6^th International Conference and Exhibition on Advances in Chromatography & HPLC Techniques August 02-03, 2018 Barcelona, Spain

Biography:

Mladen Krajacic is a Full Professor at University of Zagreb where he completed his PhD in Virology. As a Head of the Laboratory and Principal Investigator in several research projects, he has been leading the molecular biology investigations on viruses, sub-viral agents (viroids, satellite RNAs) and non-cultivable bacteria. During the last decade, he has dedicated to chromatography on monolithic supports and its appliance in separation of viral and sub-viral ribonucleic acids, as well as virus particles. His results have been published in prominent scientific journals in which he has regularly been employed as a Reviewer. He was supervising a number of young research fellows who have received positions from well recognized institutions across the world.

Abstract:

The particular structure of monolithic chromatographic supports has been proved highly advantageous in analytic separation and purification of large biomolecules. Characterized by very high porosity, high binding capacity, and high flow rate based on convective mass transport, monoliths are particularly applicable in virus research. Monolith chromatography has hardly any limitation to be applied in processing of virus particles, huge macromolecular complexes, and viral genomes, especially when being distinct from nucleic acid forms present in the host cell. The majority of trials published so far, have exploited ion exchange carriers, although other chemistries are also applicable, like hydrophobic interaction when concentrating viruses from marine environment. According to papers published over the past decade, viruses of different sizes, structures and morphologies, even virus-like particles, have successfully been purified from tissue homogenates or cell lysates. Moreover, following virtual separation by overlapping chromatograms obtained from separate experiments, a real separation of three virus species, and a distant strain of one of them (four viral fractions in total) was accomplished from laboratory prepared mixture. One of the most recent achievements is a proof of principle that virus chromatographic feature could easily be modified without abolishing its structural stability, or its biological activity. As adenoviruses have been used in almost a quarter of human gene therapy trials, the experiment was conducted with an adenoviral vector. The deletion of just two negatively charged amino acids from the main structural protein was efficient in shortening chromatographic retention of the recombinant adenovirus. In this way, it would be possible to shift virus particles away from particular interfering substances present in the crude lysate. The concept might be followed to facilitate chromatography-approach purification by engineering modifications contributing to virus separability, besides those contributing to its therapeutic functionality.

Biography:

Fotios Tsopelas is a Lecturer in the School of Chemical Engineering at National Technical University (NTUA), Greece. He studied Chemical Engineering in the NTUA (1999) and Pharmacy (2004) in the National and Kapodistrian University of Athens. He completed his PhD in 2007 in Environmental Analytical Chemistry at NTUA (scholarship from Onassis Foundation) and his Postdoctoral research was focused on biomimetic chromatography for novel drug design. He has more than 25 publications in peer-reviewed scientific journals and more than 40 contributions in international conferences. He has acted as a Referee in more than 20 international journals. He has participated as a member of scientific committee in two international conferences. He has coordinated five national and international funded research projects. His research interest is mainly focused on the development of biomimetic chromatographic approaches for the evaluation of pharmacokinetic properties of candidate drugs and ecotoxicity of emerging pollutants.

Abstract:

Pidgeon and his coworkers described for the first time in 1989, the immobilization of phosphatidylcholine to propylamino-silica skeleton and up to now immobilized artificial membrane (IAM) chromatography have been employed for simulation of the environment of cell membranes. In particular, IAM chromatography constitutes a valuable tool for medicinal chemists for prioritization of drug candidates in the early drug development stages. The retention outcome on IAM stationary phases encodes lipophilicity, electrostatic and other secondary interactions in contrary to traditional octanol-water partitioning. An increasing number of publications in recent years suggest that IAM indices, including isocratic logk(IAM) or extrapolated logkw(IAM) retention factors, hydrophobicity index CHI(IAM) which corresponds to the percentage of acetonitrile required for equal partitioning of the solute between mobile and stationary phase (i.e. logk=0) or the polarity parameter Δlogkw(IAM) can successfully model the passage of xenobiotics through biological membranes and barriers and predict pharmacokinetic properties, often in combination with additional descriptors. More recently, IAM chromatography is applied to estimate toxicological endpoints in regard to drug safety, such as the phospholipidosis potential, or in regard to chemicals risk hazard including the bio-concentration factor and aquatic organisms’ toxicity. The presentation will be devoted to applications of IAM chromatography to medicinal chemistry and environmental sciences. Examples referring to modeling of human oral absorption, blood-brain penetration, skin partition as well as bioconcentration factor and median toxicity (LC50) in aquatic organisms will be discussed. The combination of promising results in both medicinal chemistry and in environmental science with the speed, reproducibility and low analyte consumption suggest that a broader application of IAM chromatography in early drug discovery process and in ecotoxicity is expected in initial drug candidate selection and contribute to reduced risk hazard of chemicals.

Biography:

Monika Šuleková has completed her Graduation at the University of Pavol Josef Šafárik in Košice, Slovakia. During her university studies, she spent half a year at Friedrich-Schiller University in Jena, Germany where she studied Analytical Chemistry. Nowadays she works as a Teacher at the University of Veterinary Medicine and Pharmacy in Košice, Slovakia as well as a Researcher in the field of desorption of drugs from mesoporous silica modified by different functional groups, and in determination of synthetic dyes in pharmaceutical products by the RP-HPLC method.

Abstract:

Naproxen, a non-steroidal anti-inflammatory drug (NSAID) is widely used to moderate pain relief in the treatment of many diseases. Naproxen has analgesic and antipyretic properties. Mesoporous silica SBA-15 was prepared to evaluate its application as naproxen drug delivery system. The amount of naproxen released from the pores of mesoporous silica SBA-15 into the solutions was determined by the method of a reverse-phase high performance liquid chromatography (RP-HPLC). SBA-15 having 3-aminoprophyl-, methyl-, fenyl-and cyclohexyl-surface groups was successfully prepared by the grafting of SBA-15 with the corresponding alkoxysilanes. The release of the drug was performed in two different media, in a simulated body fluid (pH 7.40) and in a simulated gastric fluid (pH 2.06). The HPLC system Dionex Ultimate 3000 RS (Thermo Fisher Scientific, Germany) consisted of a quaternary pump, a degasser, an automated injector, a column oven and a diode array detector DAD. HPLC system was used, with stationary phase ODS Hypersil C18 column (150x4.6 mm, 3 μm). To determine the concentration of naproxen, the calibration curve has been established based on five solutions of different concentrations of naproxen. The linearity was determined by threefold repeating measurement of each concentration step. The mixture of acetonitrile and water (55:45, v/v) adjusted with ortho-phosphoric acid to pH 3 was selected as the best mobile phase. The flow rate was 1 mL/min and detection was carried out at a wavelength of 229 nm. During the chromatographic separation, the mobile phase was kept isocratic. The release of the drug was studied as a function of time and the results are shown in Figure.

Biography:

Clydewyn M Anthony is a Senior Scientific Liaison in the Chemical Medicines Division at The United States Pharmacopeial Convention (USP) and is currently responsible for the modernization of documentary standards for Over-The-Counter drug formulations. He completed his PhD in Analytical Chemistry at The Pennsylvania State University and BS in Chemistry in Hunter College at the City University of New York. Prior to joining USP in May 1999, he worked as a Research Chemist with Texaco Inc. where he was responsible for performing and overseeing the compositional analyses of motor oils, gasolines and all aftermarket petroleum related products on both the domestic and international markets. He also held the position of Criminalist with the New York Police Forensic Investigation Department, worked on thousands of narcotics and arson related cases, and testified as an Expert Witness on behalf of the New York City Police Department.

Abstract:

A few years ago there was a potentially serious health issue in Pakistan and Paraguay which spurred investigation and responsive action by both the United States Pharmacopeial Convention (USP) and The Food and Drug Administration (FDA). This crisis resulted in the deaths of adults and children who had ingested Dextromethorphan Cough Syrup. It was later determined and confirmed that toxic levels of the controlled substance, levomethorphan, an enantiomer of dextromethorphan, was found in the drug formulation and was responsible for the resulting deaths. USP has thus charged with developing a quantitative procedure for monitoring levomethorphan and simultaneously incorporating this method as a revision to the documentary standard within its compendium. At the time of the public health issue, the existing USP Dextromethorphan monograph did not include a quantitative procedure for the determination of its enantiomer, levomethorphan. Hence a chiral HPLC method was developed to bring the monograph up-to-date and simultaneously address obvious safety concerns associated with the enantiomer. The proposed HPLC method separates levomethorphan and dextromethorphanone (another impurity, dextromethorphan Related Compound C) from dextromethorphan; and allowed quantitation to satisfy acceptance criteria requirements for these impurities (0.10%). Hence, manufactured lots which test higher than the specified limit of levomethorphan can be rejected thus helping to prevent potential safety issues in the future. A complete overview of the issues encountered in the development of this chiral HPLC method along with the challenges associated with the implementation of a global procedure which utilizes a schedule 11 controlled substance as a public standard will be presented.

Biography:

Yuichi Negishi has completed his PhD at Keio University, Japan. He is a Professor of Tokyo University of Science, Japan. He has over 140 publications that have been cited over 7,200 times. In his publications, 10 papers are categorized to top 1% cited papers. His publication H-index is 45. He has been awarded several prizes, including the PCCP Prize (2007), CSJ Award for Young Chemists (2008), Japan Society of Molecular Science Award for Young Chemists (2012), and Yagami Prize (2017).

Abstract:

Small thiolate-protected gold clusters have attracted considerable attention as new functional nanomaterials because they have size-specific properties and functions that are not found for bulk gold. In particular, hydrophilic thiolate-protected gold clusters (hereinafter referred to as hydrophilic gold clusters) exhibit high biocompatibility and luminescence quantum yield in addition to pollution-free properties. Therefore, hydrophilic gold clusters are expected to be used in biomedical and environmental applications. Replacing some of the Au atoms in these clusters with different elements may impart them with even more useful functions. However, the synthesis of hydrophilic metal clusters has been less studied because of the complexity involved in evaluating the mass distributions of product mixtures. In this work, we found two hydrophilic interaction liquid chromatography (HILIC) columns for high-performance liquid chromatography (HPLC) suitable for the high-resolution separation of hydrophilic metal clusters. The mass distributions of the product mixtures of hydrophilic metal clusters were evaluated via HPLC mass spectrometry (LC/MS) using these HILIC columns. Consequently, we observed multiple clusters that had not been previously reported for glutathionate (SG)-protected gold clusters (Aun (SG)m). Additionally, we demonstrated that Aun−xMx(SG)m alloy clusters (M=Ag, Cu, or Pd) in which part of the Au in the Aun (SG)m cluster is replaced by a hetero element can be synthesized, similar to the case of hydrophobic alloy clusters. It is easy to evaluate the mass distributions of hydrophilic metal clusters using this method. Thus, remarkable progress in the synthesis techniques of hydrophilic metal clusters through the use of this method is anticipated, as is the situation for hydrophobic metal clusters.

Biography:

Roberto Biondi completed his undergraduate degree in Medicine and Surgery from University of Perugia, Italy; pursued PhD in Biological Sciences from Pisa University, Pisa, Italy. He is currently a Visiting Professor at the Maternal-Fetal Assistance Excellence Unit, of Alessandru Rusescu National Institute for Health of Mother and Child, Bucharest, Romania. He has developed a method on the detection and scavenging of hydroxyl radical via D-phenylalanine hydroxylation in human fluids at the same institute. He has worked on the detection of tetrahydrobiopterin oxidation products in myocardial ischemia and also during chemical oxidation at both Johns Hopkins University in Baltimore, Maryland, USA and Ohio State University in Columbus, Ohio, USA.

Abstract:

Living organisms have developed a complex antioxidant system to counteract reactive oxygen species (ROS) and oxidative stress (OS) damage. An array of small antioxidant molecules (vitamin C, vitamin E, and flavonoid) were part of the antioxidant barrier and were evaluated using a detection system based on Fenton reaction-mediated D-phenylalanine (D-Phe) hydroxylation. This reaction in turn generated o-tyrosine, m-tyrosine and p-tyrosine that were separated by HPLC (high performance liquid chromatography) with fluorescence detectors. The addition of antioxidants competed with D-Phe on .OH attack, thus allowing to determine the .OH scavenger capacity. Using a kinetic approach, the hydroxyl radical scavenger capacity was applied to biological fluid samples but also to antioxidants as N,N dimethylthiourea. In order to define the process of ROS-mediated tetrahydrobioptern (BH4) degradation in isolated rat hearts subjected to ischemia, a direct HPLC assay of BH4 and its pteridine derivatives was used. The lack of oxygen due to myocardial ischemia determined ROS generation and consequently caused a BH4 cardiac content decline while the dihydroxanthopterin was produced. Contrarily other researchers found a BH4 depletion with BH2 increased levels in the vessels of a range of cardiovascular disease including hypercholesterolemia, diabetes, atherosclerosis, hypertension and heart failure. HPLC estimation was based on indirect method in which BH2 and BH4 were oxidized to biopterin by iodine and then evaluated by fluorescence detector. It is debated whether the provision of OHSC (OH scavenging capacity) assay to the industries producing artificial human milk, types of tea and wine might better improve the knowledge of their antioxidant properties. Regarding to pterin biosynthetic pathway, it is discussed if the ischemia- and OS-determined BH4 alteration could result more comprehensive using a sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) method. This technique could be applied to amniotic fluid samples collected from pregnant women suffering from preeclampsia in whose pathogenesis the main role is due to ischemia/reperfusion- caused ROS generation.

Biography:

Madhuri Singhal has completed her PhD at Dr Hari Singh Gaur University, Sagar and Postdoctoral work at Allahabad University, Allahabad. Her research area is the role of natural products from medicinal plants in drug discovery and development. She has presented research papers in international conferences in Australia in 2005 and was invited as visiting academic in 2006 at Australian University. She has presented papers in USA in 2009 and 2015. In 2010, she has presented research paper at Ubon Ratchathani University, Thailand. In 2011, she has presented paper in Hong Kong. She is an Editorial Board Member of an international research journal. More than 30 research papers have been published. At present, she is a Professor of Chemistry in Government Motilal Vigyan Mahavidyalaya, Bhopal.

Abstract:

Chromatographic techniques are extensively used for isolation and separation of bioactive constituents present in different plants having medicinal value. Plant extracts after cold and hot extraction process were subjected to column chromatography for the separation of different bands corresponding to class of compounds using different solvents in various proportions. Portions having sufficient compound, after evaporation are separated to a single compound by using thin layer chromatography. In her research work author has isolated novel isoflavone glycoside, prenylated flavone glycoside and novel triterpenoidal saponin from the plant Pithecellobium dulce. She has also isolated different compounds from plants Terminalia bellerica, Moringa oleifera and other indigenous medicinal plants. The extract of Terminalia bellerica was found to possess anticancer activity while the extract of plant Moringa oleifera was found to have anti-diabetic activity. The application of chromatography technique on natural products is quite useful in order to form herbal based drug with no side effects and toxicity.

Biography:

Mariana G Selener has completed her Bachelor degree in Pharmacy in the Faculty of Pharmacy and Biochemistry at University of Buenos Aires (UBA) in 2011. She is pursuing her PhD at Chair of Pharmacognosy at the same University. She has expertise in analytical development. The aim of her PhD research work is the isolation and identification of bioactive compounds from Argentine Asteraceae species. She is an Assistant at the Chair of Pharmacognosy at the Faculty of Pharmacy and Biochemistry, University of Buenos Aires (UBA) for undergraduate courses.

Abstract:

Chagas disease is a parasitic disease caused by the protozoan Trypanosoma cruzi. According to the World Health Organization (WHO) this parasitosis affects 6-7 million people worldwide. In Argentina it is estimated that approximately 1.5 million people are infected. The current available drugs used for its treatment, nifurtimox and benznidazole, have limitations due to host toxicity, side effects and low efficacy. In this context, it is extremely necessary to develop new drugs. Nature has provided useful drugs that are used nowadays to treat different pains. Asteraceae species have been a rich source of active compounds and have been attractive for drug discovery. In previous work the trypanocidal activity of the dichloromethane extract of Gymnocoronis spilanthoides (Asteraceae) [GSDE] has been demonstrated. The aim of this investigation was to isolate and identified the active compounds present in GSDE. GSDE was purified by liquid-liquid partition and fractionated by column chromatography using Silicagel-60 and a gradient of CH2Cl2 and EtoAc. From fractions eluted with CH2Cl2: EtoAc (2:1) a pure compound was isolated (compound A). The GSDE as well as compound A were analyzed by HPLC (C18 column, linear gradient elution mode and UV/Vis absorbance detector). The structure elucidation of the isolated compound was performed by spectroscopic methods. The trypanocidal activity of compound A was evaluated on T. cruzi epimastigotes (RA) by the [3H]-thymidine uptake assay. The cytotoxicity of this compound on mammalian cells was performed using mouse splenocytes. Compound A presented a significant trypanocidal activity (IC50= 1.6 μg/ml). This compound showed some toxicity to mammalian cells (CC50=4.9 μg/ ml). The compound A presented 98% purity (by HPLC) and was identified as the ent-11α-hydroxy-15-oxokaur-16-en-19-oic acid. The trypanocidal activity of ent-11α-hydroxy-15-oxokaur-16-en-19-oic acid on trypomastigote and amastigote forms will be evaluated. We will also continue with the isolation and identification of other compounds present in the active extract.

Biography:

Amal Al Sabahi has completed her BSc in Science Education at Sultan Qaboos University (SQU) in 1996 and MSc in Chemistry in Department of Chemistry, College of Science at SQU in 2003. Currently, she is pursuing PhD in Chemistry in the same department. She worked as a Chemistry Teacher for 10 years and as Educational Researcher for six years. She worked as a Lab Instructor in SQU for three years.

Abstract:

Statement of the Problem: Many labeling reagents are commercially available for quantifying primary alcohols; however, these reagents show some drawbacks, such as toxicity, lack of sensitivity and selectivity, low solubility, and high cost. Using fluorescent heterocyclic compounds as labels is an outgrowing field in analytical chemistry. Pyrazolines are well known heterocycle owning fascinating photophysical properties that qualify them for sensing and imaging of bioorganic molecules. The applications of these dyes span many areas from photodynamic cancer therapy, organic light emitting diodes, to fibers whitening and brightening. However, recently it has been evaluated as potential candidates for pre-column derivatization of amino acids and neurotransmitters. The purpose of this study is to evaluate the adequacy of the newly synthesized pyrazoline, 4-(1-(4-trifluoromethyl) phenyl)-4, 5-dihydro-3-(naphthyl)-1H-pyrazole-5-yl) benzoic acid (TFNPB) as a label for primary alcohols and to develop a pre-column derivatization method for quantifying these analytes in different matrices.

Methodology & Theoretical Orientation: TFNPB was synthesized by the conventional method, which involves two steps, an aldol condensation reaction between acetyl-naphthalene and 4-formylbenzoic acid followed by Michael addition of the phenyl-hydrazine. The photophysical properties including absorption, emission, and lifetime measurements have been studied in different solvents. Primary alcohols were then derivatized by this reagent, and LC-MS was used to assess the produced derivatives. The derivatization procedure was optimized, and the assay of alcohols by this method was validated.

Findings: TFNPB shows excellent photophysical properties including high fluorescence intensity and quantum yield. It emits in the visible region at 460 nm in acetonitrile. It is used to derivative hydroxyl groups instantly at low temperature and in short reaction time. Alcohol derivatives show strong well-separated peaks (resolution μ1.5) on C8 column using 75% ACN in water. The produced derivatives were stable at room temperature for more than one month. Excellent linear relationships were obtained for four alcohols in the range 1.25-94 μmol L-1 (R2≥0.991). All figures of merit were calculated and the method developed was validated for the quantitative analysis of primary alcohols.

Biography:

Abstract:

Biopartitioning micellar chromatography (BMC) utilizes micelles formed by a surfactant, such as polyoxyethylene (23) lauryl ether (Brij35), in a concentration higher than its critical micelle concentration and a reversed-phase stationary phase in order to gain insight into drug-membrane interactions, by rapid, friendly and reproducible measurements. The chromatographic column, modified by the surfactant, resembles the ordered array of the membranous hydrocarbon chains in regards to hydrophilic/hydrophobic character and the interactions of xenobiotics with the H-bonding groups of the adsorbed surfactant similar to the membrane ones. Therefore, the characteristics of the BMC are similar to biological barriers. Up to now, the reported studies of BMC to model toxicity (LD50), blood-brain barrier penetration, plasma clearance, volume of distribution as well as oral absorption are based on limited datasets and therefore, further investigations are needed. The aim of the present study was the evaluation of the potential of biopartitioning micellar chromatography to estimate cell permeability. For this purpose, retention indices (logkw) of an extended set of structurally-diverse drugs were measured on a discovery RP- 18 column using as eluent phosphate buffer in the presence of Brij at a concentration of 0.04 M. The effect of the addition of NaCl in a concentration of 9.2 g/L was studied as well as the effect of increase of temperature from ambient to 37oC. Retention factors were compared with octanol-water partitioning and retention factors obtained in immobilized artificial membrane (IAM) chromatography. Retention factors were subsequently compared with Madin Darby Canine Kidney (MDCK) cell lines permeability data taken from literature and they were used to model % Human Oral Absorption (% HOA) data, compiled from literature sources. For reasons of comparison, the constructed models were compared with those derived by octanol-water partitioning.

Biography:

Tomasz Tuzimski is an Adjunct Professor in Department of Physical Chemistry at Faculty of Pharmacy with Medical Analytics Division, Medical University of Lublin, Poland. He has completed MA degree and PhD degree in 1995 and 2002, respectively, and Doctor of Sciences in Habilitation in 2012 at the Faculty of Pharmacy with Medical Analytics Division, Medical University of Lublin. His scientific interests include the theory and application of liquid chromatography, extraction (QuEChERS) and detection techniques. He has authored or co-authored more than 62 scientific papers (IF=100, H-index=17). He is a member of the Editorial Board of the Acta Chromatographica. He is Quest Editor of special sections on pesticide residues analysis in the JAOAC Int. He has co-authored and coedited with Professor Joseph Sherma the book entitled “High Performance Liquid Chromatography in Pesticide Residue Analysis” and the book “Determination of Target Xenobiotics and Unknown Compounds Residue in Food, Environmental and Biological Samples”.

Abstract:

Bisphenols are a class of chemicals with two hydroxyphenyl functionalities, which include bisphenol A (BPA) and several analogues such as bisphenol S (BPS), bisphenol F (BPF), and bisphenol B (BPB). As industrial chemical, BPA is widely used in the production of polycarbonate plastics (used in food contact materials, such as food containers, baby food and water bottles), epoxy resins (used as internal coating in canned food and beverage to prevent the food to get in contact with the metal wall and the deterioration of cans) and as an antioxidant in polyvinyl chloride (PVC) plastics in materials intended to come into contact with food (packaging cling films). Considering the similarity in the structure of various analogues can migrate from food containers. Due to their fat content, both breast and dietary milk could be polluted by many xenobiotics characterized by lipophilic properties. Breast milk, milk of lactating cows and dairy products are widely consumed by infants, children and many adults throughout the world and occurrence of quantifiable amounts of BPA represents a matter of public health concern. Due to the ability to accumulate in organisms and in the food chain, BPA may also affect the development of subsequent generations. The breastfeeding could represent a source of exposure to infants, because BPA tends to persist for extended periods of time and infants can be more susceptible to adverse effects resulting from chemical exposures, due to the rapid mental and physical changes that take place during prenatal and neonatal periods. Despite the reported levels in food are generally considered safe, the occurrence of BPA residues in breast milk and dairy products should be considered significant in terms of potential human-health risk. The aim of the paper was determination of xenobiotics (bisphenols, pesticides, dyes, medicines) in breast milk samples and food products.