Supercritical Fluid and Subcritical Fluid

7/31/2019 Supercritical Fluid and Subcritical Fluid

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2010-2011

SU PERCRITICAL FLU ID AN DSU BCRITICAL FLU ID

Keyu r Va sava. .


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SUPERCRITI CAL FLUI D AN D SUBCRIT ICAL FLUI D

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Introduction

Pharma science is witnessing a lot of research targeted at meeting delivery and

manufacturing demands of the new age molecules. Entirely new technologies and modifications

of existing technologies are being developed to address these needs. Supercritical fluid

technology (SFT) has been used in many fields for decades, such as the food industry, chemical

processing, polymers, textile, forest product industries, and in the cleaning of precision parts

other than pharmaceuticals. In the pharmaceutical field it has been widely used for the extraction

of natural products like aromatic oils and caffeine, etc. Newer areas of their application have

appeared, such as particle size reduction, and designing of novel drug delivery systems.

Important Definitions:

1) Critical Temperature:If the temp. is elevated sufficiently, a value is reached above which it is impossible to

liquefy a gas irrespective of the pressure applied.This temp. above which a liquid can nolonger exist, is known as critical temperature.

2) Critical Pressure:The pressure required to liquefy a gas at its critical temperature is the critical pressure,

which is also the highest vapour pressure that the liquid can have.

Basis of SF Technology:

The first report which describing a potential process using a supercritical fluid as medium for

particles production was published by Hannay and Hogarth: We have then, the phenomenon of

a solid with no measurable gaseous pressure, dissolving in a gas. When the solid is precipitated

by suddenly reducing the pressure, it is crystalline, and may be brought down as snow in the gas,

or on the glass as a frost, but it is always easily redissolved by the gas on increasing the pressure

(Hannay and Hogarth, 1879).A fluid reaches the supercritical region when heated and pressurized above its critical

pressure and temperature; the critical point represents the end of the vaporization curve in the PT

phase diagram (Fig. 1). The supercritical status does not represent a specific aggregation state,

but it corresponds to a region where the physico-chemical properties of the material are

intermediate between those of the liquid and the gas. The macroscopic appearance of a fluid at


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the critical point is that of a homogeneous and opalescent system without apparent phase

separation because, at this point, the density of the gas and liquid are identical.

Like a gas the SCF shows lower viscosity and higher diffusivity relative to the liquid.

These properties facilitate mass transfer phenomena, such as matrix extraction or impregnation.

Like a liquid, the SCF shows a density value high enough for exerting solvation effects. A SCF

is dense but highly compressible, thus, any pressure change results in density alteration and,

consequently, in solvent power variation (Brunner, 1994). In the vicinity of the critical point, the

compressibility is high, and a small pressure change yields a great density modification.

FIG.-1 Phase Diagram of Super critical regio n.

Choice of SFs:

All gases can form SCF above specific sets of critical conditions (P, T), but it should be

kept in mind also that, for many cases, the transition to the supercritical state occurs at high

temperatures not compatible with pharmaceutical compounds (e.g., SC water, Table 1). The

critical P, T values increase with the molecular weight or intermolecular hydrogen bonding or

polarity [18]. Not only mild processing conditions, but also safety and affordable economics are

valid criteria to choose the supercritical fluid. For example, Xenon and SF6 (when sufficiently

purified) have low critical values, but remain too expensive for commercial use. Gases such as

N2O or ethane have low critical values, but can generate explosive mixtures and are therefore

unsafe to handle. SCF trifluoromethane (CHF3), which is chemically inert, nonflammable, has

low toxicity and a low critical temperature and pressure. Furthermore, CHF3 has a strong


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permanent dipole moment (1.56 D), and can thus solubilize pharmaceutical compounds.

However, more than 98% of applications have been developed using carbon dioxide as the SCF

because of its low critical temperature (31.18 C) and pressure (7.4 MPa), inexpensiveness, non-

flammability, non-toxicity, recyclability and environmental benignity.

The field of supercritical fluid is still young and the SCF technologies are not yet

widespread throughout the pharmaceutical industry, which is likely due to the high cost of the

equipment and the weak knowledge of this area.

MATERIAL Tc (.C) Pc(MPa)

H2O 374 22

Xenon 16.6 5.9

SF6 45.5 3.8

N2O 36.5 4.1

CHF3 25.9 4.7

CO2 31.3 7.4

C2H4 9.1 5.1


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FIG.-2 Phase Diagram of CO2

PROCESSING USING SUPERCRITICAL FLUID:

Supercritical fluids have been proposed for many different unit operations; nevertheless all

processes are based on some techniques which can be divided in four groups:

Operations where the SCF acts as a solvent (RESS, RESOLV);

Operations where the SCF acts as an antisolvent (GAS, PCA, ASES, SEDS);

Particles from a gas-saturated solutions (PGSS, DELOS, CPCSP);

CO2-assisted spray-drying (CAN-BD, SAA).

1. Operations where the SCF acts as a solvent:A) RESS process (Rapid Expansion of Supercritical Solution)

The RESS process (Rapid Expansion of Supercritical Solution) consists of the

saturation of the supercritical medium with a solute followed by a rapidly depressurization

(expansion) of the solution through a heated nozzle at supersonic speed. The rapid and

uniform pressure drop, obtained by passing from supercritical to ambient conditions, leads to

a dramatic and instantaneous decrease of solvent power (solute supersaturation) and as aconsequence, to a rapid nucleation of the solute in form of very small particles with uniform

size. These particles are completely dry, solvent free, and they do not need further

processing.

The morphology and size distribution of the precipitated material is a function of its

pre-expansion concentration and expansion conditions. The pre-expansion concentration is


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dependent on the choice of SF, nature of solute, addition of co-solvents and operating

pressure and temperature. The higher the pre-expansion concentration, the smaller the

particles and narrower the particle size range.

ADVANTAGES OF RESS:

The simple control of process parameters The relatively easy implementation on lab-scale when a single nozzle is used The absence of organic solvents

DISADVANTAGES OF RESS:

Limited the development of this technique: the difficulty in scaling-up The possible particles aggregation and nozzle blockage The need of large amount of SCF The poor solubility of most pharmaceutical compounds in supercritical CO2.

FIG.-3 RESS Equ ipm ent Concep t.

B) RESOLV processRESOLV (Rapid Expansion of a Supercritical Solution into a Liquid Solvent)

represents a variation of RESS. This technique was studied in order to minimize the

particles aggregation during the jet expansion. Here, the supercritical solution is

depressurized through anorifice into a collection chamber containing an aqueous solution at

room temperature. Various water-soluble polymers or surfactantsare added to the aqueous

medium for stabilizing the obtainednanoparticle suspension.


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2. Operations where the SCF acts as an antisolvent:These techniques were proposed to process molecules with poor solubility in

SCFs. Here the pressurized CO2 acts as an antisolvent for precipitating a solute from an

organic solvent solution.

The solute is first dissolved in an organic solvent, and then the pressurized

CO2 is put in contact with the solution. The principle is based on (i) the ability of the

organic solvent to dissolve a large amount of gas; (ii) the mutual miscibility of the organic

and SCF phases; and (iii) the low affinity of the SCF for the solute. The CO 2 diffuses into

the organic solvent leading to the solvent evaporation into the gas phase; then, the volume

expansion determines a density reduction, which lowers the solvent power of the organic

solvent which leads to the precipitation of the solute.

One of the requirements for this approach is that the carrier solvent and the SF

antisolvent must be at least partially miscible. This process works in a semi batch mode,

with the supercritical solvent introduced into an already existing stationary bulk liquid

phase. This mode offers better control over the particle characteristics as governed by the

rate of addition of the SF. However, the liquid phase cannot, in general, be completely

removed, and requires additional processing steps before a dry product can be recovered.

The effect of processing variables such as temperature, pressure, stirring rate,concentration of the injection solution, rate and temperature of the carrier solution, nature of

liquid solvents and choice of the SF on the physical properties of the end product have to be

optimized when looking to obtain any product of desired characteristics.

Different processes based on the different mixing modes between the organic

solution and the SCF were designed. In the Gaseous Antisolvent (GAS) the precipitation

vessel is loaded with the solution then, the SCF is introduced into a vessel until the final

pressure is reached.


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FIG.-4 GAS equipm ent concept.

In the Particles by Compressed Antisolvent (PCA) and Supercritical

Antisolvent(SAS), the CO2 (supercritical for SAS, or subcritical forPCA) is first pumped

inside the high-pressure vessel until the systemreaches the fixed pressure and temperature,

then, the organic solution is sprayed through a nozzle into the SCF bulk determining the

formation of the particles that are collected on a filter at the bottom of the vessel.

FIG.-5 SAS equi pm ent concept


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FIG.-6 ASES equ ipm ent concept

The Aerosol Solvent Extraction System (ASES) is similar to the SAStechnique except that, in this case, the solution and the antisolvent are simultaneously

sprayed into the precipitation vessel.

FIG.-6 SEDS equ ipm ent concept

Simultaneous spraying of the solution and the antisolvent occurs in the case of

Solution Enhanced Dispersion by SupercriticalFluids (SEDS) as well. Compared with the

other antisolvent techniques SEDS is characterized by the use of a co-axial nozzle designed


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with a mixing chamber, which allows the attainment of better mixing by enabling a

turbulent flow. SEDS was also successfully used to process aqueous solution of peptides

and proteins despite the limited solubility of water in SC-CO2. SEDS overcomes this

problem by using a co-axial three-component nozzle, in which the organic solvent, the SCF,

and the aqueous solution, as separate streams, meet in the mixing chamber before being

introduced in the precipitation vessel.

The temperature and pressure together with accurate metering of flow rates of

drug solution and SF through a nozzle provide uniform conditions for particle formation.

This helps to control the particle size of the product and, by choosing an appropriate liquid

solvent, it is possible to manipulate the particle morphology.

Although the antisolvent techniques are very promising for particles

engineering, the complete solvent removal still represents an issue: a final wash-out

treatment with supercritical CO2 through the precipitated particles may be required.

3. Particles from a gas-saturated solutionsParticles from gas-saturated solutions (PGSS), in its most interesting

application for pharmaceutics, implies the dissolution of the compressed gas (super- or

subcritical) into a melted material (usually a polymer) followed by the rapid depressurization

of the gas-saturated solution through a nozzle that causes the formation of particles.The interaction between CO2 and the polymer reduces inter chain polymer

bonds giving rise to an enhanced polymer segmental mobility that in turn results in a

reduction of the glass transition or melting point. The extent of the glass transition or melting

point depression depends on the amount of CO2 dissolved in the polymer. In the rubbery or

liquid state the polymer can be used as coating agent, to produce foams, or to incorporate

drugs, affording a molecular dispersion which can be extruded or sprayed at lower pressure

to obtain drug loaded microparticles. Obviously, polymer plasticization can be also induced

thermally or by means of organic solvents, however major drawbacks of these approaches are

the difficulty to process heat-sensitive materials and the possible presence of residual solvent

in the final product. These are points of major concern especially in pharmaceutical,

biotechnological and food applications.


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This technique offers numerous advantages: the pressure used is lower than

that necessary for RESS; the required amount of gas is relatively small; no organic solvents

are required; it can work in continuous mode; the process affords a good yield.

On the other hand, the poor control of the particles size and particle size

distribution represents an issue that has still to be addressed.

Nonetheless the balance between advantages and drawbacks should be

considered positive, and this technique might play a prominent role in the future of drug

delivery especially for the preparation of microparticulate systems.

Several PGSS plants are already running with capacities of some hundred

kilograms per hour (Natex, Austria, Thar Technologies, USA, Uhde HPT)

(http://www.natex.at, http://www.thartech.com, http://www.UhdeHPT.com).

Based on the principle of PGSS also DELOS and CPCSP processes have been

developed.

In the Depressurization of an Expanded Liquid Organic Solution (DELOS)

the CO2 expands in an autoclave where an organic solutionof the solute to be micronized is

dispersed. Then the ternarymixture solutesolventcompressed gas is depressurized by rapid

reduction of the system pressure to atmospheric conditions in an expansion chamber. During

the expansion the mixture cools down:the temperature drop is the driving force that causes

the nucleation and precipitation of the drug. In this process the CO2 does not act as anantisolvent, but as a co-solvent tonebulize and cool the organic solution. The process is not

necessarily supercritical; in fact the operative pressure does not exceed the critical point of

the CO2/solvent mixture. The Continuous PowderCoating Spraying Process (CPCSP) was

proposed by for coating powders. In the CPCSP, the main components (binder and hardener)

are melted in separated vessels to avoid a premature interaction with the polymer. The

molten polymer is fed into a static mixer, and homogenized with compressed carbon dioxide.

Then, the different components are intensively mixed and the formed solution expanded

through a nozzle into a spray tower.


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FIG.-7 PGSS equ ipm ent concept

4. CO2-assisted spray-dryingIn these aerosolization techniques the supercritical CO2 is used to assist the

nebulization of the solution of the compound to be processed. The mechanism of this process

is close to classical micronization by spray-drying. The substance is dissolved or suspended

in water or ethanol or both, and the solution or suspension is intimately mixed with the SC-

CO2. The formed emulsion is rapidly decompressed through a suitable device. In the case of

CAN-BD (Carbon dioxide Assisted Nebulization witha Bubble Dryer) the near-critical or

supercritical CO2 and the solution are pumped through a near zero volume to give rise to an

emulsion which expands through a flow restrictor into a drying chamber at atmospheric

pressure to generate aerosols of microbubbles and microdroplets that are dried by a flux of

warm nitrogen.

In the case of SAA (Supercritical Fluid-Assisted Atomization) the

supercritical CO2 and the solution are mixed into a vessel loaded with stainless steel

perforated saddle which assures a large contact surface between liquid solution and the SCF;

then the mixture is sprayed in a precipitator at atmospheric pressure under a flow of hot N 2.

The main difference between CAN-BD and SAA processes is represented by

the mixing part of the equipment and, therefore, by the extent of solubilization of the SC-CO2

in the liquid solution.


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In summary many different techniques were developed based on the

characteristics of the material to be treated and/or the final product. This underscores the

versatility of the SCF-based techniques: a process limitation in one area often opens up new

research and solutions in other directions. For instance, if the starting material is soluble in

the SCF the RESS will be preferred while in case of the low solubility an antisolvent process

will be used. When organic solvent has to be avoided, i.e. in the case of biological products,

processes such as CAN-BD or SEDS with three co-axial nozzle should be selected. Finally,

the characteristics of the desired product (micronized, coated, encapsulated, etc.) would drive

the selection of the technology as well.

APPLICATION OF SUPERCRITICAL FLUID IN PHARMACY FIELD:

1. DRUG DELIVERYA) Particle and Crystal Engineering (Size Reduction and Solid State Chemistry)B) Particle CoatingC) Particulate Dosage Form

Cyclodextrin Inclusion Complexes Extrusion

Liposomes Preparation Microspheres

2. Sterilization3. Solvent Removal4. Extraction5. Supercritical and Subcritical Chromatography

DRUG DELIVERY:

A) Particle and Crystal Engineering (Size Reduction and Solid State Chemistry)Standard Micronisation Processes: Comparison with Supercritical Fluid base Technique

The standard micronisation processes comprise crushing/ milling, air

micronisation, sublimation and recrystallization from solvents. These techniques can often

undergo several practical problems. The mechanical treatments can damage, degrade


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particles due to high stresses (thermal and mechanical) generated by attrition. Moreover,

changes of drug crystallinity could originate during stress, leading to chemical or physical

instability or to an increase of surface energy with increased tendency to adhesion and

agglomeration. In the conventional crystallisation process the products are more or less

contaminated with the solvents and waste streams are produced.

An alternative particle engineering technique is the spray drying process that

can partially alleviate some of the problems of the mechanical techniques, but it is difficult to

use with poorly water-soluble drugs and usually requires high operating temperatures, which

could cause degradation of heat sensitive materials.

Furthermore, all these techniques share the disadvantage of a poor control of

the particle size distribution. Some of the practical problems associated with the use of

conventional micronisation processes can be overcome by means of supercritical fluids

technologies, allowing the preparation of micro- or even nanoparticles with a narrow particle

size distribution.

The main established advantages of supercritical CO2 based techniques are:

Mild operating temperatures;

Single-step process; Recovery and recycle of fluid;

Green technology;

Solvent-free products.

Furthermore, traditional methods for the production of drug microparticles

require numerous manufacturing steps that can be avoided, in case of SCF techniques,

bringing to a simplification and better control of the process (Fig. 8).


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FIG.-9 Process simp lificatio n by SCF relat ive to conven t ional m icron-sized crystall ine po w der

product ion .

Crystallization is a transient non-equilibrium process associated with a phase

change leading to crystal formation. The driving force for this process is supersaturation.

A high degree of supersaturation is generated in non-equilibrium conditions, in the area

above the saturation boundary in the concentration/temperature diagram (spinodal curve,

Fig. 8). In this area, the crystallization process, referring to the simultaneous occurrence

of nucleation, crystal growth, and agglomeration, takes place.

Traditional crystallization methods include sublimation, crystallization from

solutions, evaporation, thermal treatment, desolvation, or grinding/milling. The

crystallization process is governed by both thermodynamic and kinetic factors, which can

make it highly variable and difficult to control. For a compound existing in various solid-

state forms, thermodynamic factors influence the conditions and direction in which atransformation from one polymorph to another takes place, while kinetics influences the

time in which this transition occurs. The relative thermodynamic stability of solids and

the driving force for a transformation at constant pressure is given by the Gibbs equation:


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FIG.-8 Spinodal curve: A) undersaturated area, decreasing temperature, the solut ion

goes to the satur at ion B), C) solut ion ent ers the m etastable zone , D) fast n ucleation, E)

concentrat io n decreases w ith crysta l grow th.

The relative stability is given by the algebraic sign of G; if it is negative the

phase transformation occurs naturally and the change continues as long as the free energy

of the system decreases. G=0 when the system is in equilibrium with respect to

transformation; G is positive when the free energy increases and the transformation

does not occur. The operative conditions that can be changed to control the process are

temperature range, cooling/heating rates, choice of solvents, and variation of solute

concentration (which depends on the temperature). Additional factors such as impurity

level, mixing regime, vessel design, and cooling profiles can deeply impact the size,

number, and shape of the crystals produced.

With respect to traditional processes, the use of SCFs implies the possibility

of more precise control of the operative conditions, but also of taking advantage of some

peculiar conditions, such as pressure, rate of solvent evaporation and fine-tuning of the

density. As discussed in some detail below, the kinetics of the crystallization process is

mainly driven by diffusion. Therefore, the density of the solvent greatly impacts this

aspect of the phenomenon, as this parameter strongly influences the diffusivity of the

solvent, or the antisolvent and, therefore, on the transient concentration gradient.

Additionally, the possibility of modulating the rate of solvent evaporation deeply

influences the kinetics of the process because of its relation to the rate of accumulating

supersaturation.


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RESS is an attractive process, simple and relatively easy to implement on a

small scale when a single nozzle is used. The main limitation of the use of RESS

technique is the poor solubility of pharmaceutical products in supercritical CO2. The low

solubility can, in some instances, be overcome with appropriate tuning of the fluid

density and the use of an adequate cosolvent. Other limitations are poor predictive control

of particle size, morphology along with the difficulty to scaling-up the process because of

particle aggregation and nozzle blockage caused by cooling effects due to the rapid

expansion of the supercritical solution.

Analogous RESS experiments were performed with supercritical CO2 and

ibuprofen to investigate the effects of extraction pressure, pre-expansion temperature,

capillary length, spraying distance, and collision angle on the size and morphology of the

precipitated particles. Micronisation of ibuprofen was successfully obtained with average

particle size of the powder below 8m all the different conditions tested. No clear

dependence of particle size on the pressure (1317MPa) was found, while a slight

decrease in the particle size was observed increasing the pre-expansion temperature. The

particle size decreased also when the capillary length (812mm) and the collision angle

(from 45 to 90) were increased, and the spraying distance (26 cm) was decreased.

RESS is usually performed on a binary system made up by the supercritical

fluid and the solid. Sometimes a cosolvent can be added to improve the solvent power ofthe SF. Polar organic solvents or capable of H-bonding (e.g. methanol) can enhance the

solubility of polar solutes in supercritical CO2. Enhancement factors (solubility in the SF-

cosolvent system divided by the solubility in the SF alone) values of 104106 are

common and even values as high as 1012 have been reported.

Besides cosolvents, also cosolutes can enhance the solubility of the solute in

supercritical CO2. The effects of addition of benzoic acid (BA) on the solubility in

supercritical CO2 of salicylic acid or phenanthrene were analyzed. The solubility of both

compounds was enhanced by the presence of BA while the crystal size decreased.


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TABLE-2 Volume diameter at 50% frequency (D[0.5]) and span for ASA powders

obtained by RESS at different temperature and pressure values (S.D.), n=3

Experimental Conditions:

T(

.

C), P(MPa)

D(0.5) (m) Span

40-15 5.39(0.06) 1.89(0.27)

40-20 4.75(0.06) 1.79(0.05)

40-25 4.02(0.53) 2.94(2.05)

50-15 4.70(0.16) 1.37(0.19)

50-20 4.60(0.40) 3.40(1.25)

50-25 4.04(0.93) 1.28(0.76)

Untreated Acetylsalicylic Acid D(0.5)=520m

Micronized particles of acetylsalicylic acid (ASA) were obtained with the

RESS process. The solid state characteristics of micronized ASA, as well as the

correlation between process parameters, namely temperature and pressure, and solid-state

properties were investigated. A remarkable reduction of particle size was obtained, which

linearly correlated to pressure (Table-2), while temperature apparently did not influence

the particle size of the products. No polymorphism was induced by the RESS process,

although a significant melting temperature decrease for micronized ASA was found and

linearly correlated with the reciprocal of the mean particle radius, by applying the Gibbs-

Duhem-Laplace equation.

GAS/SAS Process

Different SAS injection rates were used with sub- and supercritical carbon

dioxide to recrystallize sulfamethizole from solutions in acetone or N, N-

dimethylformamide (DMF). Sulfamethizole crystals with tabular and platy habits were

produced by different operating conditions; in particular, low CO2 injection rates

produced large crystals with irregular tabular habit, while higher injection rates gave rise

to small particles with thin platy habit. Also the solvent choice affected the crystal habit:

large platy crystals, independently of injection rate, were obtained from DMF; low

injection rates afforded tabular crystals, whereas higher injection rates resulted in thin


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platy crystals from acetone. Ultrasounds application to the nozzle was investigated: 22.5

kHz sonication accelerated nucleation, producing a larger number of small crystals.

Particle size increased with temperature increase, both in sub- and supercritical

conditions.

The effect of pressure on particle size depended on the solvent choice:

from ethanol and ethanol/methanol solutions, a significant decrease of particle size with

pressure increase was observed, while the opposite occurred from acetone solutions. The

different behavior was explained taking into account the difference in vapor pressure of

the solvents and the different phase equilibria of the ternary system. In case of lower

solution flow rates (0.3 L/h), more concentrated solutions afforded smaller particles,

whereas with high flow rates (0.6 L/h) the reverse tendency was found. The particle size

was more or less independent of the nozzle diameter when similar nozzles were used.

The nozzle design affected the particle size: for example, using a two-flow nozzle,

instead of a Laval nozzle, the mean particle size was reduced, probably because the two-

flow nozzle enhanced atomization energy and mixing efficiency.

ASES Process

Insulin, albumin, lysozyme, and recombinant human deoxyribonuclease

(rhDNase) were precipitated using the ASES process and CO2 modified with ethanol.

The proteins precipitated as nano-sized particles ranging from 100 to 500 nm, and theirphysical properties and biochemical stability depended on the operating conditions.

SEDS Process

The influence of SEDS process parameters mentioned above on the

recrystallization of the model drugs budesonide and flunisolide. As for budesonide, the

starting material was composed of irregularly spherical particle aggregates (typical

morphology of micronized powders with high energy operations). SEDS crystals formed

at 10MPa and 80 C from acetone were smooth, discrete and nearly spherical, while

crystals from methanol had a plate-like shape. The diameter of the particles from acetone

was within the 13m range, while those from methanol had a diameter within 530m.

The higher solubility in the SF of acetone compared with methanol influences the

saturation profile with a consequent effect both on nucleation and crystal growth,

producing particles with different size distributions and morphologies. The differences


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found for samples obtained from acetone or methanol could be partly due also to bond

formation between solvent and drug, thus affecting the extraction process. As

demonstrated by their uniformity, particle size and morphology can be effectively

controlled by the SEDS process.

These examples confirm that changes in morphology, size, and

surface properties of powders can be manipulated and designed by supercritical fluids

processes. In addition to morphology and micromeritics above discussed, particle

engineering of pharmaceuticals must take into account also another very important topic,

with both technological and biopharmaceutical implications: polymorphism,

pseudopolymorphism and solidsolid phase transitions.

The kinetics of the phase transitions can be influenced by typical

environmental parameters (temperature, pressure, relative humidity), presence of

crystalline defects, impurities and mechanical stress. Polymorphic purity is an important

parameter to consider in drug products, since the presence of differing crystal phases can

accelerate the conversion process by lowering the relevant activation energy barrier. The

phase transformation could lead to different polymorphs with unwanted physical and

chemical properties; for this reason, regulatory authorities have long recognized the need

for limiting polymorphic impurities in pharmaceutical materials.

Supercritical carbon dioxide in control of polymorphismAs explained in the previous section, different polymorphs have different

physical characteristics; thus, the identification of stable polymorphs with desired

physical properties is very important for drug products development. The solid phase

must be monitored, especially after technological unit processes potentially able to

modify the solid-state properties of the components. In particular, during the

conventional micronisation process such as milling, spray-drying and freeze drying, the

substances are usually exposed to mechanical stress, contact with solvents, heating

cooling cycles, that can often lead to alterations of the solid phase, such as new

polymorph formation, dehydration, vitrification via solid-state or melting mechanism.

The treatment with supercritical CO2 of a new alkylating anticancer drug, at

55 C and 40MPa, originated a new crystal phase via a solution-mediated process.


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Carbamazepine polymorphs were also investigated in order to obtain the pure

crystalline phase stable at ambient temperature (form III) by means of supercritical CO 2.

An equimolar mixture of carbamazepine polymorphs I and III was processed with

supercritical CO2 to obtain a crystallographically pure phase. It has been proved that the

suspension in supercritical CO2 leads to an almost quantitative conversion of form I into

form III (Table 3). The enrichment of the mixture in terms of form III is essentially due to

the conversion of form I via solubilization in supercritical CO2 followed by re-

crystallization of the less soluble and more stable polymorph.

In another example, deoxycholic acid powders were stored in a pressure

vessel purged with carbon dioxide at 12MPa and 60C for fixed time intervals. After 1h,

new peaks in the powder X-ray diffraction pattern suggested that a new polymorph was

being generated. The conversion was almost quantitative after 6h treatment.

Table 2 Percent of carbamazepine form III obtained

after static treatment of a 1:1 mixture form I: form III

in supercritical CO2 at 35MPa and 55 C (n=4)

SF Treatment (h) %Form 3(S.D.)

0 47.9(3.9)

6 88.3(0.9)

9 90.6(2.8)

23 91.2(4.7)

48 94.7(0.7)

SEDS Process

Salmeterol xinafoate (SX) is an anti-asthma drug administered by inhalation

and thus requiring particle size within 25m. Commercial SX, generated by a

conventional crystallization process, has very poor flow properties, and it is unsuitablefor size reduction with a fluid energy mill. Two different approaches to SX

recrystallization as fine particles, i.e. fast cooling crystallization and the SEDS process,

were investigated. In the fast cooling crystallization, SX was first dissolved in a hot

organic solvent and then quenched in a chilled solvent to induce supersaturation and

consequent precipitation of the drug. A wide range of solvents and experimental


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conditions was examined: SX with good flow properties, suitable for further milling

down to 25m-sized particles, was obtained. In the second approach, solutions of SX in

ethanol and in acetone were introduced in the SEDS apparatus in differing working

conditions (i.e., temperature, pressure, solution concentration, and supercritical carbon

dioxide flow rates). This process was able to provide, without extra milling operation,

microfine free-flowing crystalline particles (25 m), virtually free from retained solvent.

Interestingly, the SEDS process, unlike conventional crystallization

procedures, was able to produce specific polymorphs of SX, depending on the operating

conditions. The fast cooling crystallization gave exclusively SX form I, namely the stable

polymorph at ambient temperature. Moreover, micronised SX obtained by SEDS showed

superior crystalline purity, with respect to commercial micronized batches (MSX, form

I). Surface thermodynamic properties of both solid phases (obtained by SEDS or MSX)

were compared by inverse gas chromatography. Metastable SX form II (prepared by the

SEDS process) exhibited higher surface entropy and free energy and a more polar surface

than the stable form I (also prepared by the SEDS process), whereas MSX displayed a

higher surface free energy and enthalpy than SX form I.

B) PARTICLE COATING

Conventional coating processes are carried out by nebulizing an aqueous or

organic solution of the coating material (sugar, polymer, wax) onto the solid dosage form.The use of aqueous solutions eliminates many disadvantages associates with organic

solvents; however, it increases the drying time due to the higher latent heat of

vaporization of the water relative to organic solvents. Furthermore, the number of the

material, in particular polymers, that can be dispersed or dissolved in water is limited.

Last but not least, the coating of the small particles still represents an issue. Solvent less

coating technologies may overcome some of the above-mentioned disadvantages, and

reduce the overall cost by eliminating the slow and expensive process of solvent removal.

A fluidized-bed coating process based on the RESS was described by

Tsutsumi et al. The coating granulation process consists of three main steps: extraction,

expansion and fluidization. In a typical experiment, CO2 flows through the melted

coating material in an extraction column, where the CO2 becomes saturated with the

solute. The bed is fluidized by adding pure CO2 via a bypass line; thus the CO2 acts as


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carrier fluid as well. The CO2, saturated with the coating material, expands through a

nozzle into the fluidized bed, which consists in a column installed inside an autoclave.

The solid final product is separated from the gas by a cyclone and several filters. The

column is equipped with two pressure sensors: one at the bottom and one on the top of

the column. These allow the control of the pressure drop across the fluidized bed and

calculation of the minimum fluidization velocity. Below this last the bed is packed, while

above the bed expands and the particles move.

Process key parameters are temperature, pressure, solidification kinetics of the

coating material, and fluidization gas velocity. The relatively low pressure and

temperature as well as the absence of organic solvents enable the use of such process for

encapsulating sensitive material like proteins. These authors tried to coat with paraffin

irregularly shaped particles of bovine serum albumin (BSA) and insulin, by means of

RESS process in a high-pressure fluidized bed running with SC-CO2. The fluidization

behavior of BSA was improved by adding free flowing lactose. Lactose was mixed with

BSA in different volume percentages and the minimum fluidization velocity of each

mixture was measured at fixed pressure and temperature (812MPa and 4055 C) by

stepwise increasing the SC-CO2 flow rate. It was noticed that the minimum fluidization

velocity decreased by increasing pressure as well as by decreasing temperature; similar

effects were found also by. The obtained coated particles were tested for dissolutionbehavior in comparison to the uncoated particles. Dissolution retardation of even more

than 180min, indicating improved encapsulation, was achieved for certain experiments.

Although this technique appears to be very promising, to date only few

experiments have been done and too few data are available to express an opinion about

its future application in pharmaceutics.

A supercritical fluid coating process by using a simple autoclave equipped

with an rotating impeller was used to prepare BSA microparticles coated with two lipids.

Protein particles were coated with trimyristin (Dynasan 114) and Gelucire 50-02, two

glyceride mixtures with a melting point of 45 and 50 C, respectively. Dynasan 114

afforded a discontinuous coating constituted by disordered micro-needles. This led to a

drug release kinetics exhibiting a significantly high initial burst (35% in 5 min); then 70

and 85% of the protein was released in 30 min and 5 h, respectively. On the other hand, a


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prolonged protein release (over 24 h) was achieved with particles coated with Gelucire

50-02, which produced a more homogeneous film coating. It was also shown that BSA

did not undergo any degradation following the SC-CO2 treatment.

Noteworthy, lipids are quite soluble in pressurized CO2, while the most

commonly used coating materials including many polymers show very low solubility in

this supercritical fluid. This problem can be overcome by adding a co-solvent to the

coating solution. Compared to conventional techniques the amount of organic solvent

used here is greatly reduced. Patented an anti solvent process to prepare microcapsules

containing active ingredients with a polar polymer film (polysaccharide, cellulose

derivates, acrylic or methacrylic polymers, polymers of vinyl esters, polyesters,

polyamides, polyanhydrides, polyorthoesters, or polyphosphazenes). The procedure

implied the suspension of the active ingredient in an ethanol solution of the polymer,

followed by mixing with SC-CO2 for the ethanol extraction.

Mishima et al. used a method called rapid expansion from supercritical

solution with a non-solvent (RESS-N) for the microencapsulation of proteins (lipase and

lysozyme) with polymers such as polyethylene glycols (PEGs), poly methyl methacrylate

(PMMA), polylactic acid (PLA), polylactide-co-glycolide (PGLA) and PEG

PPG(polypropylenglycol)PEG triblock copolymer. The polymer, protein and ethanol

were placed inside an autoclave, then the mixture was stirred, and the CO2 pumped in atfixed temperature and pressure. Non-agglomerated proteins containing microparticles

were obtained upon the expansion of the polymeric solution. The thickness of the

polymer coating, the mean particle diameter as well as the particle size distribution, could

be controlled by changing the polymer feed composition.

C) PARTICULATE DOSAGE FORM:

1) CYCLODEXTRIN INCLUSION COMPLEXES:Cyclodextrins are cyclic oligosaccharides able to fully or partially include in their

hydrophobic internal cavity a guest molecule of appropriate size. This complexation

allows to improve some physico-chemical properties such as solubility, dissolution rate,


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Stability, as well as organoleptic characteristics. Conventional preparation methods of

CDs inclusion complexes include kneading, co-precipitation, co-evaporation, co-

grinding, and freeze- and spray drying.

Studies on the feasibility of supercritical fluid processes to produce inclusion complexes

between drugs and CDs in the solid-state are reported in literature. A successful

complexation (94% inclusion yield) between piroxicam and -cyclodextrin (-CD) was

obtained by. The inclusion experiments were performed by keeping a physical mixture of

-CD and piroxicam for 6 h in contact with CO2 at 150 C and 15MPa without the use of

organic solvents.

By using a similar method Moneghini et al. tried to include nimesulide into -CD. The

obtained results indicated that the drug was only partially included. However, the

physico-chemical characterization of the obtained product pointed out the existence of

interactions between drug and carrier that led to an increased in vitro drug dissolution

rate.

Charaoenchaitrakool et al. prepared a complex of methyl--cyclodextrin (M--CD) and

ibuprofen affording the M--CD solid/liquid transition as a consequence of the

interaction with SC-CO2. The same research group used an ASES process for producing

micron-sized naproxen formulation incorporating hydroxyl propylated- and methylated-

-CD.A controlled particle deposition process using SC-CO2 (CPD) was developed by Turk et

al. to produce complexes of racemic ibuprofen and -CD. In this method, ibuprofen and

-CD were filled in to two separates cartridges as pure compound and in another

cartridge, as a physical mixture: the cartridges were inserted in a high-pressure cell

heated and pressurized at the desired values (40 C and 25 or 30MPa). After 15 h under

stirring, the system was depressurized within 10 min. The maximum inclusion yield, in

the case of separate cartridges was 88%, and 60.5% in the case of the physical mixture.

Both the dissolution rate coefficient and the dissolved amount after 75min of the CPD

complexes were found to be significantly higher than those of unprocessed ibuprofen

alone and ibuprofen/-CD physical mixture.

A SEDS process (10MPa, 4080 C) was carried out to produce crystalline channel-type

and amorphous -CD particles and crystalline channel-type -CD complexes in a single-


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step process. The increase of process temperature changed the crystallinity of-CD. In

particular, at 80 C amorphous -CD was obtained, while the complexes crystallized a

tetragonal channel-type form to hexagonal channel-type form. The dissolution behavior

of budesonide/-CD complexes depended on their crystal structure: the tetragonal

dissolved faster than hexagonal form.

This represents a nice example of-CD crystallinity change with subsequent variation of

the dissolution rate of complexed budesonide stemming from the modification of the

operative process conditions.

SAS process was used for the preparation of an inclusion complex between simvastatin

(SV) and hydroxypropyl--cyclodextrin (HP--CD). The activity of the SV/HP--CD

Complexes was tested in vivo in comparison with uncomplexed simvastatin. SV/HP--

CD inclusion complex performed better than SV in reducing total cholesterol and

triglyceride levels; this could be primarily attributed to the improved solubility and

dissolution of the complex. In another study, the feasibility of the ASES process for

preparing solid-state inclusion complexes of itraconazole (ITR) with HP--CD was

investigated. ITR/HP--CD complexes with a significantly higher dissolution rate (90%

of ITR dissolved within 510min) than the unprocessed ITR and its physical mixture

were obtained.

2)

EXTRUSION:The above-mentioned capability of SC-CO2 to plasticize polymers at low

temperature can be exploited in the extrusion process. Here, the SC-CO2 can both change

the rheological properties of the material, and behave as an expansion agent. The

dissolution of a large amount of SC-CO2 determines a polymer expansion and viscosity

reduction. The viscosity reduction results in lower mechanical constraints and decreases

the required operating temperature, thus allowing processing of thermolabile compounds.

Verreck and co-workers carried out extrusion of PVP-VA (polyvinylpyrrolidone-co-vinyl

acetate), Eudragit and ethylcellulose, by means of a twin-screw extruder in which

pressurized CO2 was injected at a constant pressure rate. The physico-chemical

characteristics of the polymer before and after injection of CO2 were evaluated. The

specific surface area and the porosity of the polymers increased after treatment with

carbon dioxide, eventually resulting in enhanced polymer dissolution in water.


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The expansion of the CO2 at the nozzle determined a change of the

morphology of the system toward a foam-like structure. The same observations were

done in the case of itraconazole/polymer combinations as well. Another work illustrates

the use of SC-CO2-assistedextrusion for the preparation of a hot melt extruded monolithic

polymer matrix for oral drug delivery. The polymer used was polyethylene oxide (PEO)

while the model drug was carvedilol. The SC-CO2-assisted process gave rise to extruded

material with faster dissolution compared to that obtained with a classical extrusion

process.

In conclusion this represents an interesting example of process

contamination where the introduction of a SCF technology into a conventional

technique leads to a better performing material.

3) LIPOSOMES PREPARATION:The preparation of stable liposomes formulation on industrial scale is still amajor issue in

pharmaceutics mainly due to the need of the large amount of organic solvents and the

high energy consume. In this respect, SCF-based technologies have attracted a great deal

of interest during the past 10 years, as a green alternative to classical methods.

Pioneering work in this field was done by Frederiksen and coworkers, based on

amodification of RESS process. The apparatus was mainly composed by two parts: a

high-pressure system in which phospholipids and cholesterol were dissolved into thesupercritical phase and a low pressure system in which the supercritical phase (containing

also a various amount of ethanol) was expanded and simultaneously mixed with a water

phase containing dextran and fluorescein isothiocyanate to yield liposomes. Liposomes

with a diameter of 200 nm were obtained. Noteworthy, this technique get the same

encapsulation efficiency but required 15-fold less organic solvent, compared to the

ethanol injection method of Batzri and Korn (1973). Castor and Chu (Castor, 2005;

Castor and Chu, 1998) presented a Supercritical FluidsTM CFN (SFS-CFN) apparatus

formaking liposomes containing hydrophobic or hydrophilic drugs featuring critical,

subcritical or near-critical CO2. Such fluids were used to solvate phospholipids,

cholesterol and other raw materials.

After a specific mixing time the resulting mixture was decompressed by means of an

injection nozzle into a chamber that contained phosphate-buffered saline or another


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biocompatible solution with the hydrophilic drugs (recombinant proteins, RNA, DNA).

SCF depressurization and phase-conversion into a gas (with formation of bubbles)

created a deposition of the phospholipids in the aqueous phase and, consequently, the

formation of phospholipid bilayers encapsulating the hydrophilic drug. In the case of

hydrophobic drugs, the phospholipids and the target compounds were solvated

simultaneously in a SCF cocktail which was dispersed continuously into the aqueous

environment. In a third variation of this technique, hydrophobic drugs and phospholipids

were directly solvated in the SCF prior to injection into an aqueous solution.

The characteristics of the obtained liposomes depended on several process parameters:

size and design of the nozzle (affecting the bubble sizes and therefore the size of

liposomes formed); decompression rate which defines the characteristics of the deposited

phospholipids as well as the intensity of the mixing (slow decompression produced

smaller and more uniform liposomes); interfacial forces between the SCF and the

aqueous phase; charge distribution of the liposomes and its interaction with the

surrounding aqueous medium (pH, ionic strength and electrolyte composition); the nature

of compounds being encapsulated.

It was found that liposomes formed with a 0.5mm nozzle with lecithin at28MPa and60

Chad an average diameter of 478nm with unimodal distribution, while liposomes formed

with 0.6mm nozzle under identical condition had an average diameter of 326 nm. Theincorporation efficiency varied between 1% for small unilamellar vesicles and 88% for

some multilamellar vesicles (Castor and Chu, 1998). The stability of liposomes was also

tested, by measuring the particle size distribution as a function of time. SC-CO2

liposomes proved to be stable at 4 C over a 6-month period. This technique also afforded

liposomes incorporating molecules such as paclitaxel, camptothecin, betulinic acid,

bryostatin 1, vincristine and doxorubicin, with diameter between 100 and 300 nm.

In vitro and in vivo results suggested that these formulations are more effective than the

commercial available ones. More recently, other authors (Otake et al., 2001) produced

liposomes by using the method described by Castor and Chu. Phospholipids and ethanol

were introduced in a view cell, along with the CO 2. The cell temperature was then raised

up to 60 C, namely a temperature higher than the phospholipid phase transition

temperature, while the pressure was kept at 20MPa. An aqueous dispersion of liposomes


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was obtained through the formation of an emulsion by introducing an aqueous glucose

solution into the cell. Then, the pressure was reduced to release the CO 2, leading to a

homogeneous liposomial dispersion. Transmission electron microscopy indicated that the

vesicles obtained were large ellipsoidal unilamellar liposomes with a diameter from 0.1

to 1.2m, enabling a high entrapping efficiency (up to 20% for hydrophilic substances

and 63% for hydrophobic compounds). The same method was used to prepare chitosan-

coated cationic liposomes for DNA transfection (Otake et al., 2006). Pressurized CO2

forms carbonic acid in contact with water thus it lowers the water poly methyl

methacrylate, swollen with CO2 (Sproule et al., 2004) The experiments were carried out

with a view cell that allow in situ quantification of the polymer swelling by laser

dilatometry. Foaming was studied by means of a X-ray computed tomography, while

protein impregnation was verified by confocal microscopy. Finally, Barry et al. produced

bovine chondrocytes containing methacrylate scaffolds. Polymer foaming was achieved

by placing PEMA/THFMA discs under CO2 at 40 C and 10MPa for a time ranging from

1 to 48 h, then by rapidly depressurizing the system (Barry et al., 2004). The study

demonstrated that the change in the structure of the substrate from flat disk to foam

enhanced the cell phenotype retention, although the authors concluded that further

modifications in the SC processing were required to obtain optimum porosity for cell

migration and cartilage tissue formation.


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FIG.-9 Process Flow Sheet of Liposom e

The 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and cholesterol

were dissolved in supercritical carbon dioxide modified with ethanol. Rapid expansion of

the supercritical solution into an aqueous phase containing a marker results in the

formation of liposomes encapsulating the marker (Fig. 9). Liposomes have an average

size of approximately 4050 nm.

4) MICROSPHERES:Microparticles have a variety of structures (Fig. 10):

Particles with irregular geometry, composed of an active substance in form ofaggregates or molecularly dispersed solid embedded in to a matrix. They are

called microspheres;

Particles with spherical geometry, composed of a core of active substancesurrounded by a solid polymeric or proteic shell. They are called microcapsules.

There is no universally accepted size classification of these particles.


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FIG.-10 Stru cture of the Dif ferent M icropart ic les.

However, many workers classify spheres/capsules smaller than 1m as nano-

spheres/capsules and those larger than 1000m as macro-spheres/capsules. Commercialmicro-spheres/capsules typically have a diameter between 3 and 800m and contain 10

90 wt.% carrier/core. A wide range of materials has been embedded/encapsulated,

including adhesives, agrochemicals, live cells, active enzymes, flavors, fragrances,

pharmaceuticals, and inks. Most carrier/shell materials are natural or synthetic organic

polymers, but fats and waxes are also used.

Examples

Debenedetti et al. obtained microparticles by the RESS co-precipitation of a drug(lovastatin) and a biodegradable polymer (poly(D, l-lactic acid) (DL-PLA)).The

co-precipitation of the polymer and the drug led to a heterogeneous population of

microparticles consisting of microspheres containing a single lovastatin needle,

larger spheres containing several needles, microspheres without protruding

needles and needles without any polymer coating.

Mishima described the formation of microspheres of flavones and a polymer(Eudragit- 100 or PEG 6000) by spraying at atmospheric pressure, a suspension of

flavonoids in a supercritical solution of the polymer and a co-solvent.

Sze Tu et al. used the ASES technique for the coprecipitation of a model drug,parahydroxybenzoic acid (p-HBA) with the biodegradable polymers, poly(lactide-

co-glycolide) (PLGA) and poly(l-lactic acid) (PLA) as presented on Fig. 12. A

multiple nozzle assembly arranged coaxially was designed for the co-introduction


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of the drug and polymer solutions (Fig. 11). In this configuration, solutions are

expanded at a much faster rate than in the conventional ASES process since

minute volumes of the solution continually dispersed by the nozzle suddenly enter

a region of high anti-solvent concentration. The rapid expansion causes the solid

to precipitate out of solution almost instantaneously. In this case, supercritical

fluid is used only for its anti-solvent properties. Coprecipitation of the p-HBA and

PLGA resulted in the formation of p-HBA particles coated with the polymer

microspheres. The coprecipitation of p-HBA with PLA resulted in the formation

of a product where the drug and polymer were incorporated together in a fibrous

network.

FIG.-11 Process flow sheet b y Sze Tu et al.


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References:

1) Bansal A. K., Kakumanu V. K., Supercritical fluid technology in pharmaceuticalresearch. Business Briefing: Labtech 2004. 70-72.

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3) Giordino F., Pasquali I., Ruggero B., Solid-state chemistry and particle engineeringwith supercritical fluids in pharmaceutics. European Journal of pharmaceutical

sciences, 27(2006), 299-310.

4) Fages J., Lochard H., Letourneau J. J., Sauceau M., Rodier E., Particle generation forpharmaceutical applications using supercritical fluid technology. Powder Technology,

141(2004), 219-226.

5) Giordino F., Pasquali I., Ruggero B., Supercritical fluid technologies: An innovativeapproach for manipulating the solid state of pharmaceuticls. Advanced Drug Delivery

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6) York P., Strategies for particle design using supercritical fluid technologies.Pharmaceutical Science and Technology Today, Volume-2 Number-11, 1999, 430-

440.

7) Kiran E., Yeo S. D., Formation of polymer particles with supercritical fluids: AReview. The Journal of Supercritical Fluids, 34(2005), 287-308.

8) Perrut M., Jung J., Particle design using supercritical fluids: Literature and patentsurvey. The Journal of Supercritical Fluids, 20(2001), 179-219.

9) Giordino F., Bonassi L., Ruggero B., Castoro V., Rossi A., Zema L., Gazzaniga A.,Solubility and conversion of carbamazepine polymorphs in supercritical carbon

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Supercritical Fluid and Subcritical Fluid

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