Simultaneous determination of diethylene glycol and propylene glycol in pharmaceutical products by HPLC after precolumn derivatization with p-toluenesulfonyl isocyanate

A simple and reliable HPLC method was developed for the simultaneous quantitative analysis of diethylene glycol (DEG) and propylene glycol (PG) in pharmaceutical products by precolumn derivatization. The derivatization reagent p-toluenesulfonyl isocyanate (TSIC, 10 microL, 20% in ACN v/v) was added to 100 microL of the sample, and then 10 muL of water was added. The resulting derivatives were separated using a C(18)analytical column and a mobile phase composed of 0.01 M KH(2)PO(4)buffer (adjusted to pH 2.5 with phosphoric acid) and ACN (47:53 v/v) at 1 mL/min and 25 degrees C. For detection, UV light at 227 nm was used. The derivatization conditions including reaction time, temperature, and concentration of TSIC were optimized. The calibration curves were linear from 0.062 to 18.6 microg/mL (r(2)= 0.9999) and from 0.071 to 21.3 microg/mL (r(2) = 0.9999) for DEG and PG, respectively. The RSD values of intra- and interday assays were all below 4% for DEG and PG. The proposed method was then successfully applied to analyze two Armillarisin A injection samples and two spiked syrup samples.     

2,6-Dimethyl-4-quinolinecarboxylic acid N-hydroxysuccinimide ester: a fluorogenic hydrophilic derivatizing reagent for liquid chromatographic analysis of aliphatic amines

The use of a fluorogenic, hydrophilic, and amine-reactive reagent, 2,6-dimethyl-4-quinolinecarboxylic acid N-hydroxysuccinimide ester (DMQC-OSu) has been investigated in the procolumn derivatization for the LC separation of aliphatic amines. In pH 8.0 aqueous medium, DMQC-OSu reacted with amines at 50 degrees C within 20 min to form highly fluorescent carboxamides and the excess reagent hydrolyzed to the corresponding carboxylic acid. The separation of representative amine derivatives with DMQC-OSu has been performed using a C18 column with the fluorescence detection at 326/409 nm. The detection limits reached nanomolar level. The proposed method has been applied to the analysis of real samples with recoveries of 94-108%. Compared with other succinimidyl esters used in the derivatization of amino compounds, DMQC-OSu and its hydrolysate had negligible fluorescence (phi(fl) = 0.09 and 0.02, respectively), which implied that small peaks appeared in chromatograms and slight interference was introduced to the determination.     

3-Iodoacetylaminobenzanthrone as a fluorescent derivatizing reagent for thiols in high-performance liquid chromatography

A new thiol-reactive derivatizing reagent, 3-iodoacetylaminobenzanthrone (IAB) has been developed for thiol analysis in liquid chromatography. In aqueous methanol containing 15 mM pH 8.3 H₃BO₃-KCl-Na₂CO₃ buffer, IAB reacted with thiols at 35 °C for 15 min. The derivatives of IAB with glutathione (GSH), cysteine (Cys), homocysteine (Hcy) and N-acetylcysteine (Nac) were well separated on a C₁₈ column with the mobile phase of methanol-water (50:50, v/v) containing 15 mM pH 2.7 H₃cit-Na₂HPO₄ buffer. At λ_ex/λ_em=420/540 nm, the detection limits were 20, 20, 55 and 40 fmol (1, 1, 2.3 and 2 nM), respectively, with a signal-to-noise ratio of 3. Owing to the preferential selectivity of iodoacetamidyl moiety to SH group, amino acids, aliphatic amines, phenol and alcohols had no obvious interference with the determination. The proposed method has been applied to the determination of thiols in human blood with recoveries of 98.5-105.3%.     

Analysis of primary aromatic amines using precolumn derivatization by HPLC fluorescence detection and online MS identification

2-(2-phenyl-1H-phenanthro-[9,10-d]imidazole-1-yl)-acetic acid (PPIA) and 2-(9-acridone)-acetic acid (AAA), two novel precolumn fluorescent derivatization reagents, have been developed and compared for analysis of primary aromatic amines by high performance liquid chromatographic fluorescence detection coupled with online mass spectrometric identification. PPIA and AAA react rapidly and smoothly with the aromatic amines on the basis of a condensation reaction using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) as dehydrating catalyst to form stable derivatives with emission wavelengths at 380 and 440 nm, respectively. Taking six primary aromatic amines (aniline, 2-methylaniline, 2-methoxyaniline, 4-methylaniline, 4-chloroaniline, and 4-bromoaniline) as testing compounds, derivatization conditions such as coupling reagent, basic catalyst, reaction temperature and time, reaction solvent, and fluorescent labeling reagent concentration have also been investigated. With the better PPIA method, chromatographic separation of derivatized aromatic amines exhibited a good baseline resolution on an RP column. At the same time, by online mass spectrometric identification with atmospheric pressure chemical ionization (APCI) source in positive ion mode, the PPIA-labeled derivatives were characterized by easy-to-interpret mass spectra due to the prominent protonated molecular ion m/z [M + H](+) and specific fragment ions (MS/MS) m/z 335 and 295. The linear range is 24.41 fmol-200.0 pmol with correlation coefficients in the range of 0.9996-0.9999, and detection limits of PPIA-labeled aromatic amines are 0.12-0.21 nmol/L (S/N = 3). Method repeatability, precision, and recovery were evaluated and the results were excellent for the efficient HPLC analysis. The most important argument, however, was the high sensitivity and ease-of-handling of the PPIA method. Preliminary experiments with wastewater samples collected from the waterspout of a paper mill and its nearby soil where pollution with aromatic amines may be expected show that the method is highly validated with little interference in the chromatogram.     

Determination of primary biogenic amines in various food products using isocratic HPLC with 3-(4-chlorobenzoyl)-quinoline-2-carboxaldehyde as a new precolumn fluorogenic reagent

A simple HPLC approach has been successfully established for the sensitive determination of six biogenic amines (BAs) in food samples. The method involves derivatization with 3-(4-chlorobenzoyl)-quinoline-2-carboxaldehyde newly synthesized as a new fluorogenic reagent followed by LC isocratic elution mode. The optimization of both derivatization and separation conditions is carefully studied. Related analyses of the eluted compounds, in the presence of MeOH/THF/H(2)O (78:2.5:19.5 v/v/v) as a mobile phase containing 8 mM, pH 6.0 phosphate buffer solution, have been carried out on a C(18) column. The LOD (S/N = 3) of 2.5 nM, RSD value from 1.0 to 5.1% in peak areas, and good response linearity (R(2) >0.9936) are provided with fluorescence detection at lambda(ex)/lambda(em) = 480/545 nm. Obviously, recovery ranging from 95 to 107% in this method demonstrates its accuracy for determination of histamine, tyramine, 2-phenylethylamine, putrescine, cadaverine, and spermidine in the storage fish sample. Thus, the present method could be developed to monitor BAs in fish, cucumber, and spinach samples.     

柱前衍生高效液相色谱法测定啤酒中微量甲醛

甲醛作为消毒剂、工业助剂等已广泛应用于各行各业,也列在食品工业加工助剂推荐名单中[1]。然而,甲醛能引起头痛、恶心,肠胃不舒服、皮肤过敏等反应。美国环保署(EPA)称甲醛可能是致癌诱导物[2]。我国居室内空气中甲醛卫生标准不大于0·08mg/m3,而饮用水和饮料类中甲醛允许含量     

A means of analysing glycols,especially ethylene glycol and diethylene glycol,by a method used for the determination of carbohydrates in alcoholic beverages

Summary This paper describes the separation of carbohydrates/glycerol/ethylene glycol/diethylene glycol in alcoholic liquids by TLC. An in situ reaction already proven in carbohydrate analysis is used for the detection among other things, of diethylene glycol in low, unpermitted concentrations and is compared with vanadium pentoxide derivatisation also described in the literature. Only the vanadium pentoxide reaction yields satisfactory results for diethylene glycol. The information capacity of HPTLC chromatograms is also demonstrated by the simultaneous analysis of 19 samples. Chromatogram filing is an important consideration, since the lifespans of chromatograms derivatised by the two methods vary considerably. Plates or sheets treated with dichlorofluorescein can still be evaluated over a period of years, whereas plates treated with vanadium pentoxide are unusable after only a few weeks. An advantage of the latter procedure, however, is the amenability of the chromatograms to normal photocopying.     

Indirect enantioseparation of proteinogenic amino acids using naproxen‐based chiral derivatizing reagent and HPLC

Diastereomers of 18 proteinogenic amino acids were synthesized under microwave irradiation and by vortexing using (S)‐naproxen–benzotriazole as chiral derivatizing reagent. The diastereomers synthesized by two approaches were found to be identical in terms of their characterization and chromatographic data. A linear gradient of triethylammonium phosphate (pH 3.5)–acetonitrile (30–65%, within 35 min) was found to be successful using reversed‐phase high‐performance liquid chromatography for their separation. Detection was carried out at 231 nm and sharp peaks were obtained. The method was validated for accuracy, precision and limit of detection. Copyright © 2012 John Wiley & Sons, Ltd.     

高效液相色谱-荧光检测法同时测定水果中的3种链格孢霉毒素

建立了同时检测水果中链格孢霉毒素、链格孢酚和链格孢酚甲醚残留量的高效液相色谱-荧光分析方法。样品经乙腈提取,HLB和氨基固相萃取小柱净化,采用Chromolith Performance RP-18e整体柱分离,荧光检测器检测,以外标法进行定量分析。当3种链格孢霉毒素添加水平为20,40,100μg/kg时,方法平均回收率均在78.2%～103.6%范围内,相对标准偏差小于8.6%。方法适用于水果中链格孢霉毒素残留量的测定。     

Hollow-fiber liquid-phase microextraction for the determination of pesticides and metabolites in soils and water samples using HPLC and fluorescence detection

A new and simple method has been developed for the determination of a group of four benzimidazole pesticides (carbendazim/benomyl, thiabendazole, and fuberidazole), a carbamate (carbaryl), and an organophosphate (triazophos), together with two of their main metabolites (2-aminobenzimidazole, metabolite of carbendazim/benomyl, and 1-naphthol, metabolite of carbaryl) in soils. First, an ultrasound-assisted extraction (UAE) was performed, followed by evaporation and reconstitution in water. Then, extraction and preconcentration of the analytes was accomplished by two-phase hollow-fiber liquid-phase microextraction (HF-LPME) using 1-octanol as extraction solvent. Parameters that affect the extraction efficiency in HF-LPME technique (organic solvent, pH of the sample, extraction time, stirring speed, temperature, and ionic strength) were deeply investigated. Optimum HF-LPME conditions involved the use of a 2.0 cm polypropylene fiber filled with 1-octanol to extract 10 mL of an aqueous soil extract at pH 9.0 containing 20% (v/v) of NaCl for 30 min at 1440 rpm. Separation and quantification was achieved by HPLC with fluorescence detection (FD). The proposed optimum UAE-HF-LPME-HPLC-FD methodology provided good calibration, precision, and accuracy results for two soils of different physicochemical properties. LODs were in the range 0.001-6.94 ng/g (S/N = 3). With the aim of extending the validation, the HF-LPME method was also applied to different types of waters (Milli-Q, mineral and run-off), obtaining LODs in the range 0.0002-0.57 μg/L.     

Study of a new derivatizing reagent that improves the analysis of amino acids by HPLC with fluorescence detection: application to hydrolyzed rape bee pollen

A simple and sensitive method for evaluating the chemical compositions of protein amino acids, including cystine (Cys)₂ and tryptophane (Try) has been developed, based on the use of a sensitive labeling reagent 2-(11H-benzo[α]-carbazol-11-yl) ethyl chloroformate (BCEC–Cl) along with fluorescence detection. The chromophore of the 1,2-benzo-3,4-dihydrocarbazole-ethyl chloroformate (BCEOC-Cl) molecule was replaced with the 2-(11H-benzo[α]-carbazol-11-yl) ethyl functional group, yielding the sensitive fluorescence molecule BCEC–Cl. The new reagent BCEC–Cl could then be substituted for labeling reagents commonly used in amino acid derivatization. The BCEC–amino acid derivatives exhibited very high detection sensitivities, particularly in the cases of (Cys)₂ and Try, which cannot be determined using traditional labeling reagents such as 9-fluorenyl methylchloroformate (FMOC-Cl) and ortho-phthaldialdehyde (OPA). The fluorescence detection intensities for the BCEC derivatives were compared to those obtained when using FMOC-Cl and BCEOC-Cl as labeling reagents. The ratios I _BCEC/I _BCEOC = 1.17–3.57, I _BCEC/I _FMOC = 1.13–8.21, and UV_BCEC/UV_BCEOC = 1.67–4.90 (where I is the fluorescence intensity and UV is the ultraviolet absorbance). Derivative separation was optimized on a Hypersil BDS C₁₈ column. The detection limits calculated from 1.0 pmol injections, at a signal-to-noise ratio of 3, ranged from 7.2 fmol for Try to 8.4 fmol for (Cys)₂. Excellent linear responses were observed, with coefficients of >0.9994. When coupled with high-performance liquid chromatography, the method established here allowed the development of a highly sensitive and specific method for the quantitative analysis of trace levels of amino acids including (Cys)₂ and Try from bee-collected pollen (bee pollen) samples.     

High‐performance liquid chromatography with fluorescence detection for the rapid analysis of pheophytins and pyropheophytins in virgin olive oil

Pheophytins and pyropheophytin are degradation products of chlorophyll pigments, and their ratios can be used as a sensitive indicator of stress during the manufacturing and storage of olive oil. They increase over time depending on the storage condition and if the oil is exposed to heat treatments during the refining process. The traditional analysis method includes solvent‐ and time‐consuming steps of solid‐phase extraction followed by analysis by high‐performance liquid chromatography with ultraviolet detection. We developed an improved dilute/fluorescence method where multi‐step sample preparation was replaced by a simple isopropanol dilution before the high‐performance liquid chromatography injection. A quaternary solvent gradient method was used to include a fourth strong solvent wash on a quaternary gradient pump, which avoided the need to premix any solvents and greatly reduced the oil residues on the column from previous analysis. This new method not only reduces analysis cost and time but shows reliability, repeatability, and improved sensitivity, especially important for low‐level samples.     

Sensitive determination of aliphatic amines by high-performance liquid chromatography with a new fluorogenic probe 3-(4-fluorinebenzoyl)-2-quinoline carboxaldehyde

A new fluorogenic reagent 3-(4-fluorinebenzoyl)-2-quinoline carboxaldehyde (FBQCA) has been synthesized and used as a derivatizing reagent for the determination of aliphatic amines with HPLC. The reagent is nonfluorescent, but forms highly fluorescent isoindole upon the reaction with primary amines in alkaline medium. Eleven amine derivatives were baseline separated in 8 min using a gradient elution on a C(8) column and detected with fluorescence detection at lambda(ex)/lambda(em) = 480/546 nm. The detection limits were in the range of 0.5-2 nM (S/N = 3). The proposed method has been successfully applied to the analysis of aliphatic amines in food and environmental samples, including white wine, soybean oil, soil, and tap water with satisfactory recoveries in the range of 94-106%.     

Synthesis of nano-TiO2 assisted by diethylene glycol for use in high efficiency dye-sensitized solar cells

The performance of dye-sensitized solar cells(DSSCs) consisting of anatase TiO_2 nanoparticles that were synthesized via a hydrothermal method was studied.The synthesized TiO_2 nanoparticles were characterized by X-ray diffraction(XRD),nitrogen sorption analysis,scanning electron microscopy(SEM),high resolution transmission electron microscopy(HRTEM),and UV-vis spectroscopy.Then the J-Vcurve,electrochemical impedance spectroscopy(EIS),and open-circuit voltage decay(OCVD) measurement were applied to evaluate the photovoltaic performance of DSSCs.Compared with the commercial TiO_2nanoparticles(P25),the synthesized-TiO_2 nanoparticles showed better performance.By adding diethylene glycol(DEG) before the hydrothermal process,the synthesized TiO_2 nanoparticles(hereafter referred to as TiO_2-DEG particles) shows narrower size distribution,larger specific surface area,higher crystallinity,and less surface defects than TiO_2(DEG free) particles.The analysis of photovoltaic properties of DSSCs based on TiO_2-DEG particles showed that the recombination of electron-hole pairs was decreased and the trapping of carries in grain boundaries restrained.It was believed that the photoelectrode fabricated with the as-prepared TiO_2 nanoparticles improved the loading amount of dye sensitizers(N719).and enhanced the photocurrent of the DSSCs.As a result,the TiO_2-DEG particle based cells achieved a photo-to-electricity conversion efficiency(η) of 7.90%,which is higher than 7.53%for the cell based on TiO2(DEG free) and 6.59%for the one fabricated with P25.     

Determination of 1-aminocyclopropane-1-carboxylic acid in apple extracts by capillary electrophoresis with laser-induced fluorescence detection

A rapid and sensitive method for the determination of 1-aminocyclopropane-1-carboxylic acid (ACC) in apple tissues has been described. This method is based on the derivatization of ACC with 3-(2-furoyl)quinoline-2-carboxaldehyde (FQ), and separation and quantification of the resulting FQ-ACC derivative by capillary electrophoresis coupled to laser-induced fluorescence detection (CE-LIF). Our results indicated that ACC derivatized with FQ could be well separated from other interfering amino acids using 20 mM borate buffer (pH 9.35) containing 40 mM sodium dodecyl sulfate and 10 mM Brij 35. The linearity of ACC was determined in the range from 0.05 to 5 microM with a correlation of 0.9967. The concentration detection limit for ACC was 10 nM (signal-to-noise = 3). The sensitivity and selectivity of this described method allows the analysis of ACC in crude apple extracts without extra purification and enrichment procedure.     

Determination of phosphoamino acids by micellar electrokinetic capillary chromatography with laser-induced fluorescence detection

A micellar electrokinetic capillary chromatography (MEKC) method with laser-induced fluorescence detection (LIF) was developed for analyzing three phosphoamino acids including phosphotyrosine (P-Tyr), phosphoserine (P-Ser), and phosphothreonine (P-Thr). 3-(2-Furoyl)quinoline-2-carboxaldehyde (FQ), a fluorogenic dye, was employed for derivatization of these phosphoamino acids. Results indicated that the complete baseline resolution of each phosphoamino acid was obtained within 10 min, using 20 mmol l⁻¹ sodium borate buffer (pH 9.35) containing 20 mmol l⁻¹sodium deoxycholate (SDC) and 10 mmol l⁻¹ Brij35. Other common amino acids, especially Glu and Asp, did not disturb the assay of these phosphoamino acids. There was a linear relationship between the peak area for analyte and its concentration, with correlation coefficients in the range of 0.9966-0.9996. The concentration detection limits (signal-to-noise = 3) for P-Tyr, P-Ser, and P-Thr were 10, 40, and 75 nmol l⁻¹, respectively. The developed method was successfully applied for determining phosphoamino acids in the hydrolysis sample of a phosphorylated protein kinase.     

高效液相色谱法测定药品中的乙二醇二乙醚二胺四乙酸

建立了高效液相色谱分析方法用来测定药品中乙二醇二乙醚二胺四乙酸(EGTA)含量,通过检测EGTA与Cu²⁺的配合物EGTA-Cu来检测EGTA。采用Ultimate-AQ C18色谱柱(250 mm×4.6 mm, 5 μm),流动相为乙腈-四丁基氢氧化铵水溶液(质量分数约0.3%四丁基氢氧化铵水溶液,醋酸调pH 6.50)-醋酸钠溶液(35 mmol/L醋酸钠,醋酸调pH 6.50)(20:20:60, v/v/v),检测波长为245 nm,流速为1.50 mL/min,柱温为40 ℃,进样量为100 μL。结果表明,EGTA质量浓度在0.10~15.00 mg/L范围内线性关系良好(R=0.9998);以信噪比(S/N)为3及10确定检出限和定量限,分别为0.05 mg/L和0.17 mg/L;样品加标平均回收率为98.34%~99.03%, RSD为1.08%~3.33%(n=9)。该方法操作简便,具有分离度好、灵敏度高、重复性好、回收率高等特点,适合药品中EGTA含量的检测,为EGTA检测提供了一种有效的检测方法。     

Liquid-liquid-liquid microextraction followed by HPLC with UV detection for quantitation of ephedrine in urine

Liquid-liquid-liquid microextraction (LLLME) in combination with HPLC and UV detection has been used as a sensitive method for the determination of ephedrine in urine samples. Extraction process was performed in a homemade total glass vial without using a Teflon ring, usually employed. Ephedrine was first extracted from 3.5 mL of urine sample (pH 12) into a microfilm of toluene/benzene (50:50). The analyte was subsequently back extracted into an acidic microdrop solution (pH 2) suspended in the organic phase. The extract was then injected into the HPLC system directly. An enrichment factor of 137 along with a good sample clean-up was obtained under the optimized conditions. The calibration curve showed linearity in the range of 0.01-50 mg/L with regression coefficient corresponding to 0.998. The LODs and LOQs, based on a S/N of 3 and 10, were 5 and 10 microg/L, respectively. The method was eventually applied for the determination of ephedrine in urine sample after oral administration of 5 mg single dose of drug.     

Rapid determination of nitrite in foods in acidic conditions by high‐performance liquid chromatography with fluorescence detection

In this study, a simple, rapid, and sensitive method for the determination of nitrite (NO₂^?) in food samples by high‐performance liquid chromatography with fluorescence detection in acidic conditions had been developed. The derivatization of the nitrite with 2,3‐diaminonaphthalene was performed in acidic conditions to yield the highly fluorescent 2,3‐naphthotriazole, which was directly analyzed by high‐performance liquid chromatography with fluorescence detection without adjusting the solution to alkaline. The analysis column was reversed‐phase C₈ column. A constant flow rate of 1.0 mL/min was employed using water/acetonitrile as the mobile phase in isocratic mode (70:30, v/v). Fluorescence was monitored with excitation at 375 nm and emission at 415 nm. The standard calibration curves were linear for nitrite in different matrixes in the concentration range of 0–100 μg/L, and the correlation coefficients ranged from 0.9978 to 0.9998. The limits of detection and quantification were in the ranges of 0.012–0.060 and 0.040–0.20 mg/kg, respectively. The recoveries of nitrite from samples spiked at three different concentrations were 74.0–113.2%, and the relative standard deviations of the recovery results (n = 6) were 1.67–10.8%. The proposed method has good repeatability and is very sensitive and simple. It has been successfully used to determine nitrite in foods.