Oxytetracycline recovery from aqueous media using computationally designed molecularly imprinted polymers

Polymers for recovery/removal of the antimicrobial agent oxytetracycline (OTC) from aqueous media were developed with use of computational design and molecular imprinting. 2-Hydroxyethyl methacrylate, 2-acrylamide-2-methylpropane sulfonic acid (AMPS), and mixtures of the two were chosen according to their predicted affinity for OTC and evaluated as functional monomers in molecularly imprinted polymers and nonimprinted polymers. Two levels of AMPS were tested. After bulk polymerization, the polymers were crushed into particles (200–1000 μm). Pressurized liquid extraction was implemented for template removal with a low amount of methanol (less than 20 mL in each extraction) and a few extractions (12–18 for each polymer) in a short period (20 min per extraction). Particle size distribution, microporous structure, and capacity to rebind OTC from aqueous media were evaluated. Adsorption isotherms obtained from OTC solutions (30–110 mg L-1) revealed that the polymers prepared with AMPS had the highest affinity for OTC. The uptake capacity depended on the ionic strength as follows: purified water > saline solution (0.9 % NaCl) > seawater (3.5 % NaCl). Polymer particles containing AMPS as a functional monomer showed a remarkable ability to clean water contaminated with OTC. The usefulness of the stationary phase developed for molecularly imprinted solid-phase extraction was also demonstrated

Does size matter? Study of performance of pseudo-ELISAs based on molecularly imprinted polymer nanoparticles prepared for analytes of different sizes

The aim of this work is to evaluate whether the size of the analyte used as template for the synthesis of molecularly imprinted polymer nanoparticles (nanoMIPs) can affect their performance in pseudo-enzyme linked immunosorbent assays (pseudo-ELISAs). Successful demonstration of a nanoMIPs-based pseudo-ELISA for vancomycin (1449.3 g mol) was demonstrated earlier. In the present investigation, the following analytes were selected: horseradish peroxidase (HRP, 44 kDa), cytochrome C (Cyt C, 12 kDa) biotin (244.31 g mol) and melamine (126.12 g mol). NanoMIPs with a similar composition for all analytes were synthesised by persulfate-initiated polymerisation in water. In addition, core-shell nanoMIPs coated with polyethylene glycol (PEG) and imprinted for melamine were produced in organics and tested. The polymerisation of the nanoparticles was done using a solid-phase approach with the correspondent template immobilised on glass beads. The performance of the nanoMIPs used as replacement for antibodies in direct pseudo-ELISA (for the enzymes) and competitive pseudo-ELISA for the smaller analytes was investigated. For the competitive mode we rely on competition for the binding to the nanoparticles between free analyte and corresponding analyte-HRP conjugate. The results revealed that the best performances were obtained for nanoMIPs synthesised in aqueous media for the larger analytes. In addition, this approach was successful for biotin but completely failed for the smallest template melamine. This problem was solved using nanoMIP prepared by UV polymerisation in an organic media with a PEG shell. This study demonstrates that the preparation of nanoMIP by solid-phase approach can produce material with high affinity and potential to replace antibodies in ELISA tests for both large and small analytes. This makes this technology versatile and applicable to practically any target analyte and diagnostic field

Leucocytes, urea and glucose levels in Albino Wistar rats exposed to doses of isolated Achatina achatina snail lectin

There are five (5) types of mature White blood cells (WBC) or Leucocytes found in the peripheral blood viz, Neutrophils (NEU), Eosinophils (EOS) and Basophils (BAS) (granulocytes); Monocytes (MON) and Lymphocytes (LYM) (agranulocytes). Urea is an organic chemical compound, and is essentially the waste produced by the body after metabolizing protein. Urea levels can be used to detect diseases and disorders that affect the kidneys. A common disease related to irregular management of glucose is diabetes. Lectins are proteins that recognize specifically and bind reversibly the carbohydrate-containing molecules of foreign cells and that elicit diverse physiological responses in various organisms. A total of 120 samples of Nigeria Achatina achatina snail specie were collected, authenticated at the Zoology Department of University of Nigeria, Nsukka and 80mls of pooled crude Lectin extract was obtained. Purifications were performed on 20mls of the crude extract in three steps viz, Ammonium sulphate precipitation and Dialysis (Partial purifications), Con A Sepharose 4B affinity Chromatography column (Complete purification). The affinity purified Lectin was used for all the tests conducted in this research. The crude, partially and complete/affinity purified Lectin extracts were subjected to Haemagglutination tests. The Lectin was further assessed to determine its effects on Leucocytes, Urea and Glucose as follows: A total of Thirty-five (35) male Albino Wistar Rats weighing 101-180g and aged 2-3 months obtained from the Animal house of University of Nigeria, Nsukka, were used in this research. The animals were Grouped into 5 (A-E) and allowed for 2 weeks acclimatization. Graded doses of 0.04ml, 0.05ml and 0.06ml of the Affinity purified Lectin were injected intra-peritoneally into each of the Rats in Groups A-D (test groups) according to their body weights at intervals of 2 days for 1 week. Group E served as the control. Two (2) mls of blood was collected from each of the Rats before and 24 hours after the last day of Lectin Doses injections for the following tests: WBC-Total and Differential counts (using Sysmex Corporation, 1999 automated equipment), Urea and Glucose estimations (performed by means of Urease-Berthelot and GOD-PAP Randox Monza automated analyser methods respectively). The results of the research showed as follows: On complete/affinity purification, 15mls of pure sample containing only the high molecular weight Lectin was obtained. The haemagglutination tests conducted showed on standardization preferential agglutination with Blood group A type. Bar Charts statistics show that there was Post Lectin Doses injections mean increase in Total WBC, NEU, LYM and decrease in MON, EOS, BAS, Urea and Glucose levels. However, the differences in Pre and Post Lectin Doses injections mean values of these parameters were further subjected to One way analysis of variance (ANOVA) test statistics to determine if statistically significant. The ANOVA statistics show that the effects of the Lectin on all the assessed Leucocytes parameters viz, Total WBC, and Differential LYM, NEU, MON, EOS, BAS, the Urea and Glucose levels were found to be statistically insignificant. However, the EOS values of only group A was statistically significant. This research has therefore succeeded in Assessment of Activities of the A. achatina snail Lectin on Leucocytes, Glucose and Urea levels

Purification and characterisation of lectin isolated from Nigeria achatina achatina snail

Lectins are carbohydrate-binding proteins that are highly specific for sugar moieties of other molecules. They perform recognition on the cellular and molecular level and play numerous roles in biological recognition phenomena involving cells, carbohydrates, and proteins. Blood groups are inherited characters which give rise to antigen-antibody reaction. A total of 120 samples of local (Nigeria) Achatina achatina snail specie were collected, authenticated at the Zoology Department of the University of Nigeria, Nsukka and 80mls of pooled crude lectin extract was obtained. Purifications were performed on 20mls of the crude extract in three steps viz, Ammonium sulphate precipitation and Dialysis (Partial purifications), Con A Sepharose 4B affinity chromatography column (Complete purification). The affinity purified lectin was used for all the actual tests conducted in this research. The crude, partially and complete/affinity purified lectin extracts were subjected to Haemagglutination tests, Protein Assay and Specific Sugar determinations. The molecular weight was assessed by Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) method. The results of the research showed as follows: On complete/affinity purification, 15mls of pure sample containing only the high molecular weight lectin was obtained. The respective haemagglutination tests on the crude, partially and affinity purified lectin showed on standardisation, preferential agglutination with Blood group A type. The Protein contents of the lectin was deduced to be as follows: The crude extract contains 13.5mg/dl, Dialysed precipitate – 5.7mg/dl, Dialysed supernatant – 5.0mg/dl and the Affinity purified Lectin – 0.422mg/dl. Galactose N-acetyl amine (Gal NAc) residue was determined to be its specific sugar. The SDS-PAGE analysis showed the molecular weight of the lectin to be 250 KDaltons. This research has therefore succeeded in the Purification, Characterisation and illustration of the lectinic properties of the local Nigeria snail - Achatina achatin

Liver function tests values in albino wistar rats administered with isolated Nigeria Achatina achatina snail lectin

Achatina achatina snail specie are considered by many people in Nigeria, Ghana and other parts of West Africa to be the most prized snail for eating. In general, lectins bind to sugar moieties in cell walls or membrane, thereby change the physiology of the membrane to cause agglutination, mitosis or other biochemical changes in the cell. It has been deduced that lectins could be toxic and can as well be used as potent administrations that could be used or serve as substitutes for routine treatment or management of many disorders. Based on these, the toxicity of the Achatina achatina snail lectin in animals was investigated with a view to determining the nutritional value of the snail as food stuffs by carrying out tests to determine the blood values of Liver Function Tests (LFT) parameters in Albino Wistar Rats administered with the lectin. A total of 120 samples of the Nigeria Achatina achatina snail specie were collected, authenticated at the Zoology Department of the University of Nigeria, Nsukka and 80mls of pooled crude Lectin extract was obtained. Purifications were performed on 20mls of the crude extract in three steps viz, Ammonium sulphate precipitation and Dialysis (Partial purifications), Con A Sepharose 4B affinity Chromatography column (Complete purification). The affinity purified lectin was used for all the tests conducted in this research. The crude, partially and complete/affinity purified Lectin extracts were subjected to Haemagglutination tests. The Lectin was further assessed to determine its effects on Liver Function Tests (LFT) parameters viz, Total bilirubin (TB), Conjugate bilirubin (CB), Alkaline phosphatase (ALP), Aspartate transaminase (AST) and Alanine transaminase (ALT) as follows: A total of Thirty-five (35) male Albino Wistar Rats weighing 101-180g and aged 2-3 months obtained from the Animal house of University of Nigeria, Nsukka, were used in this research. The animals were Grouped into 5 (A-E) and allowed for 2 weeks acclimatization. Graded doses of 0.04ml, 0.05ml and 0.06ml of the Affinity purified Lectin were administered intra-peritoneally to each of the rats in Groups A-D (test groups) according to their body weights at intervals of 2 days for 1 week. Group E served as the control. Two (2) mls of blood was collected from each of the rats before and 24 hours after the last day of lectin administration for the following tests: TB, CB, ALP, AST and ALT (performed by means of Roche Cobas C111 automated chemistry analyser). The results of the research showed as follows: On complete/affinity purification, 15mls of pure sample containing only the high molecular weight lectin was obtained. The respective haemagglutination tests on the crude, partially and affinity purified Lectin showed on standardization preferential agglutinations with Blood group A type. Bar charts statistics show that there was Post lectin administration mean increase in TB, CB and AST when the Post administrations values were compared with the Pre values. The Bar charts statistics show that there was Post lectin administration mean decrease in ALP and ALT. However, the differences in the Pre and Post administration mean values of these parameters were further subjected to one way analysis of variance (ANOVA) test statistics aimed at determining whether the mean increases or decreases in these assessed parameters were statistically significant. The ANOVA statistics show that the effects of the lectin on all the assessed LFT parameters viz, TB, CB, ALP, AST and ALT were statistically insignificant (P > 0.05). The results obtained in this research has succeeded in demonstrating that the A. achatina snail lectin is non-toxic, non-carcinogenic and therefore point to its nutritive value as food stuff, hence supports the snail eating education

Molecularly Imprinted Polymers for Ochratoxin A Extraction and Analysis

Molecularly imprinted polymers (MIPs) are considered as polymeric materials that mimic the functionality of antibodies. MIPs have been utilized for a wide variety of applications in chromatography, solid phase extraction, immunoassays, and sensor recognition. In this article, recent advances of MIPs for the extraction and analysis of ochratoxins are discussed. Selection of functional monomers to bind ochratoxin A (OTA) with high affinities, optimization of extraction procedures, and limitations of MIPs are compared from different reports. The most relevant examples in the literature are described to clearly show how useful these materials are. Strategies on MIP preparation and schemes of analytical methods are also reviewed in order to suggest the next step that would make better use of MIPs in the field of ochratoxin research. The review ends by outlining the remaining issues and impediments

Development of novel molecularly imprinted solid-phase microextraction fibers and their application for the determination of antibiotic drugs in biological samples by SPME-LC/MSn

Novel molecularly imprinted polymer (MIP)-coated fibers for solid-phase microextraction (SPME) fibers were prepared by using linezolid as the template molecule. The characteristics and application of these fibers were investigated. The polypyrrole, polythiophene, and poly(3-methylthiophene) coatings were prepared in the electrochemical polymerization way. The molecularly imprinted SPME coatings display a high selectivity toward linezolid. Molecularly imprinted coatings showed a stable and reproducible response without any influence of interferents commonly existing in biological samples. High-performance liquid chromatography with spectroscopic UV and mass spectrometry (MS) detectors were used for the determination of selected antibiotic drugs (linezolid, daptomycin, amoxicillin). The isolation and preconcentration of selected antibiotic drugs from new types of biological samples (acellular and protein-free simulated body fluid) and human plasma samples were performed. The SPME MIP-coated fibers are suitable for the selective extraction of antibiotic drugs in biological samples

ELISA-based detection of gentamicin and vancomycin in protein-containing samples

BACKGROUND: Orthopaedic implant infections are treated by surgical debridement, systematic antibiotic treatment or local antibiotic treatment with antibiotic-loaded beads. Currently antibiotic concentrations in wound exudate, serum, urine or tissue samples are determined with HPLC or fluorescent spectrometric assays. Both methods are heavily influenced due to proteins in the samples. QUESTIONS/PURPOSES: Is ELISA capable to detect gentamicin and vancomycin in protein-containing samples like serum and wound exudate. METHODS: Two specific competitive ELISA-assays were set-up to detect either gentamicin or vancomycin in protein-rich samples. An antibiotic-BSA hapten was generated as a coatable antigen and commercially available antibodies were applied for downstream immunodetection. RESULTS: The developed ELISAs perform at a detection range of 2–500 ng/ml gentamycin and 20–5000 ng/ml vancomycin. Both ELISAs were capable of detecting these antibiotics in human serum and wound exudate without being compromised by the presence of proteins. We did not detect cross-reactivity for gentamicin in the vancomycin ELISA or vice versa. CONCLUSIONS: The antibiotic ELISAs detect gentamicin and vancomycin at low concentrations in protein-rich samples and they can be used as a high throughput and cost-effective alternative for chromatographic or fluorescent methods. CLINICAL RELEVANCE: These ELISAs can be used to detect very low gentamicin or vancomycin concentrations in clinical samples or assess novel orthopaedic antibiotic release systems in in vitro and in vivo studies. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s40064-015-1411-y) contains supplementary material, which is available to authorized users