for 1–24 h, followed by conventional distillation
or extraction. EAE has been applied to cardamom, ginger, turmeric, fennel, caraway, cumin, rose
petals, and citrus peel.
Advantages: mild conditions, specific action, environmentally friendly, higher yield and improved
quality.
Limitations: enzyme cost, sensitivity to pH, temperature and inhibitors, longer incubation time
compared with physical methods, and difficult scale-up. Combination approaches (enzyme +
ultrasound or enzyme + microwave) are common.
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