INTRODUCTION:
Microwave
heating was first introduced commercially in 1947. It took some time to catch
on but it is now a standard item in most kitchens for cooking uses. Its use in
the chemistry laboratory has significantly lagged behind domestic applications.
Microwaves were first used in 1975 as a heating source for acid digestion under
atmospheric conditions. The sample preparation step was reduced from 1-2 h to
15 min, producing an overall reduction in analysis time. This work initiated
the use of microwave energy as a heating source for the chemistry laboratory.
Other applications include distillation, organic and inorganic synthesis,
evaporation and solvent extraction. The early applications for MAE (microwave
assisted extraction) were for the extraction of compounds with nutritional
interest from plant and animal tissues. from 1986 to 1990 Ganzler1 published a series
of four papers exploring the use of microwave energy to partition various
compounds from soils, seeds, food and animal feeds as a sample preparation
method prior to chromatographic analyses. The extraction step was performed
under atmospheric conditions.
They
extracted a variety of compounds that included nutritive, crude fat and
pesticides. They used solvent schemes similar to their traditional Soxhletion
technique to allow a direct comparison of the recoveries between the two
techniques. The heating programme consisted of multiple 30s microwave heating
cycles (up to seven cycles) followed by a cooling step. This approach allowed
observation of the samples in the microwave cavity to prevent sample boil-over.
The microwave extracts gave recoveries that were 100-120% of the Soxhlet
technique. However, the MAE times were a factor of 100 less than the
traditional Soxhlet approach. The authors postulated that improved extraction
efficiency was due to the polar nature of the extracted compounds or the water
contained in the materials. They also noted a decrease in extraction efficiency
for the recovery of nonpolar compounds when a nonpolar solvent was used. The
published work of Freitag and John2 in 1990 expanded the application
and technique. These authors explored the use of microwave heating at elevated
temperatures and pressure for the extraction of additives from polyolefin. The
additives were antioxidants, Irganox 1010 and Irgafos 168, and the light
stabilizer Chimassorb 81. The polyolefin
matrices were polyethylene and polypropylene with a particle size of 20 mesh
and a sample size of 1 g. The MAE was performed with 30 mL of
1,1,1-trichloroethane or a mixture of acetone and n-heptane. They obtained
90-100% recoveries of the additives with extraction times of 3-6 min without
degradation of the analytes at the elevated temperatures of the extraction.
This compared favourably with the conventional 16h Soxhlet or 0.5- 2 h
reprecipitation techniques. Pare et al.3 from the Ministry of
Environment of Canada introduced the microwave-assisted process (MAP) through a
US patent in 1991. In this work, Pare used microwave energy to extract a
variety of essential components from natural products and foods, such as
biological and consumer products. The primary examples are the extraction of
essential oils from peppermint, garlic and cedar. The major feature of this
work is the release mechanism for the compounds of interest from the substrate.
The microwave energy was used to disrupt the glandular and vascular system of
the tissue without damaging the surrounding tissue. The solvent was used to
trap and dissolve the compounds released from the tissue.
In
general, the mechanism involves localized heating of the free water present in
the sample. Once the water is at or above its boiling point, the water causes
the cell membrane to rupture. This water, as steam, transports the target
analyte from the solid to the non absorbing solvent. In this type of work, the
sample is a good dielectric while the solvent is a poor dielectric. The
microwave process gave higher yields than the traditional steam distillation
process. The authors postulated that the improved efficiency was due to the
lower bulk temperatures and shorter extraction times. In 1992, (Bichi et.al2)
published work from the pharmaceutical area based on MAE using closed-vessel
technology with temperature-feedback control. They extracted pyrrolizidine
alkaloids from dried plants using 25-50 mL of methanol. The extractions were
performed over a temperature range of 65-1003C for 20-30 min. The MAE technique
gave qualitatively and quantitatively identical chromatographic results
relative to the samples extracted by the Soxhlet procedure with a significant
reduction in extraction time and solvent consumption. The temperature feedback
control provided highly reproducible extractions. In 1993, Onuska and Terry3
published the first data on the use of MAE for pollutants from environmental
samples. They successfully extracted organo chlorine pesticide residues from
soils and sediments using a 1:1 mixture of isooctane-acetonitrile using sealed
vials. The samples were irradiated for five 30 s intervals. This produced
faster and more reliable results than the conventional methods. They also
studied the use of a nonpolar solvent, iso-octane, for the extraction of wet
sediment samples. Pesticide residue recoveries increased as the moisture
content increased to a maximum of 15% and then levelled off. This shows the
importance of a polar co-solvent when using a nonpolar extracting solvent for
MAE techniques that are temperature-dependent. In 1994 (Lopez-Avila et al.4 ) Published their work to expand the
use of MAE to 187 volatile and semi volatile organic compounds from soils. The
compounds included polyaromatic compounds (PNA), phenols, organ chlorine
pesticides and organophosphorus pesticides. This work was performed using a
temperature feedback- controlled microwave heating system with closed vessels. This demonstrated the viability of
the technique for the extraction of many compounds of interest to the US
Environmental Protection Agency with relatively small volumes of solvent and
extraction times of only 10 min. This culminated in 1998 with the approval of
the microwave extraction technique by the SW-846 Organic Methods Workgroup of
incorporation into SW-846 as the latest of the ‘green’ methods.
In
1997 there were a number of significant extensions of the technology.
Incorvia-Mattina et al. investigated the use of MAE to extract taxanes,
used in ovarian and breast cancer research, from Taxus biomass. MAE offered
significant time savings versus the standard shaking technique and improved
yields versus synthetic approaches. Stout et al. Combined MAE with
liquid chromatography electrospray ionization mass spectrometry for the
determination of imidazolinones in soils at concentrations of less than 1
p.p.b. McNair et al.5 reported the combination of MAE with
solid-phase micro extraction for the analysis of Savour ingredients at concentrations
of 2-10 p.p.b. in solid food samples. This technique showed good selectivity
for the target analytes in a variety of foods.
Basic Principles
Mechanism of Microwave Extraction
The
fundamentals of the microwave extraction (MAE) process are different from those
of conventional methods (solid–liquid or simply extraction) because the
extraction occurs as the result of changes in the cell structure caused by
electromagnetic waves6.
In
MAE, the process acceleration and high extraction yield may be the result of a
synergistic combination of two transport phenomena: heat and mass gradients
working in the same direction (Fig.1).
On the other hand, in conventional extractions the mass transfer occurs from
inside to the outside, although the heat transfer occurs from the outside to
the inside of the substrate. In addition, although in conventional extraction
the heat is transferred from the heating medium to the interior of the sample,
in MAE the heat is dissipated volumetrically inside the irradiated medium7.
During
the extraction process, the rate of recovery of the extract is not a linear
function of time: the concentration of solute inside the solid varies, leading
to a non stationary or unsteady condition. A series of phenomenological steps
must occur during the period of interaction between the solid-containing
particle and the solvent effectuating the separation, including (1) penetration
of the solvent into the solid matrix8; (2) solubilisation and/or
breakdown of components; (3) transport of the solute out of the solid matrix;
(4) migration of the extracted solute from the external surface of the solid
into the bulk solution9; (5) movement of the extract with respect to
the solid; and (6) separation and discharge of the extract and solid
Figure.1
Basic heat and mass
transfer mechanisms in microwave and conventional extraction of natural
products.
(Adapted from Périno-Issartier et al6)
Therefore,
the solvent penetrates into the solid matrix by diffusion (effective), and the
solute is dissolved until reaching a concentration limited by the
characteristics of the solid. The solution containing the solute diffuses to
the surface by effective diffusion. Finally, by natural or forced convection,
the solution is transferred from the surface to the bulk solution .
The
extraction process takes place in three different steps: an equilibrium phase
where the phenomena of solubilisation and partition intervene, in which the
substrate is removed from the outer surface of the particle at an approximately
constant velocity (Fig 2). Then,
this stage is followed by an intermediary transition phase to diffusion. The
resistance to mass transfer begins to appear in the solid–liquid interface10;
in this period the mass transfer by convection and diffusion prevails. In the
last phase, the solute must overcome the interactions that bind it to the
matrix and diffuse into the extracting solvent. The extraction rate in this
period is low, characterized by the removal of the extract through the
diffusion mechanism. This point is an irreversible step of the extraction
process; it is often regarded as the limiting step of the process . Many
forces, such as the physicochemical interactions and relationships, can be
exposed during the extraction (dispersion forces, interstitial diffusion,
driving forces, and chemical interactions), and the persistence and strength of
these phenomena may be closely tied to the properties of the solvent
(solubilisation power, solubility in water, purity, polarity, etc.)11.
Fundamentals of
Microwave Extraction
Figure. 2 Schematic
representation of yield versus time in extraction processes. (Adapted from Raynie)
Mechanism of Microwave Heating
In
the microwave heating process, energy transfer occurs by two mechanisms:
Dipole
rotation and ionic conduction through reversals of dipoles and displacement of
charged ions present in the solute and the solvent .In many applications these
two mechanisms occur simultaneously. Ionic conduction is the electrophoretic
migration of ions when an electromagnetic field is applied, and the resistance
of the solution to this flow of ions results in friction that heats the
solution. Dipole rotation means rearrangement of dipoles with the applied field12.
Energy
transfer is the main characteristic of microwave heating. Traditionally, in
heat transfer of the conventional process, the energy is transferred to the
material by convection, conduction, and radiation phenomena through the
external material surface in the presence of thermal gradients. In contrast, in
MAE, the microwave energy is delivered directly to materials through molecular
interactions with the electromagnetic field via conversions of electromagnetic
energy into thermal energy The most important properties involved in microwave
processing of a dielectricare the complex relative permittivity ( ε ) and
the loss tangent (tan d)13.
Î=
=
The
material complex permittivity is related to the ability of the material to
interact with electromagnetic energy, whereas ε ′ is the real part,
or dielectric constant, and ε ′′ is the imaginary part,
or loss factor. The dielectric constant determines how much of the
incident energy is reflected at the air–sample interface and how much enters
the sample (for vacuum, ε ′ = 1); the loss factor measures the
efficiency of the absorbed microwave energy to be converted into heat. The loss
tangent (tan d or dielectric loss) is the most important property in microwave
processing; it measures the ability of the matrix to absorb microwave energy
and dissipate heat to surrounding molecules, being responsible for the
efficiency of microwave heating. As a result, a material with high loss factor
and tan d combined with a moderate value of ε ′ allows converting
microwave energy into thermal energy14.
The
first factor one must consider when selecting microwave physical constants is
the solvent to be used. It is important to select a solvent with high
extracting power and strong interaction with the matrix and the analyte. Polar molecules
and ionic solutions (typically acids) strongly absorb microwave energy because
of the permanent dipole moment. On the other hand, when exposed to microwaves,
non-polar solvents such as hexane will not heat up. The degree of microwave
absorption usually increases with the dielectric constant. A simple comparison
between water and methanol shows that methanol has a lesser ability to obstruct
the microwaves as they pass through but has a greater ability to dissipate the
microwave energy into heat. The higher dielectric constant of water implies a
significantly lower dissipation factor, which means that the system absorbs
more microwave energy than it can dissipate. This phenomenon is called superheating:
it occurs in the presence of water in the matrix. This strong absorption
provides an increase of the temperature inside the sample, leading to the
rupture of cells by the in situ water. In some cases it can promote the
degradation of the target compound or an “explosion” of solvent, and in other
cases it can increase the diffusivity of the target compound in the matrix.
Therefore, the microwave power must be sufficient to reach the boiling point of
the water or other solvent, setting the separation temperature. The second
factor to be considered is the solid matrix. Its viscosity affects its ability
to absorb microwave energy because it affects molecular rotation. When the
molecules are “locked in position” as viscous molecules, molecular mobility is
reduced, thus making it difficult for the molecules to align with the microwave
field. Therefore, the heat produced by dipole rotation decreases, and
considering the higher dissipation factor (d), the higher is this factor, the
faster the heat will be transferred to the solvent15.
Instrumentation
MAE
is the process of heating solid sample-solvent mixtures with microwave energy
and the subsequent partitioning of the compounds of interest from the sample to
the solvent. The most common approach is to perform the extraction in a sealed
vessel that is microwave-transparent. This allows a temperature elevation
significantly above the atmospheric boiling point of the solvent (Table 1) and hastens the
extraction process16.
Microwave Instrumentation
Figure 4 shows the major components of the microwave system, including the
magnetron, isolator, wave guide, cavity and mode stirrer. Microwave energy is
generated by the magnetron, propagated down the wave guide and introduced into
the cavity. The mode stirrer distributes the energy in various directions while
the cavity acts as containment housing for the energy until it is absorbed by
the sample load within the cavity. The isolator protects the magnetron from
reflected energy that would decrease its power output. A good analogy is a
one-way mirror it allows energy to go from the magnetron to the cavity but will
not allow it to go from the cavity to the magnetron. A turntable can be used to
rotate the sample load within the cavity to ensure even energy distribution.
Microwave heating is significantly different from conductive heating methods18
Figure 4 Microwave system
components.
Conductive
heating is sample-independent. All samples placed inside a conduction heating
oven will equilibrate to the programmed temperature. This can take quite some
time. Microwave heating is sample-dependent. The temperature rise rate of
samples will depend on their microwave-absorbing characteristics. The microwave
design provides the ability to heat uniformly a large number of samples in a
short period of time based on the sample load characteristics.
Solvent Safety Features
Due
to the flammable characteristics of many organic solvents, there is a major
safety issue when heating a solvent in a microwave field. This safety issue is
magnified when heating these solvents in sealed vessels at temperatures up to
100C above their atmospheric boiling point. The microwave system should have
redundant safety features, each acting as a backup to prevent possible fire or
explosion from occurring inside the cavity. Instruments should be designed to
eliminate possible ignition sources, to detect solvent leaks and to remove
leaking solvent. Figure 5 is an illustration of the interior of a
commercially available microwave extraction system. The safety aspects are an
exhaust fan which evacuates the cavity air volume approximately once per
second. If the exhaust fan fails or there is a block downstream of the fan, the
air Sow switch shuts down the system. The solvent detector monitors the cavity
for the presence of solvent. The detector shuts the system down if solvent
concentrations reach one-tenth of the lower explosive limit for acetone, sets
off an alarm and posts a message for the operator. The cavity is Teflon coated
to minimize the potential of high energy discharges. The system door is
designed to withstand an event equivalent to the explosion of 1 g of TNT. It
will partially open, allowing gases to escape, and then the compression springs
will pull it closed. This will contain any of the contents associated with a
vessel related event inside the system’s cavity19.
. 
Figure 5 Safety exhaust
and solvent detector
Vessel Technology
The
closed vessels used for MAE are designed for temperatures up to 2003C and
pressures of 200 psi (14 bar). The materials of construction for the components
that are in contact with the sample solvent mixture, either Teflon or glass,
are inert to solvents. However, since these materials are relatively weak, an
outer body is used that is much stronger either a reinforced thermoplastic or a
frame of polypropylene, or both. In addition, these materials of construction
absorb a minimal amount of microwave energy. The vessels are composed of glass
or Teflon liners, Teflon, PFA_ seal cover, polyetherimide load disc and sealing
screw, glass- filled polyetherimide sleeve and polypropylene support frame. The
vessel has a built-in pressure relief mechanism for safety purposes (Fig 6a and b). If the pressure inside
the vessel exceeds the operating limits, the vessel will automatically vent.
The control vessel is modified to accept a probe to monitor and control the
extraction temperature. A turntable of 14 extraction vessels is placed into the
instrument’s cavity for batch processing. The ability to rapidly achieve
elevated solvent temperatures under controlled conditions for a large batch of
samples is a major advantage of the MAE technique20.
Figure 6 illustrates a
standard extraction vessel: A)
components of vessel, B) Assembly of vessel
Temperature Control System
Temperature
control is necessary to optimize the extraction efficiency, prevent thermal
degradation of the target analytes, and to provide reproducible operating
conditions. This is achieved with a temperature measurement system that is
microwave-transparent so it does not cause any self-heating. The temperature
probe is inserted directly into the control vessel to measure the temperature
of the solvent -sample mixture. It is then used in a feedback control loop to
regulate the microwave power output to achieve and maintain the
operator-selected extraction temperature. This approach provides temperature
control for one of the samples in the batch and assumes equivalent reaction
conditions for all the other samples. This control technique is augmented with
an indirect infrared temperature measurement system. The infrared sensor is
located underneath the cavity floor21. It monitors the temperature
of each vessel as it passes over the sensor. This temperature reading is
correlated to the direct temperature reading from the control vessel to provide
temperature data for all of the samples in the batch.
Indirect Heating Source
In
some extraction applications, the solvent of choice for the target analytes is
non-polar and therefore does not heat when exposed to the microwave field. This
would normally preclude the use of the MAE technique unless the analyst is
willing to alter the solvent scheme to include a polar co-solvent. This is not
desirable since it alters the extraction efficiency (or selectivity). This
problem is overcome with the use of an insert that is a chemically inert
Suoropolymer filled with carbon black, a strong microwave absorber. The insert
is placed into the vessel with the solvent-sample mixture. The insert absorbs
microwave energy and transfers the thermal energy it generates to the mixture.
The performance characteristics of the
heating inserts for heating with n-hexane in a microwave field.
The use of the heating insert allows transfer of existing methods to the
MAE technique without a change in the solvent scheme22.
Stirring Mechanism
Stirring
increases the surface area contact between the sample and solvent. This offers
the benefit of improved extraction efficiency and decreased solvent
consumption. Stirring is achieved with the use of a rotating magnet below the
cavity floor and the placement of a magnetic spin bar in the extraction vessel.
The magnet creates a rotating magnetic field that couples with the spin bar in
the vessel to create a stirring effect. The spin bars are either coated with an
unfilled fluoropolymer for applications using polar solvents or a carbon black
filled fluoropolymer for applications with nonpolar solvents.
These
components create a system to perform MAE. This technique has the capability to
reduce extraction time, reduce solvent consumption and improve extraction
efficiency. However, it does not come without a cost. The extraction step is
but one of many necessary steps to obtain the final analytical result. The
analyst must take into account the differences in using the MAE technique
versus their existing approach. The primary difference is the finished sample
form. It is the same sample -solvent mixture originally placed into the vessel.
The analyst needs to separate the sample from the solvent at the completion of
the extraction step. If the analyst can work with an aliquot of the solvent for
analysis, this objection can be overcome with the use of a syringe filtration
technique. A secondary consideration is the vessel manipulations used with MAE.
These manipulations will be new for the analyst23.
Important Parameters in Microwave-Assisted
Extraction and Mechanism of Action
The
optimization of MAE conditions has been studied in several applications. The
efficiency of the process is directly related to the operation conditions
selected. Special attention should be given to usually studied parameters that
may influence the performance of MAE such as solvent composition,
solvent-to-feed ratio, extraction temperature and time, microwave power, and
the characteristics of the matrix its water content. Comprehension of the
effects and interactions of these factors on the MAE process is significant.
Thus, this topic emphasizes some of the parameters that affect MAE, presenting
guidelines regarding the selection of proper operation conditions, and also
discusses the interaction between these parameters24.
Effect of Solvent System and
Solvent-to-Feed Ratio (S/F)
The
most important factor that affects MAE process is solvent selection. A proper
solvent choice will provide a more efficient extraction process. Solvent
selection depends on the solubility of the compounds of interest, solvent
penetration and its interaction with the sample matrix and its dielectric
constant, and the mass transfer kinetics of the process. The solvent should
preferably have a high selectivity toward the solutes of interest excluding
undesired matrix components. Another important aspect is that the optimal
extraction solvents cannot be selected directly from those used in conventional
extractions: it depends on the capacity of the solvent to absorb the microwave
energy and consequently heat up. In general, the capacity of the solvent to
absorb microwave energy is high when the solvent presents high dielectric
constant and dielectric loss. Solvents that are transparent to microwaves do
not heat when submitted to them. Hexane is an example of microwave-transparent
solvent whereas ethanol is an excellent microwave- absorbing solvent. Both
polar and non-polar solvents can be used in MAE, and solvents such as ethanol,
methanol, and water are sufficiently polarized to be heated by microwave energy25.
In this context, the properties of the solvent can be modified when combining
different solvents, which allow varying the solvent selectivity for different
compound. The addition of salts to the mixture can also increase the heating
rate, because besides dipole orientation the ion conductivity is the main
origin of polarization and corresponds to losses to heat in dielectric heating
. Studies have shown that small amounts of water in the extracting solvent make
possible the diffusion of water into the cells of the matrix, leading to better
heating and thus facilitating the transport of compounds into the solvent at
higher mass transfer rates. In the case of volatile compounds, the addition of
a solvent with relatively low dielectric properties can be used to ensure that
the solvent temperature is kept lower to cool off the solutes once they are
liberated into the solvent. Generally, hexane is used for the extraction of
volatile oils. In addition, the solvent-free MAE(SFMAE) process has been
designed for aromatic herbs rich in volatile oils; in this case, the moisture
content within the plant matrix itself serves for extraction and no solvent is
used . Studies have reported that ethanol or water can be added into poor
microwave absorbers, such as hexane, to improve the extraction efficiency. One
of the most used solvent mixtures is hexane-acetone, and only a small amount of
water (about 10%) must be added in non-polar solvents such as hexane, xylene,
or toluene to improve the heating rate . Zhou and Lui10 evaluated
different mixtures of Ethanol and hexane in the extraction of solanesol from
tobacco leaves; the 1:3 ratio gave the best yield. Comparing isopropanol and
hexane for rice bran oil extraction, hexane at 40°C extracted approximately 40%
more oil than isopropanol. Although by increasing the temperature hexane did
not extract significantly more amount of oil, isopropanol extracted about 25%
more rice bran oil at 120°C .Some authors studied the use of combined solvents
in MAE according to the polarity of the target compounds. A methanol–water
(85:15) combination proved to be a good solvent for MAE of gymnemagenin from Gymnema
sylvestre R. Br. Higher water concentration reduced the extraction yield
because high water content increases the mixture polarity to a degree where it
is no longer is favourable for extraction. The same was observed by Taleba et
al.20 when extracting paclitaxel from Taxus baccata : a methanol–water
(90:10) mixture was the best combination. Song et al., extracting sweet potato
leaves, found that 60–80% (v/v) ethanol concentration in water was optimal
within proportions of 40% and 80% (v/v).
The
solvent-to-solid (feed) ratio (S/F) is an important parameter to be optimized.
The solvent volume must be sufficient to guarantee that the entire sample is
immersed in the solvent throughout the entire irradiation process, especially
when using a matrix that will swell during the extraction26.
In
conventional extractions, the use of large volumes of solvent increases the
extraction recovery. Studies reported that the extraction solution must not
exceed 30–34% (w/v). In many applications, a ratio 10:1 (ml/mg) to 20:1 (ml/mg)
was found to be optimal. In addition, the solvent volume is an important factor
to be considered because too much of the extracting solvent means more energy
and time is required to condense the extraction solution in the later step and
purification process. On the other hand, MAE may give lower recoveries because
of non-uniform distribution and exposure to microwaves. In some cases, small
amounts of solvent are sufficient to extract the compounds of interest. The
phenol and methyl phenol extracted from oils had optimal conditions when S/F
reached. A different behaviour was observed in the MAE of artemisinin from Artemisia
annua L.: a higher extraction rate was achieved by a greater amount of
solvent . In Ganoderma atrum , the yield of triterpenoid saponins
increased with the increase of amount of solvent until the S/F reached 25, and
then it decreased rapidly27.
Effect of Extraction Time and Cycle
In
MAE the period of heating is another important factor to be considered.
Extraction times in MAE are very short compared to conventional techniques and
usually vary from a few minutes to a half-hour, avoiding possible thermal
degradation and oxidation, which is especially important for target compounds
sensitive to overheating of the solute–solvent system. Overheating occurs
because of the high dielectric properties of the solvent, especially ethanol
and methanol, and further dilution with water that increases the heat capacity
of the solvent combination. Higher extraction time usually tends to increase
the extraction yield. However, this increase was found to be very small with
longer time . Irradiation time is also influenced by the dielectric properties
of the solvent. Solvents such as water, ethanol, and methanol may heat up
tremendously on longer exposure, thus risking the future of thermo labile
constituents28.
Occasionally,
when longer extraction time is required, the samples are extracted in multiple
steps using consecutive extraction cycles, which are also an example of the use
of a larger amount of solvent and higher microwave application time . In this
case, the fresh solvent is fed to the residue and the process is repeated to
guarantee the exhaustion of the matrix. With this procedure, the extraction
yield is enhanced, avoiding long heating. The number of process cycles will depend
on the type of matrix and the solute. According to Li et al.15,
Three cycles of 7 min were appropriate for MAE of triterpene saponins from
yellow horn, whereas in optimization of triterpenoid saponins MAE from Ganoderma
atrum, cycles of 5 min each were recommended. Yan et al.16 found
that three extraction cycles of 5 min each are optimal for extracting astragal
sides from Radix astragal. They also found that increasing the
irradiation time from 1 to 5 min increases the extraction yield rapidly;
extraction reaches its maximum at 5 min, and then the yields decreased with the
extension of the irradiation time. In the case of flavonoids extraction from R.
astragali, there was an increase in yield with time up to an exposure of 25
min and then the extraction yield started to decrease29. In the work
of Chen et al29. it was observed that triterpenoid saponins yield
from Ganoderma atrum reached its maximum at 20 min; after this time, the
target compounds easily decomposed because of long exposure to high temperature.
The same behaviour was found by Song et al. 14.
Effect of Microwave Power and
Extraction Temperature
Microwave
power and temperature are interrelated because high microwave power can bring
up the temperature of the system and result in the increase of the extraction
yield until it becomes insignificant or declines. It is known that the
temperature is controlled by incident microwave power that controls the amount
of energy provided to the matrix, which is converted to heat energy in the
dielectric material. At high temperatures the solvent power increases because
of a drop in viscosity and surface tension, facilitating the solvent to
solubilise solutes, and improving matrix wetting and penetration . In addition,
when MAE is performed in closed vessels, the temperature may reach far above
the boiling point of the solvent, leading to better extraction efficiency by
the desorption of solutes from actives sites in the matrix. However, Rout ray
and Orsat28 state that the
efficiency increases with the increase in temperature until an optimum
temperature is reached and then starts decreasing with the further increase in
temperature: this happens because the selection of ideal extraction temperature
is directly linked with the stability and, therefore, with the yield of the
target compound30.
Microwave
power is directly related to the quantity of sample and the extraction time
required. However, the power provides localized heating in the sample, which
acts as a driving force for MAE to destroy the plant matrix so that the solute
can diffuse out and dissolve in the solvent. Therefore, increasing the power
will generally improve the extraction yield and result in shorter extraction
time. On the other hand, high microwave power can cause poor extraction yield
because of the degradation of thermally sensitive compounds. Also, rapid
rupture of the cell wall takes place at a higher temperature when using higher
power, and as a result impurities can also be leached out into the solvent
together with the desired solute.
Therefore,
it is important to properly select the MAE power to minimize the time needed to
reach the set temperature and avoid a “bumping” phenomenon in temperature
during the extraction. Moreover, the overexposure to microwave radiation, even
at low temperature or low operating power, was found to decrease the extraction
yield because of the loss of chemical structure of the active compounds.
Knowing that power level alone does not gives sufficient information about the
microwave energy absorbed into the extraction system, Alfaro et al.17 created a term to study the effect of
microwave power on MAE: energy density, de fined as the microwave irradiation
energy per unit of solvent volume for a given unit of time (W/ml). According to
Li et.al15 , the energy density should be considered as a parameter
as power level alone. In this study, the anthocyanin extraction rates from
grape peel were different under the same microwave power level, extraction
time, and S/F because the energy density levels were different. Raner et al.18
reported that variation of power from 500 to 1,000W had no significant effect
on the yield of flavonoids. The decrease in extraction yield was found at
temperatures higher than 110°C because of instability of flavonoids and
consequent thermal degradation. In another case, higher microwave power led to
thermal degradation of phenols when it was higher than 350 W (between 150 and
550 W). The temperature behaviour was the same in other studies. In extracting
astragalosides from Radix astragali , Yan et al16. Also found
that yield increased remarkably with temperature increase from 50°C to 70°C;
above 70°C, the yields of astragalosides increased slowly and even decreased.
Effect of
Contact Surface Area and Water Content
Not
only the parameters already discussed but the characteristics of the sample
also affect the MAE process. It is known that in a higher contact surface area
the extraction efficiency increases. Also, finer particles allow improved or
much deeper penetration of the microwave. On the other hand, very fine
particles may pose some technical problems; consequently, centrifugation or
filtration is applied to prepare the matrix. In the preparation step the sample
is grinded and homogenized to increase the contact area between the matrix and
the solvent. The particle sizes are usually in the range of 100 mm to 2 mm. In
some cases soaking of the dried plant material in the extracting solvent before
MAE has resulted in improved yield. This procedure is called pre-leaching
extraction25.
In
many cases the extraction recovery is improved by the matrix moisture, which
acts as a solvent. The moisture in the matrix is heated, evaporated, and
generates internal pressure in the cell, which ruptures the cell to release the
solutes, hence improving the extraction yield. When increasing the polarity of
the solvent, water addition has a positive effect on the microwave-absorbing
ability and, hence, facilitates the heating process. Moreover, the additional
water promotes hydrolyzation, thus reducing the risk of oxidation of the
compounds. In extraction of astragalosides from Radix astragali,
extraction efficiency was improved by the addition of water27. The
possible reason for the increased efficiency is the increase in swelling of
plant material by water, which enhances the contact surface area between the
plant matrix and the solvent
Effect of Stirring
The
effect of stirring is directly related to the mass transfer process in the
solvent phase, which induces convection in the headspace. Therefore, equilibrium
between the aqueous and vapour phases can be achieved more rapidly. The use of
agitation in MAE accelerates the extraction by enhancing desorption and
dissolution of active compounds bound to the sample matrix. Through stirring,
the drawbacks of the use of low solvent-to-solid ratio (S/F) can be minimized,
together with the minimization of the mass transfer barrier created by the
concentrated solute in a localized region resulting from insufficient solvent.
In the work by Kovács et.al18. It is possible to observe the
difference between suspensions with and without stirring. The authors found
that when the suspensions were agitated with magnetic stirrers the temperature
reached its maximum value within a shorter time, and the temperature differences
inside individual vessels were not significant30.
Applications
MAE
has been applied to a wide variety of samples in Which traditional Soxhlet
extractions are performed.
The application of the these extraction for better extraction of plastics
and polymers, pesticides and environmental samples.
Plastics and Polymers
The
additive package used in the production of polyolefin is designed to improve
processing efficiency or to impart specific performance characteristics. The
package can contain antioxidants, antistatic agents, slip agents, anti-block
agents, UV stabilizers or antifogging agents. It is important for production
efficiency and product quality that the appropriate amount of each additive is
present. A fast and reliable method is needed to determine the additive
concentration level. The conventional extraction approach is a reflux technique
with an appropriate solvent solvent for 1-48h followed by high performance
liquid chromatography (HPLC) analysis. An alternative extraction technique is
sonication for 30-60 min, but the gain in speed is offset by a loss in
extraction efficiency. MAE has the ability to address the time and extraction
deficiencies of the reflux and sonication techniques. Freitag and John first
demonstrated the potential for MAE when they obtained excellent antioxidant
recoveries from polypropylene and polyethylene.
Pesticides and Herbicides
Pesticides
and herbicides are used to protect a wide variety of agricultural commodities.
There is an interest in pesticides, herbicides and their degradation product
concentrations in plant and animal tissues and soil and sediment samples. The
underlying assumption is that extractable compounds are labile in the
environment and constitute a threat to the environment if they are hazardous.
Specific examples are from the work of Fish and Revesz on chlorinated
pesticides from soils and the work of Stout’s group on imidazolinone herbicides
in plant tissues. Fish and Revesz19 showed chlorinated pesticide
recoveries from a Certified Reference Soil greater than or equal to those
achieved using the standard Environmental Protection Agency Soxhlet technique,
method 3540. This was achieved with extraction times of only 20 min and solvent
volumes of 50 ml. Stout et al.25 incorporated the use of MAE with liquid
chromatography electrospray ionization-mass spectrometry to shorten the
clean-up procedure and method development time of residue methodologies for
determining the imidazolinones and their metabolites in crops. This application
area is of concern not only to the traditional commercial testing laboratory,
but to agro-chemical producers.
Environmental
The
organic side of the environmental laboratory market constitutes the majority of
the analytical testing load. The extraction of priority pollutants, as well as
other organic molecular species, from solid samples is a primary concern. The
workload in the environmental laboratory is expected to increase significantly
and will thus require extraction techniques that offer increased throughput,
reduced solvent consumption, improved efficiency and reproducibility. MAE has
the potential to address these needs. McMillin of US Environmental Protection
Agency- Region VI demonstrated this in a comparison of various soil extraction
techniques for semi volatile analysis. He used an abbreviated MAE technique
consisting of a small medication to regular MAE. He worked with only 10 mL of
solvent versus the conventional 30 ml. This eliminated the subsequent
concentration step and allowed the sample to be injected straight from the
extraction vessel into a GC. The abbreviated MAE provided better extraction
efficiencies and reproducibility than the three conventional techniques. The
extraction time averaged 16 min per sample with a solvent use of 10 mL per sample.
One difficulty in the use of MAE for the environmental laboratory market is
Environmental Protection Agency approval. The methodology has been approved by
the SW-846 Organic Methods Workgroup for incorporation into SW-846. However,
the method has not been promulgated and thus can only be used when regulations
do not specifically require SW-846 methods.
Future Developments
The
instrumentation for MAE will continue to evolve, as will its potential
applications. There will be developments in the vessel technology to address
the separation issue of the sample and solvent after the extraction step. This
will allow the technique to be a true replacement for the Soxhlet. There is
also a need for larger vessel sizes. The current vessel has a working volume of
100 ml. There is a need to increase this to 250 mL and even higher for bulky
samples and the inevitable push for lower detection limits. Finally, the
microwave system’s use should be extended to concentration of the sample after
the extraction step. This will create a multi-tasking tool for the analytical
laboratory. MAE has focused on extraction applications from solid matrices.
However, its speed and efficiency suggest that this technique will be used for
isolating pharmaceutical compounds during the drug discovery process. The
recent addition of sample stirring suggests that it can be extended to
liquid-liquid extraction applications. It could also be coupled with
solid-phase micro extraction to lower detection limits significantly. As MAE
becomes more widely accepted and instrumentation evolves, we should see a
significant increase in its applicability.
CONCLUSION:
There
has been much research and many advances in development in the microwave
assisted extraction of a number of plant compounds. This chapter showed the
phenomena of mass and heat transfer of the MAE process as well the parameters
that influence MAE extraction of bioactive compounds. Therefore, optimized
operating parameters can improve MAE performance. Also, MAE is better or
comparable with other techniques. The main advantage of MAE resides in the
performance of the heating source. The high temperatures reached by microwave
heating reduces dramatically both the extraction time and the volume of solvent
required as a concluding remark, the MAE system is considered a promising
technique for plant extraction because of its use of different physical and
chemical phenomena compared to those in conventional extractions. Finally the
costs of the specialised equipment may also influence the choice of the
extraction technique (Camel, 2001).
ACKNOWLEDGMENT:
The
author wants to acknowledge the library of Rungta College of Pharmaceutical
Sciences and Research, Kohka-Kurud road Bhilai, for providing necessary
literature for the compilation of the work. The authors also want to thanks
Shri Santosh Rungta, Chairman, Santosh Rungta Group of Institution for
providing necessary facility and infrastructure for the completion of the work.
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