Quantitative Investigation of Terbinafine Hydrochloride Absorption into a Living Skin Equivalent Model by MALDI-MSI.

The combination of microspotting of analytical and internal standards, matrix sublimation, and recently developed software for quantitative mass spectrometry imaging has been used to develop a high-resolution method for the determination of terbinafine hydrochloride in the epidermal region of a full thickness living skin equivalent model. A quantitative assessment of the effect of the addition of the penetration enhancer (dimethyl isosorbide (DMI)) to the delivery vehicle has also been performed, and data have been compared to those obtained from LC-MS/MS measurements of homogenates of isolated epidermal tissue. At 10% DMI, the levels of signal detected for the drug in the epidermis were 0.20 ± 0.072 mg/g tissue for QMSI and 0.28 ± 0.040 mg/g tissue for LC-MS/MS at 50% DMI 0.69 ± 0.23 mg/g tissue for QMSI and 0.66 ± 0.057 mg/g tissue for LC-MS/MS. Comparison of means and standard deviations indicates no significant difference between the values obtained by the two methods.

Determining suitable surfactant concentration ranges to avoid protein unfolding in pharmaceutical formulations using UV analysis.

Protein stability is fundamental to maintain pharmaceutical efficacy in the nascent field of biologics. One particular property that is essential for therapeutic effect is retention of the folded 3-dimensional conformation, i.e. once unfolding has occurred the biologic is often rendered inactive. In this work we propose a modified form of a recently published UV spectroscopic method that identifies protein unfolding. In this study we determine concentration limits to avoid protein unfolding of two model surfactants, namely polysorbate 20 and polysorbate 80, by correlating surfactant concentration with percentage 'unfolded' for three model proteins. For each scenario two distinct regions were observed, firstly surfactant concentrations at which no unfolding had occurred, followed by a second region whereby unfolding steadily increased with surfactant concentration. In general for the combinations analysed in this study, this second region began to appear around ten times below the critical micellar concentration of each surfactant, regardless of the protein or polysorbate chosen. It is therefore proposed that this adapted method could be used by researchers in the early stages of formulation development as a convenient and simple screening tool to confirm the 'onset of unfolding' concentration for protein-surfactant formulations, thus helping to optimise surfactant concentration selection in pharmaceutical formulations to maintain the benefits of surfactants yet avoid inadvertent unfolding.

CMC determination using isothermal titration calorimetry for five industrially significant non-ionic surfactants.

Surfactants are used in a vast array of products including pharmaceuticals, cosmetics and household formulations. From an industrial perspective, non-ionic surfactants are ideal for inclusion within such products as they are non-toxic, simple to formulate and economic to use. This study considers five non-ionic surfactants (Tween 20, Tween 80, Crodasol, Croduret and Etocas 35) to determine the critical micellar concentration (CMC) for each using isothermal titration calorimetry, thus avoiding issues regarding poor accuracy found with other techniques. Furthermore, this methodology has not previously been applied to this group of surfactants. For the most commonly used non-ionics (Tween 20 and Tween 80) a further study was undertaken to consider the influence of surfactant purity on the CMC determined, using standard grade (Tween 20 and 80), high purity (Tween 20 HP and Tween 80 HP) and Super Refined (SR PS20 and SR PS80). Results permitted calculation of the CMC for the surfactants whereupon the values were determined to range from 1.0 mM for Tween 20 HP to 2.9 mM for Tween 80 HP. Such information regarding the CMC event is useful from a formulation perspective as it can ensure that the most optimum concentration of surfactant is included within a formulation to maximize its efficacy.

Adaptation of the Kirkstall QV600 LLI Microfluidics System for the Study of Gastrointestinal Absorption by Mass Spectrometry Imaging and LC-MS/MS.

Absorption studies on oral drugs can be difficult due to the challenge of replicating the complex structure and environment of the GI tract. Drug absorption studies can be conducted using in vivo and ex vivo animal tissue or animal-free techniques. These studies typically involve the use of Caco-2 cells. However, Caco-2 cells do not incorporate all the cell types found in intestinal tissue and lack P450 metabolizing enzymes. The QV600 LLI system is a microfluidics system designed for use with cell culture. Here, it has been adapted to house appropriate sections of ex vivo porcine tissue to act as a system that models the duodenum section of the small intestine. A pH regulated solution of Atorvastatin was flowed over the apical layer of the GI tissue and a nutrient solution flowed over the basal layer of the tissue to maintain tissue viability. The tissue samples were snap-frozen, cryosectioned, and imaged using MALDI Mass Spectrometry Imaging (MSI). A proof-of-concept study on the effect of excipients on absorption was conducted. Different concentrations of the solubilizing agent were added to the donor circuit of the QV600 LLI. The amount of Atorvastatin in the acceptor circuit was determined to study the effect of the excipient on the amount of drug that had permeated through the tissue. Using these data, Papp, pig values were calculated and compared with the literature.

Hand-assisted laparoscopic donor nephrectomy: are hand port devices really necessary?

BACKGROUND: Hand port devices (HPD) are used routinely for hand-assisted laparoscopic surgery including hand-assisted laparoscopic donor nephrectomy (HALDN). However, the cost of such devices may prove prohibitive, particularly in centers with financial constraints. The authors aimed to identify any adverse effects of performing device-free HALDN. METHODS: A retrospective analysis was performed of patients undergoing HALDN at the authors' unit over a 3-year period (2007-2010). Eighty-four patients underwent device-free HALDN, whereas in 80 patients a HPD was used. The primary endpoint was duration of operation, with secondary endpoints including postoperative wound infections and incisional hernias. RESULTS: here was no difference in duration of operation for the device free (98 minutes; range = 43-215 minutes) compared with the HPD group (94 minutes; range = 36-180 minutes; P = .37). A device was required in 3 (3.6%) patients in which a device-free approach was attempted. There was no difference in either group in terms of rates of postoperative wound infections (0% vs 2.5%, respectively; P = .24) or incisional hernia incidence (1.5% vs 1.4%, respectively; P = .50). CONCLUSION: Device-free HALDN can be performed with no discernable compromise in operating time or patient outcome. This has implications in both cost benefit and translation of this technique to developing units.

Understanding polysorbate-compound interactions within the CMC region.

Non-ionic surfactants such as polysorbates, known as Tween™ 20 and Tween™ 80, are routinely used within the healthcare and pharmaceutical industry to enhance solubility. This work focuses on analysing the two aforementioned polysorbates, each considered at three purity levels with four model compounds, across the critical micellar concentration (CMC) range for each surfactant. Such data is of interest to investigate the influence of micelle formation upon compound-polysorbate interaction. Two analytical techniques were utilised, namely spectroscopic solubility determination and micellar liquid chromatography (MLC). In all cases it was apparent that the maximum solubility for all four compounds increased substantially at concentrations greater than the CMC and that, in most cases, a different retention profile was observed using MLC once the CMC had been exceeded. This paper is the first to have used such techniques to investigate the behaviour of these polysorbates over a series of concentrations and three levels of polysorbate purity. The findings indicate that the solubilisation potential of polysorbates differs once the CMC has been surpassed and is dependent upon the level of purity selected, i.e. compound-surfactant interactions are partially a consequence of the presence of micelles rather than monomer as well as polysorbate purity. Thus, formulators should include such polysorbates at optimised concentrations and purity if they wish to maximise their solubilisation potential.

Risk factors for acute rejection in renal transplant recipients experiencing delayed graft function.

Acute rejection (AR) superimposed upon delayed graft function (DGF) following renal transplantation worsens graft outcomes. However, risk factors for AR in patients displaying DGF remain unclear. In this study, 71 patients displaying DGF >/= 5 d were investigated. All received cyclosporine, adjunctive azathioprine or mycophenolate mofetil (MMF), and corticosteroids, with 43 receiving anti-CD25 monoclonal antibody induction. AR episodes were seen in 20 of 71 (28%) patients. Higher C2 levels at days 3 and 5 and the use of MMF were associated with a reduced incidence of AR, with increased HLA-DR mismatch associated with an increased risk for AR. C2 levels at days 3 and 5 below 885 and 1096 ng/mL, respectively, showed best discriminatory values for AR. C2 levels showed no correlation with DGF duration. This study suggests that optimizing immunosuppression in patients with DGF (by ensuring adequate calcineurin inhibitor exposure and the use of potent adjunctive immunosuppression) may reduce the incidence of AR without prolonging the duration of dialysis requirement.

Experiences From Developing and Upgrading a Web-Based Surveillance System for Malaria Elimination in Cambodia.

Strengthening the surveillance component is key toward achieving country-wide malaria elimination in Cambodia. A Web-based upgraded malaria information system (MIS) was deemed to essentially act as the central component for surveillance strengthening. New functionality (eg, data visualization) and operational (eg, data quality) attributes of the system received particular attention. However, building from the lessons learned in previous systems' developments, other aspects unique to Cambodia were considered to be equally important; for instance, feasibility issues, particularly at the field level (eg, user acceptability at various health levels), and sustainability needs (eg, long-term system flexibility). The Cambodian process of identifying the essential changes and critical attributes for this new information system can provide a model for other countries at various stages of the disease control and elimination continuum. Sharing these experiences not only facilitates the establishment of "best practices" but also accelerates global and regional malaria elimination efforts. In this article, Cambodia's experience in developing and upgrading its MIS to remain responsive to country-specific needs demonstrates the necessity for considering functionality, operationalization, feasibility, and sustainability of an information system in the context of malaria elimination.

The novel use of infrared thermal imaging as an adjunct for the management of haemodialysis access induced distal ischaemia.

INTRODUCTION: Haemodialysis access-induced distal ischaemia (HAIDI) whilst unusual may be a difficult complication to diagnose and manage particularly in patients with small vessel disease.Infrared thermal imaging (IRTI) has been used for several decades to monitor cutaneous temperature distribution and has been used in medical applications. The use of IRTI in HAIDI is attractive as it can be used as a non-invasive measure of tissue perfusion. METHOD AND RESULTS: To assess the applicability of IRTI in HAIDI, three cases were assessed and are described. IRTI was well tolerated and provided useful clinical information in all three cases, which was confirmed with further studies. CONCLUSIONS: This small case series reports on the successful use of infrared thermal imaging in the diagnosis and management of possible HAIDI and may support wider study and use.

Focused Screening and Treatment (FSAT): a PCR-based strategy to detect malaria parasite carriers and contain drug resistant P. falciparum, Pailin, Cambodia.

Recent studies have shown that Plasmodium falciparum malaria parasites in Pailin province, along the border between Thailand and Cambodia, have become resistant to artemisinin derivatives. To better define the epidemiology of P. falciparum populations and to assess the risk of the possible spread of these parasites outside Pailin, a new epidemiological tool named "Focused Screening and Treatment" (FSAT), based on active molecular detection of asymptomatic parasite carriers was introduced in 2010. Cross-sectional malariometric surveys using PCR were carried out in 20 out of 109 villages in Pailin province. Individuals detected as P. falciparum carriers were treated with atovaquone-proguanil combination plus a single dose of primaquine if the patient was non-G6PD deficient. Interviews were conducted to elicit history of cross-border travel that might contribute to the spread of artemisinin-resistant parasites. After directly observed treatment, patients were followed up and re-examined on day 7 and day 28. Among 6931 individuals screened, prevalence of P. falciparum carriers was less than 1%, of whom 96% were asymptomatic. Only 1.6% of the individuals had a travel history or plans to go outside Cambodia, with none of those tested being positive for P. falciparum. Retrospective analysis, using 2010 routine surveillance data, showed significant differences in the prevalence of asymptomatic carriers discovered by FSAT between villages classified as "high risk" and "low risk" based on malaria incidence data. All positive individuals treated and followed-up until day 28 were cured. No mutant-type allele related to atovaquone resistance was found. FSAT is a potentially useful tool to detect, treat and track clusters of asymptomatic carriers of P. falciparum along with providing valuable epidemiological information regarding cross-border movements of potential malaria parasite carriers and parasite gene flow.

The brachiobasilic arteriovenous fistula: effect of patient variables.

PURPOSE: The hemodialysis population is constantly expanding as patients on dialysis have increased longevity and the number of kidneys available for transplantation remains static (1). After radiocephalic and brachiocephalic fistulas have been exhausted the use of the autologous brachiobasilic fistula (BBAVF) should be considered prior to use of a synthetic graft. We present our single center experience of 140 brachiobasilic fistulas in a five-year period and examine any factors that influence patency and long-term function. METHODS: Patients who had undergone formation of a BBAVF between January 2004 and January 2009 were identified; a review of all case notes and databases was undertaken. Details on demographics, cause of renal failure, co-morbidities (including diabetes, cardiac morbidity, hypertension, peripheral vascular disease), dialysis status at the time of fistula creation, hemoglobin, anti-coagulation regimens, and complications from surgery were recorded. RESULTS: Patency (defined as use of AVF for dialysis) was 83% at 3 months, 77% at 6 months, and 69% at 12 months. Length of patency ranged from 0 to 1918 days (at study cut-off) with a mean patency of 532 days. Factors found to significantly affect fistula patency included age over 60 (P=<0.001) and presence of peripheral vascular disease (P=0.048). CONCLUSIONS: Our brachiobasilic fistula patency rates are comparable with published literature and other fistulas. Within our population patient variables including age over 60 and the presence of peripheral vascular disease are associated with worse outcomes as would be expected. In spite of these factors we feel the brachiobasilic fistula is an excellent option for patients with more challenging access and should certainly be undertaken prior to the use of prosthetic grafts.

Calciphylaxis following kidney transplantation: a case report.

INTRODUCTION: Calciphylaxis occurring after kidney transplantation is rare and rarely reported. It results in chronic non-healing wounds and is associated with a poor prognosis and is often fatal. We present a case of proximal lower limb calciphylaxis that occurred early after kidney transplantation. The patient had no classic associated risk factors. He had previously had a total parathyroidectomy but had normal serum calcium-phosphate product and parathyroid hormone levels. The clinical outcome of this case was favorable and highlights some fundamental issues relating to management. CASE PRESENTATION: A 70-year-old British Caucasian man with end-stage renal failure secondary to IgA nephropathy presented six months post kidney transplantation with cutaneous calciphylaxis lesions involving the medial aspect of the thigh bilaterally. CONCLUSION: To the best of our knowledge, this is the first reported case of rapid onset cutaneous calciphylaxis occurring soon after kidney transplantation that was associated with a favorable outcome. Cutaneous calciphylaxis lesions should be promptly managed with meticulous wound care, antimicrobial therapy and the correction of calcium-phosphate product where indicated.

Predicting early renal allograft function using clinical variables.

BACKGROUND: Suboptimal early graft function following renal transplantation remains a significant challenge. It is suggested that clinical variables (or scoring systems based thereon) may predict the occurrence of delayed graft function (DGF), defined as post-operative dialysis requirement. However, data is conflicting, and suboptimal renal function not requiring dialysis has been little investigated. This study tested the ability of clinical variables to predict suboptimal early function variably assessed by: (i) DGF (dialysis requirement during the first week); (ii) DGF duration; (iii) slow graft function (creatinine>3 mg/dl on day 5); (iv) creatinine reduction ratio on day 2. METHODS: Details on 217 consecutive renal transplant recipients were collected. All received ciclosporin-based immunosuppression. Multiple regression analysis was used to assess the association between individual clinical variables and suboptimal early graft function. Also tested were three scoring systems incorporating clinical variables [US Renal Database System (USRDS score), deceased donor score (DDS) and expanded criteria donor kidneys]. Receiver operated-characteristic curve analysis was used to assess the predictive power of clinical variables and scoring systems. RESULTS: Early graft function was associated with donor age, donor body mass index, donor hypertension, donation following cardiac death, black recipient ethnicity, recipient weight and cold ischaemic time (Por=150). CONCLUSIONS: Clinical variables and scores have moderate predictive ability for early graft function and although of potential use in clinical practice, caution should be exercised before altering patient management based solely on them.