Identification and quantification of glucosamine in rabbit cartilage and correlation with plasma levels by high performance liquid chromatography-electrospray ionization-tandem mass spectrometry.

A new HPLC-ESI-MS/MS method for the determination of glucosamine (2-amino-2-deoxy-d-glucose) in rabbit cartilage was developed and optimized. Glucosamine was extracted from cartilage by cryogenic grinding followed by protein precipitation with trichloroacetic acid. The HPLC separation was achieved with a polymer-based amino column using a mobile phase composed of 10mM ammonium acetate (pH 7.5)-acetonitrile (20:80%, v/v) at 0.3 mL min flow rate. d-[1-(13)C]Glucosamine was used as internal standard. Selective detection was performed by tandem mass spectrometry with electrospray source, operating in positive ionization mode and in multiple reaction monitoring acquisition (m/z 180→72 and 181→73 for glucosamine and internal standard, respectively). Limit of quantification was 0.045 ng injected, corresponding to 0.25 µg g⁻¹ in cartilage. Linearity was obtained up to 20 µg g⁻¹ (R(2)>0.991). Precision values (%R.S.D.) were <10%. Accuracy (% bias) ranged from -6.0% to 12%. Mean recoveries obtained at 3 concentration levels were higher than 81% (%R.S.D.≤8%). The method was applied to measure glucosamine levels in rabbit cartilage and plasma after single oral administration of glucosamine sulfate at a dose of 98 mg kg⁻¹(n=6). Glucosamine was present in cartilage in physiological condition before the treatment. After dosing, mean concentration of cartilage glucosamine significantly increased from 461 to 1040 ng g⁻¹. Cartilage glucosamine levels resulted to be well correlated with plasma concentrations, which therefore are useful to predict the target cartilage concentration and its pharmacological activity.

Functional results in calcific tendinitis of the shoulder treated with rehabilitation after ultrasonic-guided approach.

Calcific tendinopathy of the rotator cuff is a chronic disease that mostly in the acute phase compromises the articular function. The aim of this study is to estimate the effectiveness of the ultrasonic-guided percutaneous treatment (UGPT) in association with the rehabilitative treatment. We evaluated 106 patients with calcific tendinopathy, treated by UGPT. They underwent clinical evaluation by a physiatrist at T0 (the same day of UGPT), and were reassessed at follow-up 1 month (T1) after treatment. The assessment at T0 and T1 was done by the Constant-Murley scale. Analyzing the results, we found that at T0, the average Constant score was 43.5 out of 100; at T1 it was 83.2 out of 100. The improvement was statistically significant (P < 0.0005). We found that UGPT and rehabilitation associated with the multidisciplinary management of the patient (orthopedic surgeon-radiologist-physiatrist) was able to prevent adhesive bursitis, and to achieve clinical cure in most of the treated cases.

Humeral shaft aseptic nonunion: treatment with opposite cortical allograft struts.

Plate fixation with cortical allograft struts has been used at our Institute for decades to treat aseptic shaft nonunion. The aim of this study was to assess the results of this technique in humeral nonunion. We retrospectively reviewed 57 consecutive patients with humeral diaphyseal nonunion treated by internal fixation combined with cortical allograft struts in the last 7 years in our Department. The patients were followed-up for a mean of 48 months. We had union in 53 cases out of 57. There were 3 cases of infection out of 15 patients previously treated with an external fixator. In our experience the cortical allograft strut is a well standardised and reproducible technique that enables the treatment of severe atrophic non-union with a relatively low complication rate and quick functional recovery.

Glucosamine binding to proteins in plasma and synovial fluid and blood cell/plasma partitioning in mouse and man in vitro.

Protein binding of [14C]glucosamine (400, 1000 and 4000 ng/ml) was evaluated in human and mouse plasma and in human synovial fluid. Blood cell/plasma partitioning in human and mouse was also determined. There was no measurable protein binding of [14C]glucosamine. Its association with human and mouse blood cells ranged from 43-47% and from 27-29%, respectively. Therefore, the unbound (pharmacologically active) fraction of glucosamine in plasma and at the site of action (the joint) is the same. Protein binding displacement drug-drug interactions are unlikely during the clinical use of crystalline glucosamine sulfate. No corrections are needed, either for unbound fraction when comparing human and mouse pharmacokinetic data or for blood cell/plasma partitioning to assess glucosamine total blood clearance from plasma data in these two species.