A Small-Scale Process for Predicting Donnan and Volume Exclusion Effects During Ultrafiltration/Diafiltration Process Development

Achieving the desired final protein formulation using ultrafiltration/diafiltration (UF/DF) operations is an essential component of many protein purification processes. It is well documented that differences in the excipient and buffer concentrations exist between the DF and retentate solutions when they have achieved equilibrium. Several publications have proposed ways to calculate these differences. However, the accuracy of these methods has been limited by the use of an estimated protein charge value. In this article, a small-scale system is described, which can accurately determine the protein charge by making buffer and excipient concentration measurements and applying the determined values to the Donnan and volume exclusion equations. This information can be utilized to generate a standard curve, which in turn can be applied to at-scale UF/DF operations. For 2 different antibodies, the standard curve generated by the small-scale system yielded buffer concentrations and pH values that agreed well with those generated after UF/DF operations, whereas using the theoretical protein charge caused a departure from the measured results. This model also provides good estimates as to how the final formulation pH and buffer concentration vary as a function of the pH and buffer concentration in the DF buffer. This information is of important utility for the accurate formulation of high-concentration protein solutions (>100 mg/mL) where the coconcentration of buffers and the volume exclusion of certain excipients are amplified. The low material requirements of the small-scale system are a major benefit for early phase formulation and process development when sufficient time and material may not be available, in particular to ensure successful UF/DF operations for the development of high protein concentration formulations.     

Determination of oxytocin in a dilute IV solution by LC-MS(n)

The most common drug prescribed to induce labor in the United States is oxytocin, a peptide hormone composed of nine amino acids. Oxytocin is often reconstituted in intravenous (IV) saline solutions at less than 0.05 units ml(-1) (125 ng ml(-1)) to be delivered at 1-4 drops per minute. Existing LC-UV methods for oxytocin do not have sufficient detection limits to quantitate and/or confirm oxytocin in IV solutions without sample concentration. A determinative and confirmatory method for oxytocin was developed using an LC-MS(n) ion trap instrument with an electrospray ionization (ESI) interface in positive ion mode. Separation was achieved on a C-18 column using an isocratic elution of water with 50% acetonitrile (v/v) and water with 0.05% formic acid (v/v) at a flow rate of 250 microl min(-1). Data was acquired from the selected ion monitoring (SIM) of the precursor ion (m/z 1007.3) and MS(2) scans from the collision induced dissociation of m/z 1007.3 at 30% collision energy. In this method, MS(2) full scans were utilized to obtain three structurally significant ions for the unambiguous identification of oxytocin. Calibration standards, prepared in de-ionized water from 0.006 to 0.046 units ml(-1), were linear with an R(2) value of 0.9983. The methods LOD and LOQ were 0.00084 and 0.0029 units ml(-1) (2 and 7 ng ml(-1)), respectively. This LC-MS(n) method was used to determine the amount of oxytocin in a 0.04 units ml(-1) clinical sample that was prepared in 0.9% sodium chloride IV solution.     

Modification of the ultrafiltration technique to overcome solubility and non-specific binding challenges associated with the measurement of plasma protein binding of corticosteroids

Plasma protein binding (PPB) methodology suitable for application in the lead optimisation of a corticosteroid series known to demonstrate non-specific binding (NSB) and poor solubility has been established. The method involved a modification to standard ultrafiltration (UF) techniques. In parallel with each experimental plasma sample, a control plasma sample was also processed by ultrafiltration. The retentate from experimental and control plasma samples were mixed back into the filtrate of the partner sample. The resulting regenerated plasma samples, one representing the experimental filtrate and one representing the experimental retentate, were then analysed by LC/MS/MS. Varying degrees of NSB were demonstrated with a number of corticosteroids, and this effect was eliminated using the modified method. Validation using a panel of established corticosteroids showed good agreement with published PPB figures. The published PPB figure for fluticasone propionate (FP) was, however, found to be an underestimate, and this was subsequently confirmed, at clinically relevant plasma concentrations, to be 99.3%. The modified method was particularly suited to lead optimisation because it provided samples in a consistent matrix compatible with standard high throughput LC/MS/MS analysis.     

亚甲基蓝对猪凝血酶病毒灭活/去除工艺验证实验

目的验证亚甲基蓝光化学法灭活猪凝血酶中病毒的灭活效果,完善生产工艺。方法在SINV、EM-CV、PPV和PRV等4种病毒中,加入0.5 mg/L的亚甲基蓝置于病毒光照灭活仪上光照2 h和用60 kD膜超滤的灭活病毒方法来验证工艺的可行性。结果经亚甲基蓝可有效灭活病毒,经超滤法滤除可有效去除病毒。掺入到凝血酶原液中的病毒以及在原培养物中病毒都丧失了对细胞的感染性,降低的滴度TCID50高于4 Log10。结论亚甲基蓝光化学法可有效灭活猪凝血酶中病毒。     

应用超滤法从发酵液中提纯抗生素

土霉素的生产包括酸化、过滤、脱色和结晶等步骤,多年来土霉素的提纯技术一直没有大的改变。常用的树脂脱色过程成本高,收率低,还因树脂再生产生酸、碱废水而增加污水处理费用。本研究试用超滤取代树脂脱色单元操作以克服其上述缺点。在实验中使用了切割分子量为4960.5u的超滤膜,实验结果表明超滤是取代树脂脱色的有效手段,可以充分纯化土霉素结晶液并改善结晶,脱色过程收率由树脂法的95.3%提高到98.6%。如果超滤与净化剂预处理相结合,可以提高土霉素质量,并使土霉素结晶收率由90.58%提高到93.3%。     

Preparation of proteolytic enzyme extracts from Ananas comosus L., Merr. fruit juice using semipermeable membrane,ammonium sulfate extraction,centrifugation and freeze-drying processes

Crude purified bromelain extracts were obtained from 0.26% protein pineapple juice using sequential batch membrane processing systems which included microfiltration (MF) and ultrafiltration (UF) followed by ammonium sulfate extraction, ultracentrifugation and freeze drying. The membrane treatments (with an 8 μm mineral MF and a 10000 molecular weight cut-off (MWCO) organic UF membranes), combined with 60% ammonium sulfate extraction resulted in 0.75–0.8% protein concentration, with 99% protein rejection. A 70% ammonium saturation and ultracentrifugation process (27 000 × g at 2–3 ° C), prior to freeze drying, were used in the last step to remove the residual non-protein constituents. These processes achieved low-moisture freeze-dried, and light-colored extracts, free of non-protein constituents and which accounted for about 50% yielded extracts containing 98% protein. The extracts assayed for bromelain and proteolytic activity resulted in almost 100% potential recovered, at completion. However, bromelain and proteolytic activity decay during the processes described above is essentially caused by losses through adsorption on the UF membrane relative to the level of concentration reached.     

Icodextrin: a review of its use in peritoneal dialysis

Icodextrin (Extraneal) is a high molecular weight glucose polymer developed specifically for use as an alternative osmotic agent to dextrose during the once-daily long-dwell exchange in peritoneal dialysis (PD). Isosmotic 7.5% icodextrin solution induces transcapillary ultrafiltration (UF) by a mechanism resembling 'colloid' osmosis (unlike hyper-osmolar dextrose-based solutions, which induce UF by crystalline osmosis). In addition, absorption of icodextrin from the peritoneal cavity is relatively slow compared with that of dextrose; this results not only in UF of longer duration, but also a lower carbohydrate load compared with medium (2.5%) and strong (4.25%) dextrose exchanges. In randomised clinical trials of up to 2 years in duration, administration of icodextrin for the long (8- to 16-hour) overnight exchange in continuous ambulatory peritoneal dialysis (CAPD) or daytime exchange in automated peritoneal dialysis (APD) produced net UF which exceeded that with 1.5% and 2.5% dextrose solutions (thereby improving fluid balance), and was equivalent to that with 4.25% dextrose solution. Icodextrin also increased peritoneal clearances of creatinine and urea nitrogen compared with 2.5% dextrose solution. The increase in UF volume with icodextrin was enhanced in CAPD patients with high peritoneal membrane permeability (i.e. high and high-average transporters), maintained in the small number of patients followed-up for 2 years and sustained during episodes of peritonitis. Icodextrin reduced the percentage of patients with net negative UF in contrast to 1.5% and 2.5% dextrose and, in noncomparative studies, extended PD technique survival in patients who had failed dextrose-based dialysis. The use of icodextrin was also associated with some symptomatic improvements and health-related quality of life advantages, and no adverse effect on patient survival, compared with dextrose, although confirmation of these findings is ideally required in appropriately designed studies. The tolerability of icodextrin was generally similar to that of dextrose-based solutions in controlled clinical trials, although there was an approximate three-fold increase in the risk of new skin rash (5.5% vs 1.7%). However, reports of severe cutaneous hypersensitivity reactions remain rare; this possibility should not preclude the use of the polymer. CONCLUSION: 7.5% icodextrin solution offers the first feasible alternative to conventional dextrose solutions for the once-daily long-dwell exchange in PD. It is effective, generally well tolerated and appears to be most useful in situations of reduced or inadequate UF with dextrose, including in high and high-average transporters, during episodes of peritonitis and patients who have failed dextrose-based dialysis.     

高滴度特异性卵黄抗体在天花病毒检测中的研究

天花是人类第一个消灭的疾病,但生物界中仍广泛存在天花类病毒。目前人类还缺乏对其有效、快捷的诊断、检测和治疗措施。在全球面临生物恐怖袭击的时代,这方面的研究工作尤显迫切。以3种灭活的天花病毒（vaccinia virus,calpox virus和cowpox virus）为免疫原,分别免疫注射3组实验蛋鸡,以改良的聚乙二醇法将相应特异性抗体从卵黄中提取。利用免疫荧光实验、抗体中和实验及免疫电镜等实验持续检测抗体滴度变化以及抗体交叉反应。最后,以特异性的卵黄抗体与磁珠（Dynabead）相结合,用于天花病毒的纯化,并在PCR实验中验证其检测效果。抗vaccinia virus抗体和抗calpox virus抗体在高度稀释的情况下（分别以1：10^6,1：10^5稀释）,仍在免疫荧光实验中呈阳性反应,且抗体的这种高表达水平在持续5次免疫注射下,能维持达10个月。抗体的中和能力和抗体与抗原反应的超微结构也在相应实验中观察到。抗体与病毒蛋白结合的特殊条带也在免疫印记实验中观察到。实验显示,不同天花病毒之间存在强而明显的交叉反应。最后,特异性的卵黄抗体能够与磁珠结合,并在模拟样本中用于天花病毒的PCR检测。达到纯化病毒、增强PCR实验敏感度的效果。该研究显示,抗天花病毒卵黄抗体能够应用于天花病毒的诊断,有望以此为基础,开发出快速、有效的天花病毒检测手段。     

膜技术在酶法生产L-色氨酸中去除蛋白质和色素的应用研究

目的 用超滤-纳滤膜分离法去除L-色氨酸酶法转化液中的蛋白质和色素.方法 L-色氨酸转化液经适当处理后,输入超滤膜和纳滤膜设备,分别收集输出的清液和浓液,然后检测其透光率并用加入碱或乙醇方式,检查转化液中蛋白质、色素的去除效果.结果 在转化液pH调节至5.5 ～6.0、温度控制在20～25℃的膜分离工艺条件下,经超滤-...     

Evaluation of a rapid ultrafiltration technique for determination of quinidine protein binding and comparison with equilibrium dialysis

The free level ultrafiltration (UF) assay by the enzyme multiplied immunoassay technique (EMIT) for determination of unbound quinidine concentration in serum (Qf) was evaluated in 50 samples obtained from cardiac patients treated with quinidine for ventricular arrhythmias. Equilibrium dialysis (ED) at 37 degrees C and high performance liquid chromatography (HPLC) served as standard methods for comparison. A good agreement was found between EMIT and HPLC at the low range of free quinidine concentration (0.1-0.7 mg/L) observed in our patients (r = 0.959). Although the correlation between UF and ED was high (r = 0.972), Qf was systematically underestimated by UF. This bias was due to the fact that UF was performed according to the recommendations of the manufacturer at 25 degrees C. No systematic differences were found when 20 additional samples were assayed by the two methods at the same temperature (25 degrees C; r = 0.992). The quinidine binding ratio showed a correlation with the serum concentration of alpha 1-acid-glycoprotein (r = 0.61). The metabolites 3(S)-hydroxyquinidine and quinidine-N-oxide did not influence the protein binding of the parent drug. The importance of adjusting the serum pH to physiological values before measurement of Qf was confirmed in this study. Our results show that, if performed under the same conditions, ED and UF yield practically identical values. Because of easy handling, the EMIT Free Level System II should be applicable under clinical conditions.     

Binding of iralukast to serum proteins and erythrocytes: measurements using ultrafiltration and an erythrocyte partitioning method

The binding of iralukast to plasma (or serum) proteins and to erythrocytes was studied in vitro, at +37°C, using the erythrocyte partitioning method (EPM) and/or ultrafiltration (UF) with ¹⁴C-labelled iralukast. Iralukast was highly bound in human and animal serum (>99%). Similar bound fraction values were obtained with the two methods: in whole human plasma (or serum) 99.8% (EPM) and 99.9% (UF), in albumin solution 99.8% (EPM and UF), in high density lipoprotein solution 97.3% (EPM) and 98.3% (UF), and in low density lipoprotein solution 97.2% (EPM) and 98.8% (UF). Moreover, the erythrocyte partitioning method allowed the evaluation of other binding parameters. The binding capacity (l/μmol) of proteins equalled 35 for low density lipoproteins, 3.6 for high density lipoproteins, 1.0 for albumin, 0.78 for alpha-1-acid glycoprotein, and 0.03 for gamma globulins. In whole blood, iralukast was distributed between plasma and erythrocytes in the proportion (%) 90/10. At physiological protein concentrations, iralukast was primarily bound to albumin (79%).     

基于HPLC-DAD与LC-MS/MS法检测拉莫三嗪血药浓度结果分析及其经济学比较

目的：通过比较高效液相色谱法（HPLC-DAD,以下简称HPLC）和高效液相串联质谱法（LC-MS/MS）测定的拉莫三嗪（LTG）血药浓度值,考察2种方法检测值的一致性,并进行经济学评价。方法：采用HPLC法和LC-MS/MS法分别测定117份癫痫患者体内拉莫三嗪血清样本,用Passing-Bablok回归法和Bland-Altman偏差图分析检测结果的一致性和差异性,并计算2种方法的成本-效益。结果：2种方法得到的拉莫三嗪血药浓度值比较无显著性差异（P=0.707）,Passing-Bablok回归方程为LTG_HPLC=1.074×LTG_LC-MS/MS+0.071,Pearson相关系数为0.997。HPLC法检测结果比LC-MS/MS法偏高（0.216±0.347）μg·mL^-1 （95% CI：-0.465~0.897）。当样本量 > 9例/d时,HPLC法取得收益 > 56.04元/d;样本量 > 16例时,LC-MS/MS法取得收益 > 36.46元/d,HPLC法收益 > 653.77元/d。当HPLC法达到最大检测量（58例/d）,LC-MS/MS法检测量为66例/d时,2种方法产生的日收益相近（4 240.20元 vs 4305.43元）。当检测量 > 85例/d时,LC-MS/MS法的收益 > 5 927.64元/d,远高于HPLC法。结论：HPLC法和LC-MS/MS法在检测拉莫三嗪血药浓度时具有良好的一致性;HPLC法仪器成本较低,适用于检测量 < 58例/d的情况;LC-MS/MS法仪器成本高,单位时间内检测量为HPLC法的3~4倍,日检测量 > 85例时,检测效益优于HPLC法。     

Determination of a beta(3)-agonist in human plasma by LC/MS/MS with semi-automated 48-well diatomaceous earth plate

Methods for the determination of a beta(3)-agonist (A) in human plasma were developed and compared based on high-performance liquid chromatography (HPLC) with tandem mass spectrometric (MS/MS) detection using a turbo ion spray (TIS) interface. Drug and internal standard were isolated from plasma by three sample preparation methods, liquid-liquid extraction, Chem Elut cartridges and 48-well diatomaceous earth plates, that successively improved sample throughput for LC/MS/MS. MS/MS detection was performed on a PE Sciex API 365 tandem mass spectrometer operated in positive ion mode and using multiple reaction monitoring (MRM). The precursor/product ion combinations of m/z 625/607 and 653/515 were used to quantify A and internal standard, respectively, after chromatographic separation of the analytes. Using liquid-liquid extraction and Chem Elut cartridges, the assay concentration range was 0.5-100 ng/ml. Using diatomaceous earth plates, the concentration range of the assay was extended to 0.5-200 ng/ml. For all three assays, the statistics for precision and accuracy is comparable. The assay accuracy ranged from 91-107% and intraday precision as measured by the coefficient of variation (CV) ranged 2-10%. The sample throughput was tripled when the diatomaceous earth plate method was compared with the original liquid-liquid extraction method.     

Sensitivity of real time viral detection by an optical biosensor system using a crude home-made antiserum against measles virus as a ligand

Virus identification in clinical materials during acute phase infections could give necessary information for the treatment of infections by human immunoglobulin (hIg) or interferon (IF). But because of a lack of information, most virus infections have not been treated. We have tried to develop a real time detection system for viruses in general using an optical biosensor and a model virus, Herpes simplex virus Type 1 (HSV-1), and have proved that the HSV-1 virus propagated in Vero cells and diluted in minimum essential medium (MEM) with 10% fetal bovine serum (FBS) could be detected in high sensitivity close to 10 infectious units (50% tissue culture infective dose [TCID50] units) using purified cellular receptor molecules as the ligand because the receptor could be the most specific ligand. However, because ligands available for this system to identify various viral infections in general are limited, we also tested this system using a purified polyclonal antibody which contained many other antigens as the ligand, and produced sensitivity comparable to that using the receptor as a ligand. In this paper, we tested the sensitivity by this system under the worst condition. That is, we used a crude home-made rabbit antiserum against measles virus with host cell debris as a ligand. It was found that less than 500 infectious (TCID50) units of virus were required for detection in a 100 microl solution, and that the efficacy of the commercially available hIg was also estimated by this system. This result suggested that this real time viral detection and titration system was applicable for the diagnosis of all viral infections even under difficult conditions without requiring any complex skills, with high enough sensitivity for clinical purposes. The efficacy of hIg preparations could also be evaluated by this system at the same time of the clinical material sampling.     

Development and validation of a DESI‐HRMS/MS method for the fast profiling of esomeprazole and its metabolites in rat plasma: a pharmacokinetic study

The advances in pharmaceutical development and drug discovery impose the availability of reliable high‐throughput screening methods for the rapid evaluation of drug metabolism and pharmacokinetic (PK) in biological samples. Here, a desorption electrospray mass spectrometry (DESI‐MS) method has been developed and validated for the PK profiling of esomeprazole and its metabolites (5‐hydroxyomeprazole and omeprazole sulfone) in rat plasma. Rats were treated with an esomeprazole solution (2.5 mg/mL) for endovenous administration and the analyte levels were profiled over 2 h after liquid‐liquid extraction from plasma. MS and tandem mass spectrometry (MS/MS) experiments were performed by using a DESI‐LTQ‐Orbitrap XL instrument and an on‐spot fixed time analysis on PMMA surfaces. Validation was performed for the esomeprazole. The DESI‐MS/MS method exhibited for the esomepazole excellent sensitivity (limit of detection (LOD)=60 ng/mL), linearity (0.2‐20 µg/mL concentration range; y=23848(±361)X, n=15; r²=0.987) and precision (RSD<9%) by using an internal standard method. The PK results were discussed in terms of Area Under the Curve, C_max and T_max. Data reliability was demonstrated by comparison with a liquid chromatography‐tandem mass spectrometry method (p>0.05). The data achieved demonstrated that the DESI‐MS method is suitable for sensitive and fast profiling of a drug and its metabolites at the therapeutic concentration levels. Copyright © 2015 John Wiley & Sons, Ltd.     

Development of immunoaffinity solid phase microextraction probes for analysis of sub ng/mL concentrations of 7-aminoflunitrazepam in urine

We report on the development of solid phase microextraction probes for drug analysis, prepared with antibodies specific for benzodiazepines covalently immobilized to the surface. In the technique, immobilized antibody probes are exposed to a sample containing the drug for 30 min. Extracted drugs are subsequently desorbed from the probes in 500 microL of methanolic desorption solution, which is dried, reconstituted in a small volume of injection solution and analysed by LC-MS/MS. The antibodies were characterized both before and after immobilization, to facilitate the rational selection of antibodies for such analyses. Polyclonal and monoclonal antibodies were compared as was the impact of affinity purification of the polyclonal antibody to isolate the drug-specific fraction. The probes were evaluated for utility in analyzing 7-aminoflunitrazepam at sub ng/mL concentrations in urine, which is expected to be found several days after a single oral dose of 2 mg of flunitrazepam. Such analyses are required in monitoring for abuse of this drug, both in terms of 'club drug' use and in cases of drug-facilitated sexual assault. In these cases drug concentrations in blood and urine are much lower than in chronic abuse cases and are difficult to analyse by conventional methods. The method developed has a limit of detection of 0.02 ng/mL, with accuracy ranging from 1% to 27% and precision (% R.S.D.) ranging from 2% to 10% between the lower and upper limits of quantitation for the analysis of 7-aminoflunitrazepam in urine. The dynamic range of the method is from 0.02 ng/mL, which is limited by the instrument sensitivity, to 0.5 ng/mL, which is approaching the capacity of the probes. This would allow for quantitative analysis of samples at concentrations below that measurable by many other methods for general benzodiazepines analysis from urine, and a highly selective screen for samples at higher concentrations. The method has similar limits of detection to the most sensitive literature methods specifically designed for such analysis but with the advantage of significantly simplified sample preparation. This simplification makes the technique more amenable for use by both professionals and non-professionals.     

First observation of cylindrospermopsin in Anabaena lapponica isolated from the boreal environment (Finland)

The cyanobacterial cytotoxin cylindrospermopsin has been mostly associated with cyanobacteria present in tropical and subtropical regions. Cylindrospermopsin has recently been found in cyanobacterial samples in central and southern Europe but the possible presence of the toxin in northern Europe has been unknown. Fifty-eight field and laboratory culture samples of Finnish cyanobacteria were analyzed by high-performance liquid chromatography combined with UV diode-array detection, multiple reactant monitoring in a triple-quadrupole mass spectrometer (MS), and accurate mass measurements using a time-of-flight MS instrument. Cylindrospermopsin was confirmed by all three techniques in a culture sample of Anabaena lapponica at a concentration of 242 microg cylindrospermopsin per g freeze-dried cyanobacterial material.     

In vitro inhibition of influenza A virus infection by marine microalga-derived sulfated polysaccharide p-KG03

The sulfated polysaccharide, p-KG03, purified from the marine microalga, Gyrodinium impudium, is a unique compound comprising homogenous galactose units conjugated to uronic acid and sulfated groups. Although previous studies showed that p-KG03 suppresses tumor cell growth and infection by encephalomyocarditis virus, its effect against enveloped virus infection and the biological mechanism of action have not been elucidated. In this report, the inhibitory activity of p-KG03 against influenza virus was examined and compared with that of other sulfated polysaccharides (fucoidan and pentosan polysulfate) and antiviral agents (oseltamivir phosphate, oseltamivir carboxylate, amantadine, and ribavirin). The results of a cytopathic effect reduction assay using MDCK cells demonstrated that p-KG03 exhibited the 50% effective concentration (EC(50)) values of 0.19-0.48 μg/ml against influenza type A virus infection (selectivity index >200) but not all influenza type B viruses. Mechanism studies showed that inhibition of influenza virus replication was maximized when p-KG03 was added during or within 6 h after viral infection, suggesting that mainly the viral adsorption and internalization steps are targeted by this compound. The results of influenza virus binding assay to p-KG03 and fluorescence microscopy indicate that the antiviral activity of p-KG03 is directly associated with its interaction with viral particles. The sulfated polysaccharide p-KG03 is a potent and specific influenza A viral entry inhibitor and may be a candidate for antiviral drug development.     

Comparison of Flow Injection-MS/MS and LC-MS/MS for the Determination of Ochratoxin A

Two methods for measuring ochratoxin A in corn, oat, and grape juice were developed and compared. Flow injection (FI) and on-line liquid chromatography (LC) performances were evaluated separately, with both methods using a triple quadrupole tandem mass spectrometer (MS/MS) for quantitation. Samples were fortified with ¹³C uniformly labeled ochratoxin A as the internal standard (¹³C-IS) and prepared by dilution and filtration, followed by FI- and LC-MS/MS analysis. For the LC-MS/MS method, which had a 10 min run time/sample, recoveries of ochratoxin A fortified at 1, 5, 20, and 100 ppb in corn, oat, red grape juice, and white grape juice ranged from 100% to 117% with RSDs < 9%. The analysis time of the FI-MS/MS method was <60 s/sample, however, the method could not detect ochratoxin A at the lowest fortification concentration, 1 ppb, in all tested matrix sources. At 5, 20, and 100 ppb, recoveries by FI-MS/MS ranged from 79 to 117% with RSDs < 15%. The FI-MS/MS method also had ~5× higher solvent and matrix-dependent instrument detection limits (0.12–0.35 ppb) compared to the LC-MS/MS method (0.02–0.06 ppb). In the analysis of incurred corn and oat samples, both methods generated comparable results within ±20% of reference values, however, the FI-MS/MS method failed to determine ochratoxin A in two incurred wheat flour samples due to co-eluted interferences due to the lack of chromatographic separation.     

中空纤维离心超滤-HPLC法测定盐酸环丙沙星凝胶剂的含量

目的：建立中空纤维离心超滤的样品前处理技术结合HPLC法测定盐酸环丙沙星凝胶剂的含量。方法：采用中空纤维离心超滤技术对凝胶样品进行提取分离,基于中空纤维超滤膜特异性截留大分子原理,有效滤除了样品溶液中的高分子凝胶辅料卡波姆,超滤液直接进色谱分析。采用Promosil C₁₈柱（150×4.60 mm,5 μm）,流动相为甲醇-水（28∶72,含0.5%的三乙胺,用磷酸调节至pH4.0）,检测波长为278 nm。结果：盐酸环丙沙星在2.51~25.1 μg·mL^-1范围内呈良好的线性关系（r=0.999 7）,样品平均回收率为101.0%,RSD小于2%。结论：所建方法简便,快速,定量准确,色谱峰形良好,可用于盐酸环丙沙星凝胶剂的含量测定。