Hollow fiber liquid‐phase microextraction and determination of nonsteroidal anti‐inflammatories by capillary electrophoresis and sulfonamides by HPLC in human urine

In this paper two applications of three‐phase HF‐LPME for the determination of pharmaceuticals in human urine are proposed: a capillary electrophoresis with a photodiode array detection method for the analysis of seven nonsteroidal anti‐inflammatory drugs (NSAIDs) and a high‐performance liquid chromatographic with photo diode array and fluorescence detection method for the determination of four sulfonamides and their corresponding N⁴‐acetyl‐metabolites. Q3/2 Accurel® polypropylene hollow fibers were used for both procedures. Dihexyl ether was used as the supported liquid membrane for the determination of anti‐inflammatories and 1‐octanol for sulfonamides. An aqueous solution (pH 12) was used in both procedures as the acceptor phase and as the donor phase an aqueous solution (pH 2), and a 2 m Na₂SO₄ aqueous solution (pH 4) was used for the determination of the anti‐inflammatories and sulfonamides. The detection limits obtained were between 0.25 (naproxen) and 0.86 ng/mL (aceclofenac) for the determination of anti‐inflammatories and 7 × 10^?4 (sulfamethoxazole) and 0.048 ng/mL (N⁴‐acetyl‐sulfamethazine) for sulfonamides. The method was successfully applied to the determination of the analytes in human urine. Copyright © 2012 John Wiley & Sons, Ltd.     

Multiwalled carbon nanotubes as sorbent for on-line coupling of solid-phase extraction to high-performance liquid chromatography for simultaneous determination of 10 sulfonamides in eggs and pork

The sulfonamides (SAs) have been widely used as effective chemotherapeutics and growth promoters in animals' feeding, but their residues could be a potential danger to human health due to their carcinogenic potency and possible antibiotic resistance. Development of a simple and sensitive method for the determination of SAs residues in food of animal origin, therefore, is of great significance. An on-line solid-phase extraction (SPE) method using multiwalled carbon nanotubes as sorbent coupled with high-performance liquid chromatography (HPLC) for simultaneous determination of 10 sulfonamides (SAs) in eggs and pork was developed. The adsorptive potential of carbon nanotubes for solid-phase extraction of sulfonamides was investigated for the first time in the present paper. To on-line interface solid-phase extraction with HPLC, a conventional sample loop on the six-port injector valve of the HPLC was replaced by a preconcentration column packed with carbon nanotubes. The analytes in water solution were preconcentrated onto the preconcentration column and subsequently eluted with mobile phase of methanol-water (22:78). The developed on-line solid-phase extraction method for HPLC permitted the current HPLC separation and the next preconcentration proceeded in parallel, and thus allows one determination finished within 35 min. The RSD of 10 SAs for nine replicate measurements of a standard mixture of 1 microgl(-1) were in the range of 2.5-7.8%. The method was applied to the determination of trace sulfadiazin (SDZ), sulfamerazine (SMR), sulfadimidine (SDMD), sulfathiazole (STZ), sulfamoxol (SMO), sulfamethizole (SMT), sulfamethoxypyridazine (SMP), sulfachlorpyridazine (SCP), sulfadoxin (SDX) and sulfisoxazole (SIA) in eggs and pork. The results indicated that the proposed method was simple, cost-effective and sensitive.     

Molecular dynamics simulation of liquid methanol. II. Unified assignment of infrared, Raman, and sum frequency generation vibrational spectra in methyl C-H stretching region

Vibrational spectra of methyl C-H stretching region are notoriously complicated, and thus a theoretical method of systematic assignment is strongly called for in condensed phase. Here we develop a unified analysis method of the vibrational spectra, such as infrared (IR), polarized and depolarized Raman, and ssp polarized sum frequency generation (SFG), by flexible and polarizable molecular dynamics simulation. The molecular model for methanol has been developed by charge response kernel model to allow for analyzing the methyl C-H stretching vibrations. The complicated spectral structure by the Fermi resonance has been unraveled by empirically shifting potential parameters, which provides clear information on the coupling mechanism. The analysis confirmed that for the IR, polarized Raman, and SFG spectra, two-band structure at about 2830 and 2950 cm(-1) results from the Fermi resonance splitting of the methyl C-H symmetric stretching and bending overtones. In the IR spectrum, the latter, higher-frequency band is overlapped with prominent asymmetric C-H stretching bands. In the depolarized Raman spectrum, the high frequency band at about 2980 cm(-1) is assigned to the asymmetric C-H stretching mode. In the SFG spectrum, the two bands of the splitted symmetric C-H stretching mode have negative amplitudes of imaginary nonlinear susceptibility χ(2), while the higher-frequency band is partly cancelled by positive imaginary components of asymmetric C-H stretching modes.     

Quaternary ammonium chitosan derivative dynamic coating for the separation of veterinary sulfonamide residues by CE with field-amplified sample injection

A quaternary ammonium chitosan, 2-hydroxypropyltrimethylammonium chloride chitosan (HACC), has been developed for the dynamic coating material in CE for the first time. It presented many advantages such as favorable water solubility, satisfactory coating efficiency, and EOF toward the anode at pH >7.0. Using the modified fused-silica capillary, sulfonamides (SAs), an important group of veterinary drugs, were separated and detected by CE combined with field-amplified sample injection (FASI). The LODs of sulfonamides with UV detection were less than 0.5 ng/mL. The proposed method has been applied to the determination of veterinary sulfonamide residues in samples such as chicken, beef, and honey with fast separation (15 sulfonamides within 20 min), low LODs (0.1-0.5 ng/mL), and good reliability compared to the criteria of China (GB/T 18932.17-2003).     

Single-crystalline Bi(5)O(7)NO(3) nanofibers: Hydrothermal synthesis, characterization, growth mechanism, and photocatalytic properties

Semifluorinated self-assembled (FAS SA) films fabricated from trifunctional precursors are frequently used in myriad applications, yet an understanding of the effects of fabrication conditions, including deposition time, on adsorption mechanisms and molecular architectures is still being developed. In this work we prepared SA films based on the F(CF(2))(8)(CH(2))(2)SiCl(3) (FAS-17) precursor and characterized these films using a suite of surface analytical techniques. Contact angle, sum frequency generation (SFG) spectroscopy, X-ray photoelectron spectroscopy (XPS), and ellipsometry results are consistent with the formation of disordered sub-monolayer structures at short deposition times, well-ordered monolayers at intermediate deposition times, and inhomogeneous multilayers at long deposition times. Correlation of SFG and XPS results demonstrates a change in FAS-17 chain orientation as the deposition time increases from 2 s to 5 min. Group theory-based calculations, SFG studies, and Fourier-transform infrared (FTIR) results also afford additional evidence in support of the assignment of the SFG signals at ~1345 and ~1370 cm(-1) to the asymmetric stretching mode of the semifluorinated silane chain's terminal CF(3) group rather than to its axial CF(2) stretches. To our knowledge, this is the first report of SFG studies on semifluoroalkyl silane self-assembled films in the C-F stretching frequency region.     

固相萃取-高效液相色谱法同时测定鸡粪中四环素类、喹诺酮类和磺胺类抗生素

建立了一种高效、低成本的固相萃取-高效液相色谱（SPE-HPLC）同时测定鸡粪中6种常见抗生素（2种四环素类、2种喹诺酮类和2种磺胺类）的分析方法。样品经EDTA-McIlvaine缓冲液和有机混合提取液（甲醇-乙腈-丙酮，2：2：1，v/v/v）提取，过HLB固相萃取柱净化，甲醇-二氯甲烷（7：3，v/v）洗脱，高效液相色谱-二极管阵列检测器测定，检测波长λ=270 nm，柱温32℃，流动相为乙腈-0.7%（v/v）磷酸水溶液。该方法在0.5~100 mg/L质量浓度范围内的标准曲线相关系数r²在0.9999~1之间，样品加标回收率在70.0%~116.3%之间，相对标准偏差为1.2%~16.6%。方法检出限为1.3~6.7 μg/kg，定量限为3.5~9.2 μg/kg。应用该方法对辽宁省抚顺市某养鸡场当天的鸡粪进行检测，诺氟沙星和恩诺沙星（喹诺酮类）含量为未检出~9.23 mg/kg和1.57~7.69 mg/kg，磺胺二甲嘧啶（磺胺类）含量为2.02~13.05 mg/kg，磺胺甲恶唑、土霉素和四环素未测出。     

Communication: Spectroscopic phase and lineshapes in high-resolution broadband sum frequency vibrational spectroscopy: resolving interfacial inhomogeneities of "identical" molecular groups

The ability to achieve sub-wavenumber resolution (0.6 cm(-1)) and a large signal-to-noise ratio in high-resolution broadband sum-frequency generation vibrational spectroscopy (HR-BB-SFG-VS) allows for the detailed SFG spectral lineshapes to be used in the unambiguous determination of fine spectral features. Changes in the structural spectroscopic phase in SFG-VS as a function of beam polarization and experimental geometry proved to be instrumental in the identification of an unexpected 2.78 ± 0.07 cm(-1) spectral splitting for the two methyl groups at the vapor/dimethyl sulfoxide (DMSO, (CH(3))(2)SO) liquid interface as well as in the determination of their orientational angles.     

Surface characterization of 1-butyl-3-methylimidazolium Br-, I-, PF6-, BF4-, (CF3SO2)2N-, SCN-, CH3SO3-, CH3SO4-, and (CN)2N- ionic liquids by sum frequency generation

Sum frequency generation spectroscopy, SFG, was used for the surface characterization at the gas-liquid interface of the 1-butyl-3-methylimidazolium cation combined with the following anions: Br-, I-, PF6-, BF4-, (CF3SO2)2N- (imide), SCN-, CH3SO3- (MeSO3), CH3SO4- (MS), and (CN)2N- (DCN). The SFG spectra obtained for the different ionic liquids were similar independent of the anion selected; therefore, a comprehensive analysis for the surface characterization of the ionic liquids' cation was focused only on the PF6- and Br- anion combinations. For an accurate identification of the vibrational modes observed, FT-IR and Raman spectroscopy in combination with isotopic labeling with deuterium and polarized Raman spectroscopy was used. The cation orientation was determined by analysis of polarization-dependent SFG spectra. For a compound dried in a vacuum to < or = 2 x 10(-5) Torr, the cation appears to be oriented with the ring laying flat along the surface plane and the butyl chain projecting into the gas phase independent of the anion identity.     

Liquid chromatography–fluorescence detection for simultaneous analysis of sulfonamide residues in honey

A rapid, accurate LC analytical method has been developed for determination of eight sulfonamides (sulfacetamide, sulfapyridine, sulfamerazine, sulfamethoxypyridazine, sulfameter, sulfachloropyridazine, sulfamethoxazole and sulfadimethoxine) in honey. The sample was dissolved in phosphoric acid solution (pH 2). After filtration, the sample solution was cleaned by use of two solid-phase extraction (SPE) cartridges-an aromatic sulfonic cation-exchange cartridge and an Oasis HLB cartridge. The eight sulfonamides were then derivatized with fluorescamine and the derivatives were determined by LC with fluorescence detection at excitation and emission wavelengths of 405 and 495 nm, respectively. Average recoveries at three fortification levels in the range 0.02-0.50 mg kg(-1) in twelve different kinds of honey were 73.5-94.1% with coefficients of variation of 4.35-16.60%. The limit of detection (LOD) was 0.002 mg kg(-1) for sulfacetamide, sulfapyridine, sulfamerazine, and sulfamethoxypyridazine; that for sulfameter, sulfachloropyridazine, sulfamethoxazole and sulfadimethoxine was 0.005 mg kg(-1). The limit of quantitation (LOQ) was 0.005 mg kg(-1) for sulfacetamide, sulfapyridine, sulfamerazine, and sulfamethoxypyridazine; that for sulfameter, sulfachloropyridazine, sulfamethoxazole, and sulfadimethoxine was 0.010 mg kg(-1). The method is suitable for determination of multiresidue sulfonamides in the various kinds of honey.     

高效液相色谱法同时测定化妆品中的7种磺胺及甲硝唑和氯霉素

建立了化妆品中7种磺胺(磺胺醋酰、磺胺吡啶、磺胺甲基嘧啶、磺胺二甲嘧啶、磺胺甲氧嘧啶、磺胺间甲氧嘧啶、磺胺甲基异唑)和甲硝唑及氯霉素的高效液相色谱测定方法。样品经0.1%甲酸水溶液-乙腈(体积比为8∶2)混合液超声提取后进行液相色谱分析。方法的定量检测限为3～80 μg/g,7种磺胺在20～200 μg/mL时,甲硝唑及氯霉素在40～400 μg/mL时方法的线性关系良好(r≥0.9993)。加标回收率为83.8%～105.3%(7种磺胺的添加水平为50 μg/mL和150 μg/mL,甲硝唑及氯霉素的添加水平为100 μg/mL和300 μg/mL),其相对标准偏差均小于5%。     

固相萃取-高效液相色谱法同时测定牛奶中22种磺胺类兽药残留

建立了一种基于固相萃取技术同时测定牛奶中22种磺胺类兽药残留的高效液相色谱分析方法.样品经乙腈-甲酸混合溶液提取,乙腈饱和的正己烷除酯后,HLB固相萃取柱净化,以甲醇-2 mmol/L乙酸铵(含0.2％乙酸)为流动相进行梯度洗脱,XBridge C18色谱柱进行分离,采用光电二极管阵列检测器检测,外标法定量.磺胺类化合...     

A simple spectrophotometric method for the determination of copper in industrial, environmental, biological and soil samples using 2,5-dimercapto-1,3,4-thiadiazole.

A simple spectrophotometric method is presented for the rapid determination of copper at a trace level using 2,5-dimercapto-1,3,4-thiadiazole (DMTD) as a new spectrophotometric reagent. The method is based on the reaction of non-absorbent DMTD in a slightly acidic (0.002-0.014 mol dm(-3) sulfuric acid) aqueous solution with copper(II) to produce a highly absorbent greenish-yellow chelate product that has an absorption maximum at 390 nm. The reaction is instantaneous and the absorbance remains stable for 24 h. The average molar absorption coefficient and Sandell's sensitivity were found to be 5.65 x 10(4) dm3 mol(-1) cm(-1) and 10 ng cm(-2) of CuII, respectively. Linear calibration graphs were obtained for 0.1-20 microg cm(-3) of CuII; the stoichiometric composition of the chelate is 1:2 (Cu:DMTD). A large excess of over 50 cations, anions and complexing agents (e.g. tartrate, oxalate, citrate, phosphate, thiourea, SCN-) do not interfere in the determination. The method was successfully used for the determination of copper in several Standard Reference Materials as well as in some environmental water samples, biological samples, soil samples and solutions containing both copper(I) and copper(II) and complex synthetic mixtures. The method has high precision and accuracy (s = +/-0.01 for 0.5 microg cm(-1)).     

Trace determination of sulfonamides residues in meat with a combination of solid-phase microextraction and liquid chromatography-mass spectrometry

An integrated method of combining solid-phase microextraction (SPME) with liquid chromatography-mass spectrometry (LC-MS) was evaluated for determination trace amount of sulfonamides in meat products. Eight commonly used sulfonamides, sulfadiazine (SDZ), sulfathiazole (STZ), sulfamerazine (SMR), sulfamethazine (SMT), sulfamonomethoxine (SMMX), sulfamethoxazole (SMXZ), sulfaquinoxaline (SQX) and sulfadimethoxine (SDMX), were investigated in this study. Chromatography was performed on a C₁₈ reversed-phase column using an isocratic acetonitrile in water as the mobile phase. Fiber coated with a 65 μm thickness of polydimethylsiloxane/divinylbenzene (PDMS/DVB) was used to extract sulfonamides at optimum conditions. Analytes were desorbed with static desorption in an SPME-HPLC desorbed chamber for 15 min and then determined by LC-MS. The detection limits of these sulfonamides in pork were from 16 μg kg⁻¹ (SMT) to 39 μg kg⁻¹ (SMMX). According to the analysis, the linear range was from 50 to 2000 μg kg⁻¹ with relative standard deviation (R.S.D.s) value below 15% (intra-day) and 19% (inter-day). The proposed method was tested by analyzing meats from a local market for sulfonamides residues. Some sulfonamides in our study were detected in the meat samples. The concentration of these residual sulfonamides ranged from 66 μg kg⁻¹ (SDZ) to 157 μg kg⁻¹ (SQX) in a chicken sample. The results demonstrate that the SPME-LC-MS system is highly effective in analyzing trace sulfonamides in meat products.     

Heterogeneous ozone oxidation reactions of 1-pentene, cyclopentene, cyclohexene, and a menthenol derivative studied by sum frequency generation

We report vibrational sum frequency generation (SFG) spectra of glass surfaces functionalized with 1-pentene, 2-hexene, cyclopentene, cyclohexene, and a menthenol derivative. The heterogeneous reactions of ozone with hydrocarbons covalently linked to oxide surfaces serve as models for studying heterogeneous oxidation of biogenic terpenes adsorbed to mineral aerosol surfaces commonly found in the troposphere. Vibrational SFG is also used to track the C=C double bond oxidation reactions initiated by ozone in real time and to characterize the surface-bound product species. Combined with contact angle measurements carried out before and after ozonolysis, the kinetic and spectroscopic studies presented here suggest reaction pathways involving vibrationally hot Criegee intermediates that compete with pathways that involve thermalized surface species. Kinetic measurements suggest that the rate limiting step in the heterogeneous C=C double bond oxidation reactions is likely to be the formation of the primary ozonide. From the determination of the reactive uptake coefficients, we find that ozone molecules undergo between 100 and 10000 unsuccessful collisions with C=C double bonds before the reaction occurs. The magnitude of the reactive uptake coefficients for the cyclic and linear olefins studied here does not follow the corresponding gas-phase reactivities but rather correlates with the accessibility of the C=C double bonds at the surface.     

Determination of Sulfonamides in Pharmaceuticals and Rabbit Plasma by Microchip Electrophoresis with LED-IF Detection

In this paper, we describe a compact and low-cost light-emitting diode-induced fluorescence (LED-IF) detection coupled to microchip electrophoresis for the determination of sulfonamides in pharmaceutical formulations and rabbit plasma. Three fluorescein isothiocyanate-labeled sulfonamides in rabbit plasma were separated in the running buffer of 40 mM phosphate buffer (pH 7.0) at the separation voltage of 2.0 kV, and detected by LED-IF detector in which the high-power blue LED was driven at the constant current of 150 mA and the emitted fluorescence over 510 nm was collected by a planar photodiode. The linear concentration ranged from 2.0 to 125.0 μg mL^?1, both for sulfadiazine and sulfamethazine with the correlation coefficients (r ²) of 0.995 and 0.997, respectively, and from 2.0 to 100.0 μg mL^?1 with the correlation coefficients (r ²) of 0.997 for sulfaguanidine. The limits of detection for the three sulfonamides were 0.36–0.50 μg mL^?1 (S/N = 3). Intra-day and inter-day precision of migration time and peak area for the determination of sulfonamides were <4.5 %. This method has been successfully applied to the analysis of sulfonamides in pharmaceuticals, and could be used to study the pharmacokinetics of sulfonamides in rabbit.     

Sanglifehrin-cyclophilin interaction: degradation work,synthetic macrocyclic analogues,X-ray crystal structure,and binding data

Sanglifehrin A (SFA) is a novel immunosuppressive natural product isolated from Streptomyces sp. A92-308110. SFA has a very strong affinity for cyclophilin A (IC(50) = 6.9 +/- 0.9 nM) but is structurally different from cyclosporin A (CsA) and exerts its immunosuppressive activity via a novel mechanism. SFA has a complex molecular structure consisting of a 22-membered macrocycle, bearing in position 23 a nine-carbon tether terminated by a highly substituted spirobicyclic moiety. Selective oxidative cleavage of the C(26)=C(27) exocyclic double bond affords the spirolactam containing fragment 1 and macrolide 2. The affinity of 2 for cyclophilin (IC(50) = 29 +/- 2.1 nM) is essentially identical to SFA, which indicates that the interaction between SFA and cyclophilin A is mediated exclusively by the macrocyclic portion of the molecule. This observation was confirmed by the X-ray crystal structure resolved at 2.1 A of cyclophilin A complexed to macrolide 16, a close analogue of 2. The X-ray crystal structure showed that macrolide 16 binds to the same deep hydrophobic pocket of cyclophilin A as CsA. Additional valuable details of the structure-activity relationship were obtained by two different chemical approaches: (1) degradation work on macrolide 2 or (2) synthesis of a library of macrolide analogues using the ring-closing metathesis reaction as the key step. Altogether, it appears that the complex macrocyclic fragment of SFA is a highly optimized combination of multiple functionalities including an (E,E)-diene, a short polypropionate fragment, and an unusual tripeptide unit, which together provide an extremely strong affinity for cyclophilin A.     

Confirmation of sulfamethazine, sulfathiazole, and sulfadimethoxine residues in condensed milk and soft-cheese products by liquid chromatography/tandem mass spectrometry

A liquid chromatography/tandem mass spectrometry method (LC/MS/MS) is described for the simultaneous detection of 3 sulfonamide drug residues at 1.25 ppb in condensed milk and soft-cheese products. The 3 sulfonamide drugs of interest are sulfathiazole (STZ), sulfamethazine (SMZ), and sulfadimethoxine (SDM). The method includes extraction of the product with phosphate buffer, centrifugation of the diluted product, and application of a portion of the extract onto a polymeric solid-phase extraction cartridge. The cartridge is washed with water, and the sulfonamides are eluted with methanol. After evaporation, the residue is dissolved in 0.1% formic acid solution, and the solution is filtered before analysis by LC/MS/MS. The LC/MS/MS program involved a series of time-scheduled selected-reaction monitoring transitions. The transitions of MH+ to the common product ions at m/z 156, 108, and 92 were monitored for each residue. In addition, SMZ and SDM had a fourth significant and unique product ion transition that could be measured. Validation was performed with control and fortified-control condensed bovine milk with 2.5, 5, and 10 ppb sulfonamides. This method was applied to imported flavored and unflavored condensed milk and cream cheese bars. The presence of STZ and SMZ residues was confirmed in 3 out of 6 products.     

高效液相色谱-柱后衍生法检测养殖水体及沉积物中11种磺胺药物残留

建立了养殖水体及沉积物中11种磺胺化合物的高效液相色谱-柱后衍生分析方法。养殖水体过滤后采用HLB固相萃取柱进行净化、富集;沉积物采用甲醇/EDTA-Mcllvaine缓冲液(1:1, v/v)提取,HLB固相萃取柱净化富集。经高效液相色谱分离,用荧光胺衍生试剂进行柱后衍生,荧光检测器检测。对柱后衍生系统参数进行了优化,确定了荧光胺溶液的浓度、流速和反应温度分别为0.2 g/L、0.15 mL/min和50 ℃,磺胺化合物在0.01~1.0 mg/L范围内线性显著,其相关系数r²值大于0.99995。11种磺胺类药物在养殖水体和沉积物中的加标回收率分别为79.3%~100.7%和74.6%~95.3%,相对标准偏差为2.2%~11.0%和2.6%~10.3%,检出限(LOD, S/N=3)为0.9~5.5 ng/L和0.3~1.3 μg/kg,定量限(LOQ, S/N=10)为3.0~18.1 ng/L和1.0~4.4 μg/kg。该法可应用于养殖环境中磺胺类药物的定性定量检测,具有较好的实用性。     

An environmentally friendly reflectometric method for ranitidine determination in pharmaceuticals and human urine

This paper describes an environmentally friendly method for quantitative determination of ranitidine using diffuse reflectance spectroscopy. This method is based on the reflectance measurements of the colored product produced from the spot test reaction between ranitidine and p-dimethylaminocinnamaldehyde (p-DAC), in acid medium, using filter paper as solid support. Experimental design methodologies were used to optimize the optimal conditions. All reflectance measurements were carried out at 590 nm and the linear range was from 1.42x10(-3) to 3.42x10(-2) mol L(-1), with a correlation coefficient of 0.997. The limit of detection was estimated to be 1.09x10(-3) mol L(-1) (R.S.D.=1.9%). The proposed method was successfully applied to the determination of ranitidine in commercial brands of pharmaceuticals and no interferences were observed from the common excipients in formulations. The results obtained by the proposed method were favorably compared with those obtained by an official procedure at 95% confidence level. Additionally, the method was also applied to the determination of ranitidine in human urine showing excellent recoveries (99.6-100.3%).     

Stability-indicating methods for determination of tiapride in pure form, pharmaceutical preparation, and human plasma

Four different stability-indicating procedures are described for determination of tiapride in pure form, dosage form, and human plasma. Second derivative (D2), first derivative of ratio spectra (1DD), spectrofluorimetric, and high-performance column liquid chromatographic (LC) methods are proposed for determination of tiapride in presence of its acid-induced degradation products, namely 2-methoxy-5-(methylsulfonyl) benzoic acid and 2-diethylaminoethylamine. These approaches were successfully applied to quantify tiapride using the information included in the absorption, excitation, and emission spectra of the appropriate solutions. In the D2 method, Beer's law was obeyed in the concentration range of 1.5-9 microg/mL with a mean recovery of 99.94 +/- 1.38% at 253.4 nm using absolute ethanol as a solvent. In 1DD, which is based on the simultaneous use of the first derivative of ratio spectra and measurement at 245 nm in absolute ethanolic solution, Beer's law was obeyed over a concentration range of 1.5-9 microg/mL with mean recovery 99.64 +/- 1.08%. The spectrofluorimetric method is based on the determination of tiapride native fluorescence at 339 nm emission wavelength and 230 nm excitation wavelength using water-methanol (8 + 2, v/v). The calibration curve was linear over the range of 0.2-3 microg/mL with mean recovery of 99.66 +/- 1.46%. This method was also applied for determination of tiapride in human plasma. A reversed-phase LC method performed at ambient temperature was validated for determination of tiapride using methanol-deionized water-triethylamine (107 + 93 + 0.16, v/v/v) as the mobile phase. Sulpiride was used as an internal standard at a flow rate of 1 mL/min with ultraviolet detection at 214 nm. A linear relation was obtained over a concentration range of 2-30 microg/mL with mean recovery of 99.66 +/- 0.9%. Results were statistically analyzed and compared with those obtained by applying the reference method. They proved both accuracy and precision.