Objectives. Thermal stability is a key property in determining the suitability of an antibiotic agent for local application in the treatment of orthopaedic infections. Despite the fact that long-term therapy is a stated goal of novel local delivery carriers, data describing thermal stability over a long period are scarce, and studies that avoid interference from specific carrier materials are absent from the orthopaedic literature. Methods. In this study, a total of 38 frequently used antibiotic agents were maintained at 37°C in saline solution, and degradation and antibacterial activity assessed over six weeks. The impact of an initial supplementary heat exposure mimicking exothermically curing bone cement was also tested as this material is commonly used as a local delivery vehicle. Antibiotic degradation was assessed by liquid chromatography coupled to mass spectrometry, or by immunoassays, as appropriate. Antibacterial activity over time was determined by the Kirby-Bauer disk diffusion assay. Results. The heat exposure mimicking curing bone cement had minimal effect on stability for most antibiotics, except for gentamicin which experienced approximately 25% degradation as measured by immunoassay. Beta-lactam antibiotics were found to degrade quite rapidly at 37°C regardless of whether there was an initial heat exposure. Excellent long-term stability was observed for aminoglycosides, glycopeptides, tetracyclines and quinolones under both conditions. Conclusions. This study provides a valuable dataset for orthopaedic surgeons considering local application of antibiotics, and for material scientists looking to develop next-generation controlled or extended-release antibiotic carriers. Cite this article: E. Samara, T. F. Moriarty, L. A. Decosterd, R. G. Richards, E. Gautier, P. Wahl. Antibiotic stability over six weeks in aqueous solution at body temperature with and without heat treatment that mimics the curing of bone cement. Bone Joint J 2017;6:296–306. DOI: 10.1302/2046-3758.65.BJR-2017-0276.R1

Introduction. Collagen is the predominant component of extracellular matrix in various connective tissues and makes up to 25% to 35% of the whole protein content in animal bodies. Type II collagen was first introduced from chicken sternal cartilage and presents supportive function in cartilaginous tissue. Since type II collagen is the major component of cartilage in joint, this study is aiming to determine an optimal type II collagen material for the development of medical devices for articular cartilage regeneration. In order to make more effective use of underutilized food waste, type II collagens from mammalian tissue sources (porcine tracheal cartilage; auricular cartilage; articular cartilage) and marine tissue sources (cuckoo ray, blonde ray, thorn back ray, lesser spotted dogfish) were isolated through acid-pepsin digestion under 4°C and characterized by various biological, biochemical and biophysical analysis. Pepsin cleaves the telopeptide region of the collagen molecule and pepsin treated collagen extraction ensures higher collagen yield. Telopeptide-free collagen reveals cytocompatibility, biodegradability and lower toxicity. The number and size of collagen chains were revealed by SDS-polyacrylamide gel electrophoresis. Intermolecular crosslinking density was quantified by Ninhydrin assay. Thermal stability was tested by differential scanning calorimetry (DSC) and enzymatic degradation was assessed by collagenase assay. Human chondrocytes were seeded on to collagen sponges at a density of 30,000 cells per sponge. Cell morphology (DAPI/ Rhodamine Phalloidin), viability(LIVE/DEAD®), proliferation(PicoGreen®) and metabolic activity (alamarBlue®) were analysed. Quantitative morphometric analysis was carried out using ImageJ software. Conclusion. Porcine articular cartilage and cartilaginous fishes yield high purity type II collagen. Type II collagen isolated from cartilaginous fishes exhibited similar crosslinking density and thermal stability. Among various porcine cartilaginous tissues, articular cartilage was the most resistant to enzymatic degradation and female trachea exhibited the highest cross-linking density. The biological, biochemical and thermal properties of type II collagen are dependent on the tissue and gender from which the collagen was extracted

Aim. Thermal stability is a key property determining the suitability of an antibiotic agent for local application. Long-term data describing thermal stability without interference from carrier materials are scarce. Method. In this study, a total of 38 common antibiotic agents have been maintained at 37 °C in saline solution, and degradation and antibacterial activity assessed over 6 weeks. The impact of an initial supplementary heat exposure mimicking exothermically-curing bone cement has also been tested. Antibiotic degradation was assessed by chromatography coupled to mass spectrometry or immunoassays, as appropriate. Antibacterial activity was determined by Kirby-Bauer disk diffusion assay. Results. The heat exposure mimicking curing bone cement had minimal effect on stability for most antibiotics, except for gentamicin, which experienced approximately 25% degradation as measured by immunoassay. Beta-lactam antibiotics were found to degrade quite rapidly at 37°C regardless of whether there was an initial heat exposure or not. However some of them maintained relevant concentrations and activity for 2–3 weeks, particularly aztreonam. Excellent long-term stability was observed for aminoglycosides, glycopeptides, tetracyclines and quinolones under both conditions. Conclusions. This study provides a valuable dataset for orthopaedic surgeons considering local application of antibiotics. For example, tobramycin would be more suitable for application with bone cement than gentamicin, as it was found to be resistant to heat exposure mimicking curing bone cement

Sprains and strains result from collagen fibre overextension. This study investigated changes in the molecular state of collagen due to overextension damage, thereby gaining insight into tissue degeneration and cellular detection of damage. Overextension results in intermolecular and intrafibrillar sliding, detected with x-ray diffraction. Tendon rupture results in increased susceptibility to proteolytic enzymes. These observations and contemporary theory concerning collagen fibre stability lead to the hypothesis that sub-rupture overextension should result in reduced thermal stability of fibrous collagen. Tendons were harvested from steer tails. Each provided a specimen for control and for overextension. Sub-rupture overextension at 1%/s strain rate was accomplished on a mechanical testing system, under the control of custom software, until the slope of the force-deformation curve was approximately zero (before complete failure). Two loading treatments were tested: one-cycle and five-cycles. Two specimen types were tested: native tendons ± NaBH4 crosslink stabilization. Tendons in each of the four groups (2x2) were paired by originating tail. Thermal stability was assessed in terms of denaturation temperature (Td) using hydrothermal isometric tension testing. Specimens were held at constant length and heated from ambient temperature to 90degC. Td was defined as the temperature where load suddenly increased due to molecular unraveling and attempted shrinkage. Overextension of native specimens reduced the thermal stability of the collagen (p< 0.0001) and five-cycles had a still greater effect (p=0.03). Td of controls was 64.5±1.0degC (mean±SD). After one-cycle, Td dropped to 63.2±1.0degC and, after five-cycles, Td dropped to 61.8±2.0degC. For stabilised tendons, the effect of multiple cycles was lost (p=0.08) but overstretching decreased Td by ~2degC (p< 0.0001). This study confirms that the molecular state of collagen is altered by overextension damage, reducing Td by up to 10% of the expected range (37–65degC) in our experiments. This is thought to occur due to intermolecular sliding that liberates specific domains on the molecules, lowering the activation energy for uncoiling. These domains may also be key targets in degeneration and cell-collagen signaling

Alpha Lipoic Acid (L.A.) is an effective natural antioxidant discovered in the human body in 1951 from L.J. Reed and I.C. Gunslaus from liver. It is inside broccoli, spinach and red meats, especially liver and spleen. Actually it is largely used as antioxidant in antiaging products according to the low toxicity level of the product. The present study take into consideration the possibility to reduce oxidation of medical irradiated UHMWPE GUR 1050, mixing together polyethylene powder and Alpha Lipoic Acid powder. The study is composed of two parts. Part 1 Thermostability of alpha lipoic acid during polyethylene fusion Part 2 detection of oxygen level in artificially aged irradiated polyethylene. Solid pieces were made with Gur 1050 powder (Ticona Inc., Bayport, Tex, USA) and mixed with Alpha Lipoic Acid (Talamonti, Italy, Stock 1050919074) 0,1%-­‐0,3%-­‐0,5%-­‐1%-­‐2% and gamma ray irradiated with 30 kGy (Isomedix, Northborough, MA). An owen (80° Celsius) was used to produce an aging effect for 35 days in the doped and control samples (Conventional not doped polyethylene). This process simulate an aging effect of 10 years into the human body. Part 1 : THERMAL STABILITY : a Fourier Transfer Infra Red (FTIR) test was made in pieces molded in a cell at 150° and 200°Celsius and pressure of 200 MPa comparing to the UHMWPE powder mixed with alpha lipoic acid. The presence of Alpha Lipoic Acid in the polyethylene was found at any depth in the manufacts. Part 2 : OXIDATION OF THE SURFACE : After 5 weeks at 80° Celsius in a owen (ASTM standard F-­‐2003-­‐02)A FOURIER TRANSFER INFRA RED TEST (FTIR) was made in the superficial layer and deeper on the undersurface of doped 0,1% and conventional UHMWPE. The antioxidation limit is defined as the ratio of the area under 1740cm/−­‐1 carbonyl and 1370 cm/−­‐1 Methylene absorbance peaks. In conventional UHMWPE oxidation is detected on the surface and decreases in the deeper layers down to zero under 1500 Micron. In the doped UHMWPE, FTIR demonstrate a very low oxidation limit on the surface and at any depth, comparing to conventional UHMWPE. The examples show that Lipoic Acid is effective as antioxidant in irradiated UHMWPE and it is stable with respect to thermal treatment. For any figures or tables, please contact authors directly

Porcine and fish by-products in particular are rich sources for collagen, which is the main component of the extracellular matrix (ECM). Although there are studies investigating different collagen derived from various tissue sources for the purpose of creating biomaterials, the comparison of biophysical, biochemical and biological properties of type II collagen isolated from cartilaginous tissues has yet to be assessed. In addition, it has been shown from previous studies that sex steroid hormones affect the collagen content in male and female animals, herein, type II collagens from male and female porcine cartilage were assessed in order to investigate gender effects on the property of collagen scaffolds. Moreover, type II collagen has a supportive role in articular cartilage in the knee joint. Therefore, the aim is to assess the properties of type II collagen scaffolds as a function of species, tissue and gender for cartilage regeneration. Type II collagen was extracted from male and female porcine trachea, auricular, articular cartilage and cartilaginous fish through acid-pepsin digestion at 4°C. SDS-PAGE was conducted to confirm the purity of extracted collagen. Collagen sponges were created via freeze-drying. Scaffold structure and pore size were evaluated by scanning electron microscopy (SEM). Thermal stability was assessed by differential scanning calorimetry (DSC). Sponges were seeded with human adipose derived stem cells to assess chondro-inductive potential of collagen sponges after 7, 14 and 21 days of culture. In conclusion, collagen sponges support the proliferation and differentiation of human adipose derived stem cells to different extents

Thermostability is a key property in determining the suitability of local delivery of antibiotics in the treatment of orthopaedic infections. Herein, we aimed to assess the thermal stability and antibacterial activity of ciprofloxacin, ceftriaxone, gentamycine and vancomycine in high temperature conditions. Using a standardized E-test method, minimally inhibited concentration of each antibiotic substance against Staphylococcus aureus cultures were determined. The solutions of antimicrobial drugs ciprofloxacin 2 mg/ml, ceftriaxone 200 mg/ml, gentamycine 40 mg/ml and vancomycine 200 mg/ml were diluted twofold in deionised water. Acquired solutions were divided into three aliquots. The first aliquot was held at 40°C for 30 min in a waterbath, the second and the third aliquots were exposed to 80 and 100°C for 30 min in hot-air sterilizer, respectively. The treated solutions were tested for residual activity against S. aureus using a standardized disk diffusion method. Mediums with untreated antibiotic solutions and S. aureus were used as control. Plates were incubated at 37°C, at which time zones of inhibition (ZoI) were measured to the nearest whole millimeter for 14 days. The investigation indicated that the temperature elevation impacted considerably on antimicrobial activity and antibiotic stability overall. The in vitro temperature-response curves showed that ZoI diameter decreases logarithmically with elevated temperatures. Gentamicin was the only drug that was found to be affected to some extent. Results from the study provides a valuable dataset for orthopaedic surgeons considering local application of antibiotics and methods of antibiotic impregnation

Alpha Lipoic Acid (L.A.) is an effective natural antioxidant discovered in the human body in 1951 from L.J. Reed and I.C. Gunslaus from liver. It is inside broccoli, spinach and red meats, especially liver and spleen. Actually it is largely used as antioxidant in antiaging products according to the low toxicity level of the product. The present study take into consideration the possibility to reduce oxidation of medical irradiated UHMWPE GUR 1050, mixing together polyethylene powder and Alpha Lipoic Acid powder. The study is composed of two parts. Part 1 Thermostability of alpha lipoic acid during polyethylene fusion. Part 2 detection of oxygen level in artificially aged irradiated polyethylene. Solid pieces were made with Gur 1050 powder (Ticona Inc., Bayport, Tex, USA) and mixed with Alpha Lipoic Acid (Talamonti, Italy, Stock 1050919074) 0, 1% and gamma ray irradiated with 30 kGy (Isomedix, Northborough, MA). An oven (80° Celsius) was used to produce an aging effect for 35 days in the doped and control samples (Conventional not doped polyethylene). This process simulate an aging effect of 10 years into the human body. THERMAL STABILITY: a Fourier Transfer Infra Red (FTIR) test was made in pieces molded in a cell at 150° and 200°Celsius and pressure of 200 MPa comparing to the UHMWPE powder mixed with alpha lipoic acid. The presence of Alpha Lipoic Acid in the polyethylene was found at any depth in the manufacts. figure 1: A Pure Lipoic Acid. B Lipoic Acid + UHMWPE melted 150° C. Lipoic Acid + UHMWPE melted 200° C° (A And B spectra subtracted UHMWPE). OXIDATION: After 5 weeks at 80° Celsius in a oven (ASTM standard F-2003-02)A FOURIER TRANSFER INFRA RED TEST (FTIR) was made in the superficial layer and deeper on the undersurface of doped 0.1% and conventional UHMWPE. The antioxidation limit is defined as the ratio of the area under 1740 cm/−1 carbonyl and 1370 cm/−1 Methylene absorbance peaks. In conventional UHMWPE oxidation is detected on the surface and decreases in the deeper layers down to zero under 1500 Micron Fig 2. figure 2 Pure polyethylene: A Surface, B 1500 Micron, C 3000 Micron. In the doped UHMWPE, FTIR demonstrate a very low oxidation limit on the surface and at any depth, comparing to conventional UHMWPE Fig 3. figure 3 Doped UHMWPE A surface, B 700 micron deept, C 1700 micron deept. The examples show that Lipoic Acid is effective as antioxidant in irradiated UHMWPE and it is stable with respect to thermal treatment

Aims

To explore the effect of different types of articulating antibiotic-loaded cement spacers in two-stage revision for chronic hip prosthetic joint infection (PJI).

Objectives

The optimal protocol for antibiotic loading in the articulating cement spacers for the treatment of prosthetic joint infection (PJI) remains controversial. The objective of the present study was to investigate the effectiveness of articulating cement spacers loaded with a new combination of antibiotics.

Objectives

The optimal protocol for antibiotic loading in the articulating cement spacers for the treatment of prosthetic joint infection (PJI) remains controversial. The objective of the present study was to investigate the effectiveness of articulating cement spacers loaded with a new combination of antibiotics.