Development of a multicomponent immunochromatographic test system for the detection of fluoroquinolone and amphenicol antibiotics in dairy products.

BACKGROUND: Ciprofloxacin (CIP) and chloramphenicol (CAP) are relevant antibiotics of the fluoroquinolone (FQ) and amphenicol (AP) groups, respectively, widely used in veterinary practice and they contaminate agricultural products. In this study, a rapid and sensitive immunochromatographic assay (ICA) was developed for simultaneous detection of CIP and CAP in dairy products. The ICA was carried out in a direct competitive format using gold nanoparticles as a label. RESULTS: The ICA developed here allowed for the detection of CIP and CAP in Triton X-100-containing buffered saline (PBST) within 15 min with instrumental detection limits of 20 pg mL-1 and 0.5 ng mL-1 , respectively, and with a visual detection limit of 5 ng mL-1 for both antibiotics. The ICA showed cross-reactivity (69-160%) to 19 antibiotics in the FQ group and no cross-reactivity (<0.1%) to 2 antibiotics of the AP group. The ICA allowed detection of CIP and CAP in a panel of dairy products by employing a simple procedure of preliminary sample preparation. The detection limits for the two antibiotics were the same as in PBST. The analytical recoveries of CIP and CAP in dairy products ranged from 83% to 120%. CONCLUSION: The analytical characteristics of the test system allow its use for the detection of antibiotics in milk and dairy products during all steps of production. © 2019 Society of Chemical Industry.

Trigonal layered rosiaite-related antiferromagnet MnSnTeO6: ion-exchange preparation, structure and magnetic properties.

Ion-exchange treatment of Na2SnTeO6 in molten salt mixtures resulted in rosiaite (PbSb2O6)-related MnSnTeO6. Its crystal structure was refined by the Rietveld method. Of the three possible models of Sn/Te distribution, the disordered model (P3[combining macron]1m, a = 5.23093(11) Å, c = 4.62430(16) Å) was preferred based on bond distances. However, it is supposed that each individual (SnTeO6)2- layer retains complete ordering of the precursor and the apparent disorder is only due to stacking faults. Magnetic studies have shown that MnSnTeO6 orders antiferromagnetically at Neél temperature TN = 9.8 K. The effective magnetic moment reasonably agrees with theoretical estimations assuming high-spin configuration of Mn2+ (S = 5/2). Electron spin resonance reveals spin dynamics in accordance with antiferromagnetic ordering. Moreover, critical broadening of ESR linewidth indicates the low-dimensional type of exchange interactions. Based on the temperature and field-dependent magnetization studies, the magnetic phase diagram of the new compound was constructed.

Preparation and characterization of metastable trigonal layered MSb2O6 phases (M = Co, Ni, Cu, Zn, and Mg) and considerations on FeSb2O6.

MSb2O6 compounds (M = Mg, Co, Ni, Cu, Zn) are known in the tetragonal trirutile forms, slightly distorted monoclinically with M = Cu due to the Jahn-Teller effect. In this study, using a low-temperature exchange reaction between ilmenite-type NaSbO3 and molten MSO4-KCl (or MgCl2-KCl) mixtures, these five compositions were prepared for the first time as trigonal layered rosiaite (PbSb2O6)-type phases. Upon heating, they irreversibly transform to the known phases via amorphous intermediates, in contrast to previously studied isostructural MnSb2O6, where the stable phase is structurally related to the metastable phase. The same method was found to be applicable for preparing stable rosiaite-type CdSb2O6. The formula volumes of the new phases show an excellent correlation with the ionic radii (except for M = Cu, for which a Jahn-Teller distortion is suspected) and are 2-3% larger than those for the known forms although all coordination numbers are the same. The crystal structure of CoSb2O6 was refined via the Rietveld method: P3[combining macron]1m, a = 5.1318(3) Å, and c = 4.5520(3) Å. Compounds with M = Co and Ni antiferromagnetically order at 11 and 15 K, respectively, whereas the copper compound does not show long-range magnetic order down to 1.5 K. A comparison between the magnetic behavior of the metastable and stable polymorphs was carried out. FeSb2O6 could not be prepared because of the 2Fe2+ + Sb5+ = 2Fe3+ + Sb3+ redox reaction. This electron transfer produces an additional 5s2 shell for Sb and results in a volume increase. A comparison of the formula volume for the stable mixture FeSbO4 + 0.5Sb2O4 with that extrapolated for FeSb2O6 predicted that the trirutile-type FeSb2O6 can be stabilized at high pressures.

Orbitally induced hierarchy of exchange interactions in the zigzag antiferromagnetic state of honeycomb silver delafossite Ag3Co2SbO6.

We report the revised crystal structure, static and dynamic magnetic properties of quasi-two dimensional honeycomb-lattice silver delafossite Ag3Co2SbO6. The magnetic susceptibility and specific heat data are consistent with the onset of antiferromagnetic long range order at low temperatures with Néel temperature TN ∼ 21.2 K. In addition, the magnetization curves revealed a field-induced (spin-flop type) transition below TN in moderate magnetic fields. The GGA+U calculations show the importance of the orbital degrees of freedom, which maintain a hierarchy of exchange interaction in the system. The strongest antiferromagnetic exchange coupling was found in the shortest Co-Co pairs and is due to direct and superexchange interaction between the half-filled xz + yz orbitals pointing directly to each other. The other four out of six nearest neighbor exchanges within the cobalt hexagon are suppressed, since for these bonds the active half-filled orbitals turned out to be parallel and do not overlap. The electron spin resonance (ESR) spectra reveal a broad absorption line attributed to the Co(2+) ion in an octahedral coordination with an average effective g-factor g = 2.40 ± 0.05 at room temperature and show strong divergence of the ESR parameters below ∼150 K, which implies an extended region of short-range correlations. Based on the results of magnetic and thermodynamic studies in applied fields, we propose a magnetic phase diagram for the new honeycomb-lattice delafossite.

[Fluorescence polarization immunoassay of ractopamine].

A technique was developed for fluorescence polarization immunoassay (FPIA) of ractopamine, a toxic low molecular weight nonsteroidal growth regulator belonging to the most controlled contaminants of food products of animal origin. The assay is based on the competition between a sample containing ractopamine and ractopamine­fluorophore conjugate for binding to antibodies. The competition is monitored via changes in the degree of fluorescence polarization for plane-polarized excitation light, which differs for the free and antibody-bound forms of the conjugate. The optimal assay conditions were established, ensuring a high accuracy and minimal detection limit. The developed assay demonstrated a detection limit of 1 ng/mL and a range of detectable concentrations of 2.3­50 ng/mL, which met the requirements of sanitary control. The duration of the analysis was 10 min. The possible application of the developed FPIA was demonstrated with testing of turkey meat. The speed and simplicity of the proposed assay define its efficiency as a screening tool for safety of foods.

Fluorescence Polarization Immunoassay, for Express Control of Antibiotic Levels: Design and Characteristics for Chloramphenicol, as an Example.

Characteristics of the fluorescence polarization immunoassay (FPIA) as a mean for express control of antibiotic levels in various specimens and its advantages vs. other analytical tests are described. The developmental stages of the analytical procedure and its parameters are considered for chlorampnenicol as an example. The analysis is based on competitive interaction of anti-chloramphenicol antibodies with the chloramphenicol-fluorophore conjugate and the potential free chloramphenicol in the specimen. The experimental results of the comparison of the chloramphenicol FPIA with the use of two conjugates differing in the length of the bridge length between the antibiotic functional groups and fluorophore (fluorescein) are presented. The requirements to the choice of the antibody and conjugate concentrations providing highly sensitive detection are characterized. The detection limit of chloramphenicol in the FPIA was 10 ng/ml and the determination of the concentrations ranged from 20 ng/mI to 10 mcg/ml. The time of the assay was 10 min.

Highly Sensitive Immunochromatographic Identification of Tetracycline Antibiotics in Milk.

A rapid immunochromatographic assay was developed for the control of tetracycline (TC). The assay is based on the competition between immobilized TC-protein conjugate and TC in a tested sample for binding with polyclonal anti-TC antibodies conjugated to colloidal gold during the flow of the sample along a membrane strip with immobilized reactants. Conjugation of colloidal gold and the total immunoglobulin (IgG) fraction of polyclonal antibodies was used to increase the assay sensitivity to ensure low content of specific antibodies in the conjugate. This allowed effective inhibition of free TC and conjugate binding in the strip test zone. Photometric marker registration allows control of the reduction of binding, thereby enhancing detection sensitivity. The proposed assay allows TC to be detected at concentrations up to 20 ng/mL, exceeding the limit of detection of the known analogues, in a wide working range (more than two orders) of 60 pg/mL to 10 ng/mL, ensured through the use of polyclonal antibodies. The assay time is 10 min. The efficiency of the designed assay is shown to identify TC in milk; the degree of recovery of TC ranges from 90 to 112%. The precision of the concentrations measurements was no more than 10%.

A new layered triangular antiferromagnet Li4FeSbO6: spin order, field-induced transitions and anomalous critical behavior.

Structure, electrochemical, magnetic and resonance properties of new layered antimonate Li(4)FeSbO(6) were comprehensively studied using powder X-ray diffraction, cyclic voltammetry, magnetic susceptibility, heat capacity, electron spin resonance and Mössbauer spectroscopy. In the crystal structure the iron ions form the triangular network within (LiFeSbO(6))(3-) layers alternating with nonmagnetic lithium layers. The electrochemical activity studied implies an Fe(3+)/Fe(4+) redox couple at 4.3 V (ox.) and 3.9 V (red.) thereby revealing that Li can be reversibly extracted. The long-range antiferromagnetic order was found to occur at the Néel temperature, T(N) ≈ 3.6 K, confirmed both by the magnetic susceptibility data and specific heat ones. The effective magnetic moment is estimated to be 5.93 µ(B)/f.u. and satisfactorily agrees with theoretical estimations assuming high-spin configuration of Fe(3+) (S = 5/2). In the magnetically ordered state, though, the magnetization demonstrates rather peculiar behavior. An additional anomaly on the M(T) curves appears at T(2) < T(N) in moderate magnetic field. The positions of transitions at T(N) and T(2) separate increasingly with increasing external field. Multiple measurements consistently demonstrated field-sensitive moving of magnetic phase boundaries constituting a unique phase diagram for the compound under study. The complex low-dimensional (2D) nature of magnetic coupling was confirmed by the dynamic magnetic properties study. Electron spin resonance from Fe(3+) ions in paramagnetic phase is characterized by a temperature independent effective g-factor of 1.99 ± 0.01. However, the distortion and broadening of the ESR line were found to take place upon approaching the magnetically ordered state from above. The divergence of the temperature-dependent linewidth is analyzed in terms of both critical behavior close to long-range magnetic order and the Berezinskii-Kosterlitz-Thouless (BKT)-type transition. Heat capacity measurements even at zero field manifested an appearance of the additional anomaly at temperatures below the Néel temperature. The temperature dependence of ESR intensity, linewidth and shift of the resonant field imply an extended region of short-range order correlations in the compound studied. The rich variety of the anomalies in magnetic and resonance properties makes this new antimonate a very interesting system to investigate the multiple phase transitions and competing exchange interaction due to the critical role of the layered structure organization accompanied by the frustration effects in triangular antiferromagnets.

Pretreatment-free immunochromatographic assay for the detection of streptomycin and its application to the control of milk and dairy products.

A rapid pretreatment-free immunochromatographic assay was developed for the control of the streptomycin (STR) content in milk and dairy products. The assay is based on the competition between an immobilized STR-protein conjugate and STR in a sample to be tested for the binding to monoclonal anti-STR antibodies conjugated to colloidal gold during the flow of the sample along a membrane strip with immobilized reactants. It is possible to improve the cut-off level of positive and negative samples distinguished by a change in the molar STR to protein ratio in the immobilized conjugate. The cut-off level (500 ng mL(-1)) thus achieved corresponds to the stated MRL of STR in milk and dairy products. For STR concentrations in the range of 16-250 ng mL(-1) its content can be quantitatively measured based on the degree of binding of a colloidal gold label in the test strip zone with the immobilized STR-protein conjugate. The duration of the assay is 10 min. The selected sizes of membrane pores and colloidal gold particles allow the assay to be carried out at room temperature without additional reactants and pretreatment. The applicability of the assay for milk, whole milk, sour clotted milk, and kefir with different fat content (from 0.5% to 6%) was confirmed. The results of quantitative immunochromatographic assay show good correlation with traditional ELISA (r was equal to 0.935 and 0.940 for the series tested).

[Oxygen-dependent processes in the blood cells from children with arterial hypertension concomitant with biliary dyskinesia].

Age individuality is characterized by an imbalance of the molecular mechanisms of antioxidant defense in adolescents with arterial hypertension and biliary dyskinesia, as documented by an enzyme imbalance of the first line of antioxidant defense and H2O, accumulation, by a substantial increase in glutathione peroxidase activity, and by inhibition of the activity of glutathione-dependent enzymes. The considerable rise of 2,3-diphosphoglycerate suggests tissue hypoxia. With this, enhanced neutrophil elastase activity causes damage to the structural components of vascular wall connective tissue, resulting in the development of endothelial dysfunction.

[Immunochromatographic technique for express determination of ampicillin in milk and dairy products].

An immunochromatographic method for determination of beta-lactam antibiotic ampicillin has been developed. The method is based on the competitive interaction between antibiotic molecules contained in the sample and protein conjugate of penicillin immobilized on a membrane for binding with specific antibodies labeled with colloidal gold, which occurs during movement of the sample to be tested and reagents along the membrane. The completion of the test system ensures control of exceeding the maximum permissible content of the antibiotic in milk and dairy products (10 ng/ml). The possibility of testing milk, raw milk, and dairy products for 10 minutes at room temperature without sample preparation has been demonstrated.

Rapid pretreatment-free immunochromatographic assay of chloramphenicol in milk.

A pretreatment-free immunochromatographic assay for detection of chloramphenicol (CAP) in milk was developed. The assay is based on competition between CAP molecules in the sample and immobilized CAP-protein conjugate for binding to monoclonal anti-CAP antibodies conjugated with colloidal gold particles (average diameter 30nm). The assay is carried out in the course of sample flowing along test strip with immobilized reactants, and its results can be detected by the naked eye or by a photometric device. Effect of the concentration of immunoreactants on assay characteristics was studied. The assay protocol with maximal sensitivity and reliability was optimized using measured values of brightness of lines. Detection limit for CAP is 10ngmL(-1). Assay duration is 10min, and it can be carried out at room temperature without any additional devices and reactants. The developed test strip has been applied to CAP detection in dairy products.

Cellulase activity of a haloalkaliphilic anaerobic bacterium, strain Z-7026.

The cellulolytic activity of an alkaliphilic obligate anaerobic bacterium, Z-7026, which was isolated from the microbial community of soda-lake sediments and belongs to the cluster III of Clostridia with low G+C content, was studied. The bacterium was capable of growing in media with cellulose or cellobiose as the sole energy sources. Its maximal growth rate on cellobiose (0.042-0.046 h(-1)) was observed at an initial pH value of 8.5-9.0, whereas the maximal rate of cellulase synthesis, assayed by using a novel fluorimetric approach, was found to be 0.1 h(-1) at pH 8-8.5. Secreted proteins revealed high affinity for cellulose and were represented by two major forms of molecular masses of 75 and 84 kDa, whereas the general protein composition of the precipitated and cellulose-bound preparations was similar to cellulosome subunits of Clostridium thermocellum. The optimum pH of the partially purified enzyme preparation towards both amorphous and crystalline cellulose was in the range 6-9, with more than 70% and less than 50% of maximal activity being retained at pH 9.2 and 5.0, respectively.

[Effect of antioxidants on regeneration of protoplasts in the mycelial fungus Trichoderma reesei 6/16]. / Vliianie antioksidantov na regeneratsiiu protoplastov mitselial'nogo griba Trichoderma reesei 6/16.

The relative content of antioxidants in the mycelium of Trichoderma reesei 6/16 obtained by propagation of fungal protoplasts was shown to decrease (as compared to the initial culture taken for preparation of protoplasts) and restored only in the second generation of regenerated mycelium. In this respect, the effects of various antioxidants (beta-carotene, ascorbic acid, alpha-tocopherol, and ionol) on the frequency of regeneration of T. reesei 6/16 protoplasts were studied. beta-Carotene increased the viability of fungal protoplasts to the greatest extent. The effect of ascorbic acid depended on the presence of Fe ions. Ionol did not cause any measurable protective effect.