Free Doxycycline Research Articles

Doxycycline-Loaded Calcium Phosphate Nanoparticles with a Pectin Coat Can Ameliorate Lipopolysaccharide-Induced Neuroinflammation Via Enhancing AMPK

Neuroinflammation occurs in response to different injurious triggers to limit their hazardous effects. However, failure to stop this process can end in multiple neurological diseases. Doxycycline (DX) is a tetracycline, with potential antioxidant and anti-inflammatory properties. The current study tested the effects of free DX, DX-loaded calcium phosphate (DX@CaP), and pectin-coated DX@CaP (Pec/DX@CaP) nanoparticles on the lipopolysaccharide (LPS)-induced neuroinflammation in mice and to identify the role of adenosine monophosphate-activated protein kinase (AMPK) in this effect. The present study was conducted on 48 mice, divided into 6 groups, eight mice each. Group 1 (normal control), Group 2 (blank nanoparticles-treated), Group 3 (LPS (untreated)), Groups 4, 5, and 6 received LPS, then Group 4 received free DX, Group 5 received DX-loaded calcium phosphate nanoparticles (DX@CaP), and Group 6 received DX-loaded calcium phosphate nanoparticles with a pectin coat (Pec/DX@CaP). At the end of the experimentation period, behavioral tests were carried out. Then, mice were sacrificed, and brain tissue was extracted and used for histological examination, and assessment of interleukin-6 positive cells in different brain areas, in addition to biochemical measurement of SOD activity, TLR-4, AMPK and Nrf2. LPS can induce prominent neuroinflammation. Treatment with (Pec/DX@CaP) can reverse most behavioral, histopathological, and biochemical changes caused by LPS. The findings of the current study suggest that (Pec/DX@CaP) exerts a significant reverse of LPS-induced neuroinflammation by enhancing SOD activity, AMPK, and Nrf2 expression, in addition to suppression of TLR-4.Graphical

Enhanced efficacy against bacterial biofilms via host:guest cyclodextrin‐doxycycline inclusion complexes

Periodontal disease is characterized by a microbial infection and it is one of the major causes of teeth loss. The growth of pathogenic bacteria in oral cavity, such as Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans), provides a favorable enviromment for the biofilm formation, which result in periodontal diseases. The development of new technologies to potentiate existing drugs, avoiding bacterial biofilms resistance and improving chemical stability is highly desirable. Here, we report the use of host–guest chemistry to enhance the activity of antibacterial doxycycline (DOX) in A. actinomycetemcomitans bacterial strain suspension and biofilm in vitro through complexation with hydroxypropyl-β-cyclodextrin (HPβCD) using different molar ratios of DOX/HPβCD. 2D 1H NMR confirmed the host–guest complexation of DOX and HPβCD. We assessed the colloidal characteristics of the complex DOX/HPβCD via Dynamic Light Scattering (DLS) and Zeta Potential (PZ). The mixing ratio 1:2 DOX/HPβCD significantly decreased the minimum inhibitory concentration (MIC) and improved efficacy against A. actinomycetemcomitans suspensions and biofilms, respectively, when compared to free DOX and other DOX/HPβCD complexes. Further, the interaction of different molar ratio proportions of DOX/HPβCD complex with bacterial membrane was demonstrated via Isothermal Titration Calorimetry (ITC). Thus, we suggested the enhanced efficacy of the DOX/HPβCD complexes, at molar ratio 1:2, is due the higher cyclodextrin ratio, which potentiate the interaction between drug and bacterial membrane through nonionic interactions, such as hydrogen bonding or other van der Waals interactions. Collectively, the development of these complexes enables increased efficacy against bacterial biofilms, which hold promise for the treatment of aggressive and non-responsive forms of periodontitis.

Drug-Based Gold Nanoparticles Overgrowth for Enhanced SPR Biosensing of Doxycycline.

In clinical chemistry, frequent monitoring of drug levels in patients has gained considerable importance because of the benefits of drug monitoring on human health, such as the avoidance of high risk of over dosage or increased therapeutic efficacy. In this work, we demonstrate that the drug doxycycline can act as an Au nanoparticle (doxy-AuNP) growth and capping agent to enhance the response of a surface plasmon resonance (SPR) biosensor for this drug. SPR analysis revealed the high sensitivity of doxy-AuNPs towards the detection of free doxycycline. More specifically, doxy-AuNPs bound with protease-activated receptor-1 (PAR-1) immobilized on the SPR sensing surface yield the response in SPR, which was enhanced following the addition of free doxy (analyte) to the solution of doxy-AuNPs. This biosensor allowed for doxycycline detection at concentrations as low as 7 pM. The study also examined the role of colloidal stability and growth of doxy-AuNPs in relation to the response-enhancement strategy based on doxy-AuNPs. Thus, the doxy-AuNPs-based SPR biosensor is an excellent platform for the detection of doxycycline and demonstrates a new biosensing scheme where the analyte can provide enhancement.

Doxycycline-encapsulated solid lipid nanoparticles for the enhanced antibacterial potential to treat the chronic brucellosis and preventing its relapse: in vivo study.

BackgroundBrucellosis is one of the most important infection of diseases. Due to its large period of treatment and survival ability of bacteria inside the macrophages, relapse of this disease is the main challenge, especially, after the treatment.ObjectiveThe current study was carried out to evaluate the antibacterial effect of solid lipid nanoparticles loaded with doxycycline on the Brucella melitensis in in vivo conditions.MethodsThe double emulsion synthesized doxycycline-encapsulated solid lipid nanoparticles (DOX-SLN) was characterized using DLS and FE-SEM. The efficacy of the DOX-SLN on the acute and chronic Wistar rat infected brucellosis was investigated. The pathological assessments were made on the spleen and liver in the treated rates.ResultsThe in vivo experimental results demonstrated that the treated rats with DOX-SLN had significantly decreased the B. melitensis CFUs in their spleen and liver compared to that of the treated rates with free doxycycline and untreated ones. The pathologic results indicate that the improvement trend of spleen and liver tissues in rats treated by DOX-SLN was satisfactory.ConclusionAccording to in vivo results, the DOX-SLN has better effects on the treatment of chronic brucellosis. Therefore, DOX-SLN is recommended to treat the brucellosis and avoid its relapse.

Determination of Free Doxycycline Concentrations in the Plasma and Milk of Sheep and in the Plasma of Rabbits by using the HPLC Method

Abstract A fast and sensitive HPLC method was validated in order to analyze doxycycline in plasma and milk of sheep and in plasma of rabbits. The samples were processed with trifluoroacetic acid (TFA). After the centrifugation step, a supernatant containing extracted doxycycline and internal standard oxytetracycline was injected into the HPLC system with PDA detection. The method showed linearity in the range of 0.125 - 2.5 µg/mL for ovine plasma, 0.125 – 5.0 µg/mL for ovine milk, and 0.125 – 1 µg/mL for rabbit plasma. The inter-assay precision varied between 5.69 – 13.55 %. Values for intra-assay precision were between 0.62 – 8.67 %. Accuracy was higher than 90% in all of the tested concentrations in the three types of biological matrices. The mean extraction recovery was higher than 90 % for all matrices. In order to handle only with free drug concentrations, microfiltration of standard solutions with low (0.25mg/mL), medium (0.5mg/mL) and high (1.0mg/mL) concentration was performed. A percentage for correction of the quantified doxycycline was calculated. The most significant adjustments should be made at the low concentrations. The correction for rabbit plasma is 24.63±5.03%, for ovine plasma is 20.10±8.01% and for milk–16.68±0.04 %. This method can be used for routine determination of doxycycline concentrations for pharmacokinetic studies and further dosage adjustment.

Doxycycline-encapsulated solid lipid nanoparticles as promising tool against Brucella melitensis enclosed in macrophage: a pharmacodynamics study on J774A.1 cell line

BackgroundBrucellosis is a zoonotic disease caused by Brucella species. It has been estimated that more than 500,000 new cases of Brucellosis occur annually all around the world. Relapse of the disease is one of the most important challenges. The most important reason for the relapse of brucellosis is the survival of the bacteria inside the macrophages, which makes them safe from the immune system and disrupts drug delivery mechanism.ObjectivesThe present study was performed to assess the effects of Doxycycline-loaded Solid Lipid Nanoparticles (DOX-SLN) on the Brucella melitensis inside macrophages.MethodsDOX-SLN was prepared using double emulsion method. The technological characterization of DOX-SLN, including particle size, zeta potential, polydispersity index (PDI), drug loading and encapsulation efficiency were used. Fourier-transform infrared spectroscopy (FTIR) and Differential scanning calorimetry (DSC) were used to assess the interactions between Nanoparticles (NPs) components and crystalline form of doxycycline. Moreover, the effect of DOX-SLN on the bacteria were compared with that of the doxycycline using various methods, including well diffusion, Minimum Inhibitory Concentration (MIC), and investigation of their effects on murine macrophage-like cells cell line J774A.1.ResultsThe means of particle size, zeta potential, PDI, drug loading and encapsulation efficiency were 299 ± 34 nm, − 28.7 ± 3.2 mV, 0.29 ± 0.027, 11.2 ± 1.3%, and 94.9 ± 3.2%, respectively. The morphology of NPs were spherical with a smooth surface. No chemical reaction was occurred between the components. Doxycycline was located within NP matrix in its molecular form. The DOX-SLN significantly decreased the microbial loading within macrophages (3.5 Log) in comparison with the free doxycycline.ConclusionsSince the DOX-SLN showed better effects on B. melitensis enclosed in macrophages than the free doxycycline, it is recommended to use it for treating brucellosis and preventing relapse.

Nanoparticle-Stabilized Liposomes for pH-Responsive Gastric Drug Delivery

We report a novel pH-responsive gold nanoparticle-stabilized liposome system for gastric antimicrobial delivery. By adsorbing small chitosan-modified gold nanoparticles (diameter ~10 nm) onto the outer surface of negatively charged phospholipid liposomes (diameter ~75 nm), we show that at gastric pH the liposomes have excellent stability with limited fusion ability and negligible cargo releases. However, when the stabilized liposomes are present in an environment with neutral pH, the gold stabilizers detach from the liposomes, resulting in free liposomes that can actively fuse with bacterial membranes. Using Helicobacter pylori as a model bacterium and doxycycline as a model antibiotic, we demonstrate such pH-responsive fusion activity and drug release profile of the nanoparticle-stabilized liposomes. Particularly, at neutral pH the gold nanoparticles detach, and thus the doxycycline-loaded liposomes rapidly fuse with bacteria and cause superior bactericidal efficacy as compared to the free doxycycline counterpart. Our results suggest that the reported liposome system holds a substantial potential for gastric drug delivery; it remains inactive (stable) in the stomach lumen but actively interacts with bacteria once it reaches the mucus layer of the stomach where the bacteria may reside.

Formulation and Release Behavior of Doxycycline–Alginate Hydrogel Microparticles Embedded into Pluronic F127 Thermogels as a Potential New Vehicle for Doxycycline Intradermal Sustained Delivery

The aim of this work was the formulation and characterization of alginate (ALG)-doxycycline (DOX) hydrogel microparticles (MPs) embedded into Pluronic F127 thermogel for DOX intradermal sustained delivery. ALG-DOX MPs were formed by adding a solution of the drug into a 1.5% polymer solution while stirring. The MPs were cross-linked by dispersion into a 1.2% CaCl2 solution. Free MPs were characterized in terms of size, drug content, and release behavior by HPLC and UV-vis. DOX and hydrogel MPs were embedded into PF127, PF127-HPMC, and PF127-Methocel thermogels. The thermogels were characterized in terms of gelling time, morphology, and release behavior. A target release period of 4-7 days was considered optimal. The hydrogel MPs were about 20 microm in size with 90% of the population <59 microm. Drug content was about 35% (w/w). DOX released rapidly from the MPs, 90% within 2 days. An expected faster release was observed for free DOX from the thermogels with 80-90% of drug released after 3.5-4 h even in the presence of 1% HPMC or Methocel. The release was sustained after embedding the MPs into PF127 and PF127-HPMC thermogels. In particular, the PF127-HPMC thermogel showed an almost linear release, reaching 80% after 3 days and 90% up to 6 days. Although a further characterization and formulation assessment is required to optimize MP characteristics, ALG/DOX-loaded hydrogel MPs, when embedded into a PF127-HPMC thermogel, show a potential for achieving a 7-day sustained release formulation for DOX intradermal delivery.

In vitro cell compatibility and antibacterial activity of microencapsulated doxycycline designed for improved localized therapy of septic arthritis

For the treatment of septic arthritis in large animals, the local application of antibiotics as a slow release system may be an appropriate means to reach high local bioactivity and low systemic side effects and drug residues. In this study, doxycycline microspheres were developed and tested in vitro for their drug-release properties, suitability for intra-articular application and antimicrobial activity. The development of a slow release system was achieved by microencapsulation of the drug into poly(lactide-co-glycolide) microspheres by a novel ultrasonic atomization method. Drug elution was evaluated from microspheres dispersed in elution medium at pre-defined time points by HPLC. Joint-tissue compatibility was tested on cultured bovine synoviocytes by evaluating the expression of pro-inflammatory cytokine mRNA and the production of nitric oxide (NO). Finally, the antimicrobial activity of the released antibiotic was assessed with gram-negative and gram-positive bacteria exposed to release medium sampled at days 1, 7 and 12 after microsphere suspension. An adequate size of the microspheres, sufficient stabilization of doxycycline in aqueous environment and drug release (25 mg microspheres in 4 mL medium) above MIC for bacteria usually isolated in bovine and equine joints were obtained over 15 days. Although the cytokine mRNA expression reflected the excellent tissue compatibility, the results with NO yielded contradictory results. Antimicrobial tests of the release medium proved to match perfectly the activity of non-encapsulated, free doxycycline as reported in the literature. The newly developed doxycycline delivery system achieved the target specifications and is ready for in vivo testing.

Effects of cationic liposome-encapsulated doxycycline on experimental Chlamydia trachomatis genital infection in mice.

In a previous study, we demonstrated that cationic liposome-encapsulated doxycycline (CaL-Dox) was two-fold more effective than free doxycycline against Chlamydia trachomatis in vitro. Here, we evaluated the effects of two CaL-Dox regimens in comparison with unencapsulated doxycycline on the course of chlamydial genital infection in mice. Progesterone-treated, female CF-1 mice were challenged intravaginally with 1.2 x 10(5) inclusion-forming units (ifu) of C. trachomatis. Two days post-infection, the animals were divided into four treatment groups for im injection of doxycycline at 10 mg/kg body weight bd for 3 (3 Dox) or 7 days (7 Dox), or of CaL-Dox at the same dose level for 3 (3 CaL-Dox) or 7 days (7 CaL-Dox) consecutively. An infected fifth group served as a control and was given an empty CaL preparation. C. trachomatis were isolated after five blind passages from 82% of infected control mice, 61.4% of 3 Dox, 52.2% of 3 CaL-Dox, 29% of 7 Dox and 20% of 7 CaL-Dox animals. Histopathological reactions were found in various tissues of the genital tract in 79.5% of infected control mice, 80.9% of 3 Dox, 65.2% of 3 CaL-Dox, 47.1% of 7 Dox and 25.7% of 7 CaL-Dox animals. Total antichlamydial antibody titres were lower in 7 CaL-Dox mice than in all the other groups (P < 0.005). The results showed that progesterone-treated CF-1 mice are suitable for investigation of both lower and upper genital tract infection with a lymphogranuloma venereum biovar strain of C. trachomatis. Neither 7 CaL-Dox nor 3 CaL-Dox treatment was more effective than unencapsulated 7 Dox doses in the bacteriological cure of chlamydial genital infection in mice. However, 7 CaL-Dox prevented tissue damage in the genital tract significantly more than all the other regimens (P < 0.05). These results suggest that liposome-encapsulated doxycycline, particularly CaL-Dox, may have potential for the clinical treatment of chlamydial infections.

Allometric analysis of doxycycline pharmacokinetic parameters

Doxycycline serum concentration-time profiles and elimination half-lives in several species were evaluated by allometric techniques. Doxycycline elimination half-life increased with body weight. Interspecies scaling of total or free doxycycline serum concentration-time profiles was possible by use of Dedrick plots with the exception of doxycycline total concentrations in cats. The observed difference in cats was attributed to more extensive binding to plasma proteins.

Different effects of oxytetracycline and doxycycline on mitochondrial protein synthesis in rat liver after long-term treatment

The tetracyclines inhibit specifically mitochondrial (mt) and bacterial protein synthesis when they are present in low concentrations (2–10 μg/ml). There is no difference between the various members of this group of antibiotics in this respect. In the present study, however, it is shown that the inhibitory effect of doxycycline on mt-protein synthesis in rat liver is partially lost after continuous treatment for more than 1 week, whereas oxytetracycline continues to inhibit mt-protein synthesis effectively after 1 week of treatment. To find an explanation for this difference between doxycycline and oxytetracycline, a detailed study was made of the distribution and the effects on mt-protein synthesis of both tetracyclines under various conditions in rat liver. The results of the studies lead to the hypothesis that doxycycline treatment induces the formation of a doxycycline complex, and thus to a reduced amount of free doxycycline. This may explain the loss of effective inhibition of mt-protein synthesis.