nowadays , the analytical instrumentations can provide a high resolution separation and low detection limits , down to picograms or below ; the whole progressive analytical process can be wasted if an unsuitable sample preparation method has been employed .
thus , sample preparation is of crucial importance for the analysis of drugs in biological samples .
the role of sample preparation is to remove interferences and analyte preconcentration , converting the analytes to suitable form for separation and detection .
if an unsuitable sample preparation method has been employed before the injection , the whole analytical process may be wasted .
because of the low concentration levels of drug in plasma and the variety of the metabolites , the selected extraction technique should be virtually exhaustive . in recent years
, solid phase microextraction ( spme ) has been used by many researchers ever since it emerged in the early 1990s .
spme technique has been widely used in many different areas such as environmental analysis , food analysis , bioanalysis , drug monitoring , and toxicology .
spme can be used as direct immersion of the fiber into the sample to extract the analytes , or introduction of the fiber in the sample headspace to extract volatile compounds that are partitioned between gaseous and liquid phases .
many factors , such as ph , temperature , salt concentration , and stirring , affect the equilibrium constant and equilibration time [ 716 ] .
another microextraction related technique is stir - bar sorptive extraction ( sbse ) which is an extraction technique for enrichment of volatile and semivolatile organic compounds having high extraction efficiency compared to spme but has longer extraction time .
the technique has been applied effectively in environmental analysis mainly with gas chromatography - mass spectrometry ( gc - ms ) .
in addition , the sbse technique was applied to some drugs in biological samples in combination with gc - ms .
recent development in solid phase microextraction related techniques is microextraction by packed sorbent ( meps ) .
meps is the miniaturization of conventional spe and can be connected online to gc or lc without any modifications [ 1822 ] .
the extraction steps in meps are the same as in standard spe  extraction , loading , washing , and elution  and these have to be optimized to obtain the highest analyte recovery . in meps , solvent and sample volumes are significantly reduced compared to spe .
the meps technique has been used to extract a wide range of drugs and metabolites in biological fluids such as urine , plasma , and blood .
this review presents recent developments of sample preparation in drug bioanalysis of complex fluids using spme , sbse , and meps .
in spme , the extraction is based on the partitioning of the analyte between the organic phase on the fused silica fibre and the matrix .
many factors , such as ph , temperature , salt concentration , and stirring , affect the equilibrium constant and the equilibration time .
in addition , type of polymer , temperature , duration , and additives coming from the sample solution influence the stability of the coating .
it should be noted that additives such as sodium hydroxide and salt could catalyse polymer thermal degradation . in some bioanalytical studies , fibre life - time
was decreased to about 20 samplings instead of 80 [ 8 , 9 ] . in the two past decades ,
spme as a sample preparation method for drug analysis has been used with various analysis methods .
spme was used with different separation techniques such as capillary gas chromatography ( cgc ) [ 2330 ] , gc / gc - ms [ 3137 ] , lc / lc - ms [ 3850 ] , gc - icp - ms [ 5157 ] , and hplc - uv / hplc - ms [ 5868 ] . in tube
solid phase microextraction ( in - tube spme ) is a new format of spme that can be coupled online to lc for automated analysis of less volatile and polar compounds like drug metabolites .
this technique was used for the determination of drugs and metabolites in different biological matrices like urine , plasma and cell culture media from in vitro assays [ 6971 ] . in addition
fiber in - tube spme online with capillary electrophoresis ( ce ) was used for the analysis of amitriptyline , imipramine , nortriptyline , and desipramine in human urine samples . in this work ,
one is high modulus ( hm ) with heat treatment and the other is regular ( as ) without heat treatment after spinning the fibers .
in addition , db-5 capillary of 10  mm length was packed with a fiber with the same length . in order to investigate the effect of the fiber and the capillary coating on extraction efficiency , various kinds of extraction media were prepared as follows : hm fiber packed in a db-5 capillary , hm / db-5 ; as fiber packed in a db-5 capillary , as / db-5 ; hm fiber packed in an uncoated fused - silica capillary , hm / fs ; and only db-5 capillary , db-5 . for the evaluation of the effect of the packing density on the extraction efficiency ,
the packing density has been calculated by using the average diameter of the fibers as 11.5  m .
the former was packed with 123 filaments and the latter with 246 filaments in a 10  mm db-5 capillary .
this method was then applied to the analysis of amitriptyline in human urine and the results showed that the hyphenated system would be a powerful tool for the analysis of analytes in biological matrices , db-5 capillary was cut to 10  mm length and the fiber of the same length was packed into this capillary . in order to investigate the effect of the fiber and the capillary coating on extraction efficiency , various kinds of extraction media were prepared as follows : hm fiber packed in a db-5 capillary , hm / db-5 ; as fiber packed in a db-5 capillary , as / db-5 ; hm fiber packed in an uncoated fused - silica capillary , hm / fs ; and only db-5 capillary , db-5 . for the evaluation of the effect of the packing density on the extraction efficiency , 26 and
the packing density has been calculated by using the average diameter of the fibers as 11.5  m .
the former was packed with 123 filaments and the latter with 246 filaments in a 10  mm db-5 capillary .
the density is based on the volume ratio between the space and the filled part of the inner capillary in which the fibers were packed .
the running buffer was composed of 20  mm na2hpo4 buffer ( ph 9.3 ) , 0.6  mm - cyclodextrin , and 20% acetonitrile .
different strategies were developed for preparation of monolithic in tube spme ( figure 1 ) .
a hybrid organic - inorganic silica monolith with ceyanoethyl functional groups was synthesized by hydrolysis and polycondensation of precursors via a two - step catalytic sol - gel process that was used as a sorbent for in - tube spme [ 73 , 74 ] .
. 250  m ) were activated with 1  m naoh and then 1  m hcl . after rinsing with double distilled water ,
they were dried at 160c under n2 flow for 5  h. the hybrid monolith was synthesized by hydrolysis and polycondensation of precursors via a two - step catalytic sol - gel process .
the optimal preparation conditions were as follows : 180  l of methanol , 25  l of 2  m acetic acid , 110  l off cn - teos , and 110  l of teos were mixed in a 1.5  ml eppendorf vial . after thorough vortexing
, the mixture was left for hydrolysis at 60c for 5  h. after cooling to room temperature , 10  mg of n - dodecylamine was added to the solution .
then the pretreated capillary was filled to a certain length with the sol by a syringe .
the capillary was sealed at both ends with silicone rubber and then was allowed to further react at 40c for 15  h. subsequently , the capillary was rinsed with ethanol to remove the n - dodecylamine and soluble hydrolysis products and then dried at 60c for 48  h. the total and effective lengths of the hybrid silica monolith were 20 and 15  cm , respectively . as it is shown in figure 1 ,
four capillaries are connected in the modified cross connector to build the online fiber - in - tube spme - ce system . to minimize the band broadening effects
therefore , the capillaries between the two buffer reservoirs were connected using a microscope until the gap between the capillaries was less than 10  m . in addition , the sem picture ( figure 1 ) showed that the monolith is attached tightly to the inner - wall of the capillary .
the flow - through pores size distribution determined by mercury porosimeter was around 4  m with a narrow size distribution , which results in high permeability and favourable mass transfer in extraction applications .
in another work , a restricted access material ( ram ) was employed for preparation of a lab - made biocompatible in - tube spme capillary that enables the direct injection of biological fluids as well as the simultaneous exclusion of macromolecules by chemical diffusion barrier and drug preconcentration [ 75 , 76 ] . in this work , silica particles ( c1845  m )
were slurried in methanol and packed into 50  mm ( length ) of polyether ether ketone ( peek ) tubing ( i.d .
0.02 inch ) and then the capillary column was capped at both ends by a 1/16  in .
= 2.54  cm ) zero - volume union fitted with a 10  m frit . after this procedure , the capillary was conditioned with phosphate buffer ( 0.05  mol  l , ph 6.0 ) at a flow - rate of 1.0  ml  min for 20  min .
initially , 50  ml phosphate buffer solution ( 0.05  mol  l , ph 6.0 ) was percolated through the capillary at a flow rate of 1.0  ml  min , followed by 25  ml bovine serum albumin solution 1.0  mg  ml ( prepared in phosphate buffer solution ) and by 25  ml glutaraldehyde solution ( 25% , v / v ) .
after 5  h , the columns were washed with 10  ml sodium borohydride solution ( 1.0  mg  ml ) and then with 60  ml water .
the ram - bsa column was stored in phosphate buffer solution ( 0.05  mol  l , ph 7.4 ) at 4c .
in addition , monolithic molecular imprinted polymer ( mip ) fiber based solid phase microextraction ( spme ) was developed for selective and sensitive determination of different drugs and biomarkers in biological samples [ 77 , 78 ] . in situ polymerization of silica
capillary mold using e as template was reported and in some studies the mip fibers are preparedand each fiber could be used for about 50 extraction - cycles without any significant decrease in extraction capacity .  
other types of monolithic in tube spme were prepared by different kinds of monomer and cross - linker mixtures , such as poly(acrylamide - ethylene glycol dimethacrylate ) .
poly(aa - egdma ) monolith was selected as sorbent for spme of three protoberberine alkaloids ( figure 4 ) .
briefly , aa was weighed and put in a 1  ml screw capped glass vial , followed by adding isooctane , toluene , and methanol as porogen .
after aa was completely dissolved , the cross - linker egdma and the initiator aibn were added to the above solution .
finally , the prepolymerization solution was introduced into the modified peek tube carefully , and then the peek tube was sealed and put into a water bath for polymerization ( 60c , 3  h ) .
after polymerization , the monolith was washed with acetonitrile to remove porogen and unreacted reagents , poly(meta acrylic acid - ethylene glycol ) [ 8082 ] , and poly(4-vinylpyridine - co - ethylene dimethacrylate ) .
also , in - tube spme / lc method was developed and validated for rifampicin interferon 2a determination in plasma samples for therapeutic drug monitoring and in plasma samples and lidocaine and its metabolite megx in plasma samples [ 8486 ] . in summary ,
monolithic in - tube spme was shown to be an appropriate method for drug metabolism studies and routine analysis or pharmacokinetics as the parent compound and main metabolites could be monitored in various matrices of interest .
hs - spme , an alternative sample extraction technique , allows concentrating volatile and semivolatile analytes from the headspace above the sample on a coated fiber and to transfer the analytes from the fiber directly into the injector port of a gc without further manipulations .  
table 1 shows the different drugs and pharmaceutical components determined by hs - pme method .
a schematic structure of hs - pme is shown in figure 5 . in some works
in addition , the sol - gel method was used for fiber preparation in hs - pme technique . in the following part
the sol - gel process provides a useful method of preparing organic - inorganic hybrid materials through the hydrolysis and condensation of suitable metal alkoxides , particularly , silicone alkoxide , which readily allows forming three - dimensional ( 3d ) network under relatively mild conditions . 

table 2 lists the names and chemical structures of the principal ingredients of the coating solution used .
the sol - gel process may involve mainly several parts as follows : ( 1 ) ring - opening polymerization between kh-560 and dm--cd ; ( 2 ) hydrolysis and polycondensation among the product of ( 1 ) , teos , and oh - tso to generate a 3d network ; ( 3 ) chemical anchoring of the polymeric networks to the outer surface of the fused - silica fiber ; ( 4 ) deactivating residual silanol groups on the stationary phase with pmhs , aimed to reduce harmful adsorptive effects .
thus , a surface - bonded polymeric coating dm--cd / oh - tso is formed as schematically represented in figure 6 .
molecularly imprinted polymers ( mips ) have proven to be useful materials in analytical chemistry .
mips are cross - linked synthetic polymers obtained by copolymerizing a monomer with a cross - linker in the presence of a template molecule .
after polymerization , the template is removed from the porous network by washing , leaving cavities in the polymeric matrix that are complementary in size , shape , and chemical functionality to the template .
thus , the imprinted polymer is able to rebind selectively the analyte ( the template ) under certain experimental conditions .
accordingly , the combination of molecular imprinting and spme would ideally provide a powerful analytical tool with the characteristics of both technologies , simplicity , flexibility , and selectivity .
there are two strategies in this field ; the easiest way for combining both technologies was proposed by mullet , which consisted of packing a capillary with the mip particles for in - tube spme and was used for the selective determination of propranolol in serum samples .
the developed method was successfully applied and the advantages of in - tube spme were obvious ( high enrichment factors provided by multiple draw / eject cycles , ease of automation , and fast operation ) .
however , this methodology is not free of some important drawbacks such as the lack of compatibility between the solvent needed to desorb analytes from the mip and the mobile phase used ( typical drawback of online mispe protocols ) and the necessity of extra instrumentation ( pump , multiport valves ) .
thus , the preparation of silica fibers coated with a mip to perform spme would be the best option and different works have been developed in this field .
most papers that have been developed in mip - spme field are about fiber preparation for separation of different valuable targets of complex media such as clenbuterol and structural analogues , triazines , diacetylmorphine and analogous compounds , prometryn , bisphenol a [ 97 , 98 ] , anabolic steroids , 2,2-bipyridine , antibiotic drugs , and sulfamethazine .
spme conditions based on the mip - coated fibers are valuable methods that developed in recent years . as simple approach for preparation of bisphenol a ( bpa ) mip - coated spme fibers a capillary was inserted into a larger bore capillary to form a sleeve as mold .
the prepolymer solution , which comprised bpa , acrylamide ( am ) , 3-(trimethoxysilyl ) propyl methacrylate ( trim ) , aibn , and acn , was introduced into the interspace between the two capillaries , followed by polymerization under uv photoirradiation ( figure 9 ) .
this approach showed that this very simple method could become a routine preparation procedure for mip - coated fibers .
the mip coating on the silica fibers was homogeneous and porous and showed good mechanical and chemical stability .
according to the result as of this work , it was demonstrated that the mip - coated fibers had better adsorption / desorption kinetics compared with the monolith mip fiber . under the optimized spme conditions ,
selective extraction of bpa from standard mixture aqueous sample was feasible with the mip - coated fibers .
since sbse was developed in 1999 , it has already shown significance among the sorptive extraction techniques .
sbse and spme are microextraction techniques with low , or even no , consumption of organic solvents .
the analytes are extracted from the matrix into the polymer coating immobilized on a glass tube with a magnetic core .
rapid molecular - recognition equilibrium between adsorption and desorption can be established , since sampling is performed simultaneously with the stirring . as a result , competitive sorption from an additional stirrer ( e.g. , magnet essential for the spme technique )
the main differences between the two techniques are the design of the extraction system and the amount of the sorbent material .
the sorbent materials are similar , although till today the availability of commercial sbse materials is rather limited .
contrary to spme , quantitative recoveries are often achievable with sbse due to the clearly higher sample capacity .
quantitative extraction can be achieved for solutes with log ko / w values of ca .
4 , and reasonable efficiencies are obtained for solutes with log ko / w values above 3   for highly polar compounds , similar approaches as for spme can be applied ( i.e. , derivatisation ) .
same as spme in sbse various parameters such as type and thickness of the coating , extraction time , sample properties ( ph , ionic strength ) , agitation , temperature , and analyte desorption could be evaluated .
the extraction time is typically longer than in spme , because the amount of coating is greater and it takes longer to reach equilibrium .
the analyte desorption is more critical for sbse than for spme , likewise due to the greater amount of coating .
a high flow rate of gas ( up to 100  ml  min ) is recommended for fast desorption of analytes during thermal desorption .
used sol - gel technology in stir - bars to produce a partially hydroxyterminated - pdms coated stir - bar , which was used for extracting a group of pahs and organophosphorous compounds .
the sol - gel process offers a convenient , versatile pathway for preparing advanced inorganic and organic - inorganic hybrid material systems , with tunable porosity , selectivity , and thermal and chemical stability .
the schematic of sol - gel reactions is shown in figure 10 ( where methyltrimethoxysilane ( mtmos ) and hydroxy - terminated polydimethylsiloxane ( pdms ) are shown to represent sol - gel precursor and sol - gel active organic polymer , resp . ) . despite its numerous advantages over conventional spme fibers
, sbse also suffers from serious limitations   as follows : limited number of commercially - available coatings;coating is not chemically bonded to the substrate leading to the possibility of bleeding at even relatively low temperature during thermal desorption and transfer of the extracted analytes from the stir - bar to the gc system;coating is vulnerable to washing away if proper solvent is not used during solvent desorption;thermal desorption requires an expensive thermal desorption unit;thick , highly viscous polymeric sorbents used on the stir - bar require hours to reach the extraction equilibrium;there is a need for a relatively high volume of back extraction solvent , which evidently dilutes the preconcentrated analytes .
limited number of commercially - available coatings ; coating is not chemically bonded to the substrate leading to the possibility of bleeding at even relatively low temperature during thermal desorption and transfer of the extracted analytes from the stir - bar to the gc system ; coating is vulnerable to washing away if proper solvent is not used during solvent desorption ; thermal desorption requires an expensive thermal desorption unit ; thick , highly viscous polymeric sorbents used on the stir - bar require hours to reach the extraction equilibrium ; there is a need for a relatively high volume of back extraction solvent , which evidently dilutes the preconcentrated analytes .
high porous sol - gel pdms coated stir - bar with 30  m coating thickness was developed by liu et al . .
the sol - gel pdms coated stir - bars were tested for the extraction of n - alkanes , pahs , and organ phosphorus pesticides .
the sol - gel pdms coated stir - bar reached extraction equilibrium in less than 15  min .
in addition , unlike commercial pdms coated stir - bars , the sol - gel pdms coated stir - bar is equally suitable for both polar and nonpolar analytes .
a sol - gel pdms / pva coated stir - bar for the extraction of organophosphorus pesticides ( opps ) in honey samples was used .
the back - extraction solvent , which contained the analyte(s ) of interest , was then injected into the gc using large volume injection , followed by gc - fpd .
the extraction performance of sol - gel cw / pdms / pva was compared with commercial pdms stir - bar and carboxen / pdms spme fiber using headspace extraction .
sol - gel cw / pdms / pva coated stir - bar demonstrated the highest sorption capacity and ~10 times higher sensitivity . also , a sol - gel pdms/-cd coating ( 30150  m ) for the extraction of polar compounds from different matrices ( e.g. , estrogens in environmental water and bisphenol a in drinking water ) .
the same sol - gel sorbent ( sol - gel pdms/-cd ) was utilized for extracting brominated flame retardants from soil and dust samples employing ultrasound - assisted extraction followed by hplc analysis .
sbse performances were compared for four different sol - gel coatings including pdms / b - cd , pdms , cw / pdms / pva , and pdms / pva .
the sol - gel pdms/-cd coated stir - bar was found to be the most efficient for the target compounds .
in addition to superior extraction performance , sol - gel pdms/-cd coated stir - bars demonstrated excellent durability , and no discernible loss of extraction efficiency was observed even after 100 extraction cycles .
mi - sbse is based on the partitioning of target analytes between a liquid sample and a stationary phase - coated stir - bar . until now
, only polydimethylsiloxane ( pdms ) coated stir - bars are commercially available , restricting the range of applications to the extraction of hydrophobic compounds ( organochlorine and organophosphorus pesticides ) due to the polar character of pdms . besides , the mip - coated stir - bars showed not only the expected high selectivity but also rapid equilibrium adsorption , thanks to the porous nature of the imprinted polymer obtained combined with a suitable thickness of coated polymer film ( 160180  lm ) .
more recently , the use of mip - coated stir - bars prepared by chemical bonding of the mip to the stir - bar through silylation of the substrate surface and then multiple copolymerization reaction was proposed for the determination of various components in different samples [ 113117 ] .
the schematic diagrams of the preparation of mip - sbse coating using terbuthylazine as template molecule is shown in figure 11 .
the preparation of monolithic materials is very simple just by polymerization of a monomer mixture with a porogen solvent , forming a porous polymer . in this way ,
huang and yuan developed monolithic material obtained by in situ copolymerization of octyl methacrylate and ethylene dimethacrylate in the presence of a porogen solvent containing 1-propanol , 1,4-butanediol , and water with azobisisobutyronitrile as the initiator .
the results demonstrate that prepared stir - bar was suitable for preconcentration of both apolar and polar analytes .
the enrichment factors for phenanthrene , anthracene , and pyrene were 150 , 134 , and 189 , respectively .
the sbsem shows good batch - to - batch reproducibility and good stability and can be reused at least 10 times for the extraction of polycyclic aromatic hydrocarbons in seawater ( figure 12 ) . in another in situ copolymerization approach , vinylpyrrolidone and divinylbenzene in the presence of a porogen solvent containing cyclohexanol and 1-dodecanol with azobisisobutyronitrile as initiator
polycyclic aromatic hydrocarbons were used to investigate the extraction efficiencies of sbsem for apolar analytes .
hormones , aromatic amines , and phenols were selected as test analytes to investigate the extraction efficiencies of sbsem for weakly and strongly polar compounds .
the results showed that the new sbsem could enrich the above - mentioned organic compounds effectively .
it is worthy to mention that the sbsem can enrich some heavy metal ions , such as cu , pb , cr , and cd , through coordination adsorption . in another case ,
poly(vinylpyridine - ethylene dimethacrylate ) is used as sbsem combined with high performance liquid chromatography with diode array detection under the optimized experimental conditions for analysis of target compounds in wastewater samples .
the method showed good linearity and repeatability , as well as advantages such as sensitivity , simplicity , low cost , and high feasibility .
meps was recently introduced as a novel method for sample preparation , being a miniaturization of the conventional spe technique , in which the sample volume , extraction , and washing solvents volumes are greatly reduced compared to spe .
meps differs from commercial spe in that the packing is integrated directly into the syringe and not into a separate column .
moreover , the packed syringe can be used several times , more than 100 times using plasma or urine samples , whereas a conventional spe column is used only once .
meps can handle small sample volumes ( 10  l plasma , urine , or water ) as well as large volumes ( 1000  l ) .
the extraction steps in meps are the same as in standard spe extraction , washing , and elution and these have to be optimized to obtain the highest analyte recovery
. in meps extraction procedures , additional steps for postcleaning and reconditioning have to be included to enable multiple uses of the meps sorbent .
blood samples require dilution of 2025 times with water or acidic water , while 4 - 5 times dilution is needed for plasma samples . using mixed - mode ( anion - cation exchange )
meps was used with real samples from clinical institutions or pharmaceutical industry for established drugs and new drug candidature ( figure 13 ) .
the superior performance of meps was recently illustrated by online lc - ms and gc - ms assays of drugs and metabolites in water , urine , plasma , and blood samples [ 1822 , 121153 ] .
the combination of meps and liquid chromatography mass spectrometry ( lc - ms ) is a good tool for the screening and determination of drugs and metabolites in blood , plasma , and urine samples .
mass spectrometry is presently one of the most powerful detection techniques , particularly in pharmaceutical analysis , where good selectivity and high sensitivity are often needed .
this approach to sample preparation is very promising for many reasons : ( 1 ) it is easy to use , ( 2 ) it is a fully automated online procedure , ( 3 ) it is rapid , and ( 4 ) the cost of analysis is minimal compared to conventional solid - phase extraction .
plasma samples were diluted four times while blood samples diluted 20 times with water . 

the sorbent was conditioned with 150  l methanol and subsequently with 150  l water . 

sample can be loaded by multiple aspirates - dispenses cycles ( 5  100  l ) . 

the meps technique has been used to extract a wide range of analytes in different matrices ( urine , plasma , and blood ) .
hence , several drugs such as local anesthetics and their metabolites , the anticancer drugs roscovitine , olomoucine , busulphan , cyclophosphamide , and azd3409 , the -blockers acebutolol and metoprolol , the neurotransmitters dopamine and serotonin , methadone , and cocaine and cocaine metabolites have been extracted from biological samples such as blood , plasma , or urine samples using meps [ 1822 , 121153 ] .  
table 3 shows the application of meps technique in different matrices and various types of drug .
molecularly imprinted polymer was used as extracting sorbent in meps for the simultaneous determination of four local anaesthetics ( ropivacaine , lidovacaine , bupivacaine , and mepivacaine ) in human plasma and urine samples have been evaluated . in comparison with protein precipitation ,
. this may be important for increasing sensitivity and for robustness of the lc - ms system .
ion suppression was observed in protein precipitation method for lidocaine , ropivacaine , and mepivacaine .
the matrix effect ( me ) for these substances ranged from 50% to 146% using protein precipitation while it was less than 20% using mip - meps .
this suggests that cleaner extracted samples can be obtained with meps and that matrix related problems can be reduced . in comparison with conventional spe
meps technique online with lc - ms / ms was used for the quantification of snsr receptors agonist peptide bam8 - 22 and antagonist bam22 - 8 in plasma samples .
meps - c8 was used and spiked plasma sample ( 125  l ) was diluted ( 1  :  1 ) with 0.1% chooh in water ; 50  l were drawn onto the meps - syringe three times .
the sorbent was then washed once with 50  l of 5% methanol in water to remove other interferences .
the analytes were then eluted by 40  l 0.25% ammonium hydroxide in methanol - water 95  :  5 ( v / v ) directly into lc injector .
the between - batch accuracy and precision for bam8 - 22 ranged from 13 to 2.0% and 4.0 to 14% , respectively . additionally , the accuracy and precision for bam22 - 8 ranged from 13 to 7.0% and from 3.0 to 12% , respectively .
meps technique provided significant advantages such as the speed and the simplicity of the sample - preparation process . compared with other extraction techniques , such as protein precipitation and ultrafiltration , meps gave cleaner samples and higher recovery ( > 90% ) .
furthermore , the method reduced the sample preparation time for bams ( less than one minute per sample ) , which is of great importance to handle unstable analytes such as bams in plasma and blood samples .
the simplicity of meps technique will continue to attract interest among analytical chemists searching for improved analysis methods . in summary ,
the significant advantages of meps are reduction in the amount of sorbent bed , solvents , and sample preparation times and reduction in carry - over effects .
recently we introduced a 96-tips set packed with a plug of a monolithic adsorbent . using such a set ,
packed 96-tips sample preparation is a clean , highthroughput , and automated sample - preparation method .
samples are prepared in a 96-well plate format and the analytes adsorb onto the polymer - based monoliths in the extraction step . the next step purifies the sample by washing the sorbent with an appropriate washing solution . in a final step
, the analytes were directly eluted into a 96-well plate using an appropriate solvent for the analytes and the subsequent instrumental analysis .
the polymerization mixture of methacrylate monoliths consists of a solution containing glycidyl methacrylate ( 20% ) , ethylene glycol dimethacrylate ( 15.5% ) , butyl methacrylate ( 3.5% ) , aibn ( 1  wt% with respect to monomers ) , and 1-dodecanol ( 30% ) and cyclohexanol ( 30% ) was vortexed for 10  min and purged with nitrogen for 10  min in order to remove oxygen .
the pipette tips were filled with about 8  mm ( 6 - 7  l ) by the capillary action and placed vertically inside the polymerization apparatus .
the polymerization using uv light at 254  nm was allowed to proceed first for 60  min with the sharp end of the tip down and at a distance to the lamp of 15  cm and then for 25  min with the sharp end up and at a distance of 5  cm to the lamp .
after completion of polymerization , the tips were removed , inspected under microscope for bubbles , and washed with acetone to remove the porogenic solvents and other compounds remaining in the monolith ( figure 14 ) .
the key aspect of the monolithic phase is that monolithic material can offer both good binding capacity and low back - pressure properties compared to , for example , silica phases . using this device ,
evaluation of monolithic packed 96-tips for the extraction of drugs [ 154159 ] such as anticancer drugs ( busulphan , cyclophosphamide , roscovitine ) , -blocker drugs ( metoprolol , pindolol ) , and local anesthetics ( lidocaine , ropivacaine , bupivacaine ) from human plasma or blood samples has been developed and validated ( table 4 ) . utilizing plasma samples , the tips could be used several times ( 5 times ) and still get good results . utilizing blood samples ,
it was shown that small sample volumes can be handled , solvent consumption was low , and the procedure was very fast ( 2  min per 96-well plate ) ( table 4 ) .
the advantages of various kinds of spme , sbse , and meps as powerful sample preparation methods in bioanalysis have been demonstrated and illustrated in many cases .
it is clear that the number of the papers published in this research area has increased during the last decade . in addition , the results showed the ability of these techniques for determination of drugs in biological samples .
future work should be focused on finding of more selective , high throughput adsorption capacity and stable sorbents with capability for extraction of large molecules .
 in drug discovery and development , the quantification of drugs in biological samples is an important task for the determination of the physiological performance of the investigated drugs . after sampling , 
 the next step in the analytical process is sample preparation . because of the low concentration levels of drug in plasma and the variety of the metabolites 
 , the selected extraction technique should be virtually exhaustive . 
 recent developments of sample handling techniques are directed , from one side , toward automatization and online coupling of sample preparation units . 
 the primary objective of this review is to present the recent developments in microextraction sample preparation methods for analysis of drugs in biological fluids . 
 microextraction techniques allow for less consumption of solvent , reagents , and packing materials , and small sample volumes can be used . in this review 
 the use of solid phase microextraction ( spme ) , microextraction in packed sorbent ( meps ) , and stir - bar sorbtive extraction ( sbse ) in drug analysis will be discussed . 
 in addition , the use of new sorbents such as monoliths and molecularly imprinted polymers will be presented . 
1. Introduction
2. Solid Phase Microextraction (SPME) for Drug Analysis
3. Stir-Bar Sorptive Extraction (SBSE)
4. Microextraction by Packed Sorbent (MEPS)
5. Monolithic Packed 96-Tips for High Throughput Bioanalysis
6. Conclusions