Antifouling molecularly imprinted membranes for pretreatment of milk samples: Selective separation and detection of lincomycin.

As a veterinary antibiotic, lincomycin (LIN) residues in milk are raising concerns of public on account of potential harm to human health. Efficient strategy is eagerly desired for detection of LIN from milk samples. Hence, lincomycin molecularly imprinted membranes (LINMIMs) were developed for selective separation of LIN as an efficient pretreatment of milk samples. The synergistic effect of polyethylenimine and dopamine provided effective antifouling performance by improving the hydrophilicity. Based on click chemistry, specific recognition sites were facilely formed on membranes using 4-vinylpyridine as functional monomers. The satisfactory rebinding capacity (151.62 mg g-1), permselectivity (4.43), together with the linear dependence (R2 = 0.9902) of concentrations in eluents and original samples. Moreover, the method was utilized to determine LIN from milk, with good recovery and relative standard deviation. Achievements in this work will actively promote the development of efficient detection technology.

An immunochromatographic test system for the determination of lincomycin in foodstuffs of animal origin.

The aim of this study was to develop a rapid and sensitive immunochromatographic test system for the detection of lincomycin (LIN), which belongs to the lincosamide group of antibiotics and contaminates food products of animal origin. Two formats of immunochromatographic analysis (ICA) based on different approaches of introducing gold nanoparticles (GNPs) as a label were compared. It was demonstrated that an indirect ICA method where GNPs were conjugated with anti-species antibodies allowed the achievement of both instrumental and visual detection limits of LIN almost two orders of magnitude lower than those achieved in the standard direct ICA format. In the optimized conditions, the developed indirect ICA allowed for the detection of LIN within 15 min, with instrumental and visual detection limits of 8 pg/mL and 0.8 ng/mL. The assay showed 40% cross-reactivity to clindamycin (CLIN) as a structural analogue of LIN, with no interaction with antibiotics from other classes. The developed ICA was applied for LIN detection in a panel of food products. No treatment of cow milk was necessary before the analysis. For chicken eggs and honey, a simple procedure of preliminary sample preparation was developed, which fully prevented a matrix influence on the assay results. It was demonstrated that ICA could detect LIN in food products while preserving the same analytical characteristics as in the buffer. The analytical recoveries of LIN in foodstuffs were 93.8-125% with coefficients of variations of 5.3-14.0%.

Gold Immunochromatographic Assay for Rapid On-Site Detection of Lincosamide Residues in Milk, Egg, Beef, and Honey Samples.

Lincosamides (LMs), include clindamycin (CLIN), lincomycin (LIN), and pirlimycin (PIR), that are widely used as veterinary drugs. LM residues in edible animal origin foods endanger human health and are in urgent need of establishing fast, simple, and highly sensitive detection methods. A gold immunochromatographic strip is prepared to detect CLIN, LIN, and PIR residues simultaneously with a single monoclonal antibody. This antibody is obtained with the design of a novel Hapten and can simultaneously recognize CLIN, LIN, and PIR. Under optimized conditions, the strip results can be semi-quantitatively evaluated with the naked eye within 15 min, with cut-off values in phosphate-buffered saline of 1 ng mL-1 for CLIN, 10 ng mL-1 for LIN, and 25 ng mL-1 for PIR, respectively. Besides, the strip can also be quantified using a hand-held strip scanner, and the spiked samples are used for establishing matrix curves. The limits of detection for CLIN, LIN, and PIR in spiked milk, egg, beef, and honey samples can satisfy the detection requirement. The utility of this strip is also confirmed by positive honey sample. In short, this strip should be expected to be a useful tool for the rapid on-site screening of lincosamide residues in milk, egg, beef, and honey samples.

Preparation, evaluation and application of core-shell molecularly imprinted particles as the sorbent in solid-phase extraction and analysis of lincomycin residue in pasteurized milk.

In this study, core-shell lincomycin-imprinted polymers were successfully synthesized and their binding properties evaluated. The functional monomers of methacrylamide and acrylamide were used for synthesis of core and shell, respectively. The optimum synthesized core-shell molecularly imprinted polymer (MIP) was applied as a sorbet in solid phase extraction cartridge. Afterwards, the method of core-shell molecularly imprinted solid phase extraction (CSMISPE) was used for pre-concentration and clean-up of lincomycin in the milk matrix prior to analysis via high performance liquid chromatography equipped with UV detector (HPLC-UV). The linear range for analysis of lincomycin in the milk matrix using introduced method was obtained from 0.08 to 2 µg/mL with recovery range of 80%-89%. The limit of detection and limit of quantification were 0.02 µg/mL and 0.08 µg/mL, respectively. Finally, calibrated CSMISPE-HPLC-UV method was used for lincomycin residue checking and quantification in the pasteurized milk samples of Mashhad city market.

Development and validation of a simple solid-phase extraction method coupled with liquid chromatography-triple quadrupole tandem mass spectrometry for simultaneous determination of lincomycin, tylosin A and tylosin B in royal jelly.

We have developed an analytical method for the determination of lincomycin, tylosin A and tylosin B residues in royal jelly using liquid chromatography-triple quadrupole tandem mass spectrometry analysis. For extraction and purification, we employed 1% trifluoroacetic acid and 0.1 m Na2 EDTA solutions along with an Oasis HLB cartridge. The target antibiotics were well separated in a Kinetex EVO C18 reversed-phase analytical column using a combination of 0.1% formate acid in ultrapure water (A) and acetonitrile (B) as the mobile phase. Good linearity was achieved over the tested concentration range (5-50 µg/kg) in matrix-matched standard calibration. The coefficients of determination (R2 ) were 0.9933, 0.9933 and 0.996, for tylosin A, tylosin B and lincomycin, respectively. Fortified royal jelly spiked with three different concentrations of the tested antibiotics (5, 10 and 20 µg/kg) yielded recoveries in the range 80.94-109.26% with relative standard deviations ≤4%. The proposed method was applied to monitor 11 brand of royal jelly collected from domestic markets and an imported brand from New Zealand; all the samples tested negative for lincomycin, tylosin A and tylosin B residues. In conclusion, 1% trifluoroacetic acid and 0.1 m Na2 EDTA aqueous solvents combined with solid-phase extraction could effectively complete the sample preparation process for royal jelly before analysis. The developed approach can be applied for a routine analysis of lincomycin, tylosin A and tylosin B residues in royal jelly.

Preparation of novel alkaline pH-responsive copolymers for the formation of recyclable aqueous two-phase systems and their application in the extraction of lincomycin.

Aqueous two-phase systems have potential industrial application in bioseparation and biocatalysis engineering; however, their practical application is limited primarily because the copolymers involved in the formation of aqueous two-phase systems cannot be recovered. In this study, two novel alkaline pH-responsive copolymers were synthesized and examined for the extraction of lincomycin. The two copolymers could form a novel alkaline aqueous two-phase systems when their concentrations were both 6% w/w and the pH was 8.4(±0.1)-8.7(±0.1). One copolymer was synthesized using acrylic acid, 2-(dimethylamino)ethyl methacrylate, and butyl methacrylate as monomers. Moreover, 98.8% of the copolymer could be recovered by adjusting the solution pH to its isoelectric point (pH 6.29). The other copolymer was synthesized using the monomers methacrylic acid, 2-(dimethylamino)ethyl methacrylate, and methyl methacrylate. In this case, 96.7% of the copolymer could be recovered by adjusting the solution pH to 7.19. The optimal partition coefficient of lincomycin was 0.17 at 30°C in the presence of 10 mM KBr and 5.5 at 40°C in the presence of 80 mM Ti(SO4)2 using the novel alkaline aqueous two-phase systems.

Analysis of trimethoprim, lincomycin, sulfadoxin and tylosin in swine manure using laser diode thermal desorption-atmospheric pressure chemical ionization-tandem mass spectrometry.

A new extraction method coupled to a high throughput sample analysis technique was developed for the determination of four veterinary antibiotics. The analytes belong to different groups of antibiotics such as chemotherapeutics, sulfonamides, lincosamides and macrolides. Trimethoprim (TMP), sulfadoxin (SFX), lincomycin (LCM) and tylosin (TYL) were extracted from lyophilized manure using a sonication extraction. McIlvaine buffer and methanol (MeOH) were used as extraction buffers, followed by cation-exchange solid phase extraction (SPE) for clean-up. Analysis was performed by laser diode thermal desorption-atmospheric pressure chemical-ionization (LDTD-APCI) tandem mass spectrometry (MS/MS) with selected reaction monitoring (SRM) detection. The LDTD is a high throughput sample introduction method that reduces total analysis time to less than 15s per sample, compared to minutes when using traditional liquid chromatography (LC). Various SPE parameters were optimized after sample extraction: the stationary phase, the extraction solvent composition, the quantity of sample extracted and sample pH. LDTD parameters were also optimized: solvent deposition, carrier gas, laser power and corona discharge. The method limit of detection (MLD) ranged from 2.5 to 8.3 µg kg(-1) while the method limit of quantification (MLQ) ranged from 8.3 to 28µgkg(-1). Calibration curves in the manure matrix showed good linearity (R(2)≥ 0.996) for all analytes and the interday and intraday coefficients of variation were below 14%. Recoveries of analytes from manure ranged from 53% to 69%. The method was successfully applied to real manure samples.

Role of interlayer hydration in lincomycin sorption by smectite clays.

Lincomycin, an antibiotic widely administered as a veterinary medicine, is frequently detected in water. Little is known about the soil-water distribution of lincomycin despite the fact that this is a major determinant of its environmental fate and potential for exposure. Cation exchange was found to be the primary mechanism responsible for lincomycin sorption by soil clay minerals. This was evidenced by pH-dependent sorption, and competition with inorganic cations for sorptive sites. As solution pH increased, lincomycin sorption decreased. The extent of reduction was consistent with the decrease in cationic lincomycin species in solution. The presence of Ca2+ in solution diminished lincomycin sorption. Clay interlayer hydration status strongly influenced lincomycin adsorption. Smectites with the charge deficit from isomorphic substitution in tetrahedral layers (i.e., saponite) manifest a less hydrated interlayer environment resulting in greater sorption than that by octahedrally substituted clays (i.e., montmorillonite). Strongly hydrated exchangeable cations resulted in a more hydrated clay interlayer environment reducing sorption in the order of Ca- < K- < Cs-smectite. X-ray diffraction revealed that lincomycin was intercalated in smectite clay interlayers. Sorption capacity was limited by clay surface area rather than by cation exchange capacity. Smectite interlayer hydration was shown to be a major, yet previously unrecognized, factor influencing the cation exchange process of lincomycin on aluminosilicate mineral surfaces.

Biodegradability and toxicity of pharmaceuticals in biological wastewater treatment plants.

In this experimental study both biological treatability of pharmaceuticals and their potential toxic effect in biological processes were evaluated. The pharmaceuticals were selected among those that are present at higher concentration in the Italian wastewater treatment plant effluents and widely used as antiulcer (ranitidine), beta-blocker (atenolol) and antibiotic (lincomycin). The present paper is the continuation of a work already presented,[1] which used a synthetic wastewater fed to laboratory scale SBR (Sequencing Batch Reactor) operated with different sludge ages (8 and 14 days), different biochemical conditions (aerobic or anoxic-aerobic mode) and several influent drug concentrations (2, 3 and 5 mg/L). In this case a real municipal wastewater was used as influent to the SBR. In parallel, batch tests were conducted to determine the removal kinetics of drugs and nitrogen. Toxicity tests using a titrimetric biosensor to verify possible inhibition on microorganisms were also performed. Finally, the possible adsorption of the pharmaceuticals on activated sludge was evaluated. The drugs under investigation showed different behaviours in terms of both biodegradability and toxicity effect on nitrifiers. Ranitidine showed generally low removal efficiencies (17-26%) and a chronic inhibition on nitrification. Atenolol showed generally higher removal efficiencies than ranitidine, even if the fairly good efficiency obtained in the previous experimentation with synthetic wastewater (up to 90%) was not attained with real wastewater (36%). No inhibition on nitrification was observed on both acclimated and non acclimated microorganisms with a high nitrification activity, whilst it was present with activated sludge characterised by a lower nitrification activity. Consistently with his pharmaceutical properties, lincomycin showed significant inhibition on nitrification activity.

l-3,4-Dihydroxyphenyl alanine-extradiol cleavage is followed by intramolecular cyclization in lincomycin biosynthesis.

The LmbB1 protein, participating in the biosynthesis of lincomycin, was heterologously expressed in Escherichia coli, purified in its active form, and characterized as a dimer of identical subunits. Methods for purification and analysis of the LmbB1 reaction product were developed. Molecular mass and fragmentation pattern of the product revealed by capillary electrophoresis-mass spectrometry were in agreement with its proposed structure, 4-(3-carboxy-3-oxo-propenyl)-2,3-dihydro-1H-pyrrole-2-carboxylic acid. The LmbB1 is therefore a dioxygenase catalysing the 2,3-extradiol cleavage of the l-3,4-dihydroxyphenyl alanine aromatic ring. The final LmbB1 reaction product, a unique compound found in biosynthesis of lincomycin and expected in anthramycins, arises through subsequent cyclization of the primary cleavage product, 2,3-secodopa. A possible role of LmbB1 in 2,3-secodopa cyclization and alternative ways of the cyclization in the formation of biosynthetically related compounds, muscaflavin and stizolobinic acid, are discussed.

Lincomycin, cultivation of producing strains and biosynthesis.

Lincomycin and its derivatives are antibiotics exhibiting biological activity against Gram-positive bacteria. The semi-synthetic chlorinated lincomycin derivative is used in clinical practice. The chemical structure of lincosamide antibiotics, cultivation of producing strains and analytical procedures used for separation and isolation of these compounds are described in this review. Biosynthesis of lincomycin and related compounds and its genetic control are briefly discussed.

[The application of nanofiltration membrane in the concentration and separation of lincomycin wastewater].

Two spiral nanofiltration membranes, MPS-44 (1.4 m2) and DLNF2-30 (0.24 m2), were connected in series to test the concentration process of lincomycin wastewater. Results indicated when the water inflow concentration was about 200 mg/L, the lincomycin concentration can reach 2000 mg/L after being concentrated for about 10-20 times. Such concentration can reach the demand of reuse, and the concentrating time was 60-70 h. During the concentration process, the CODCr retention was always above 80%, and the lincomycin retention was always over 90%, and the lincomycin recycle rate was over 90%.

[Production of lincomycin by Micromonospora halophytica culture]. / Obrazovanie antibiotika linkomitsina ku'lturoi Micromonospora halophytica.

A Micromonospara culture designated as 991/78 with activity against gram-positive cocci and bacteria was isolated from samples of silt-covered substrates from the Amu-Darya. Directed screening on a selective medium supplemented with lincomycin in an amount of 50-100 micrograms/ml was used. Identification of the antibiotic produced by the culture showed it to be lincomycin. By its taxonomic features the culture was classified as belonging to Micromonospora (subgroup II, Cinnamomea) and in particular to M. halophytica (Weinstein, Luedemann, Oden, Wagman, 1968). Up to now, it was known that lincomycin was produced only by Streptomyces cultures.