Simultaneous quantification of cyclosporin, tacrolimus, sirolimus and everolimus in whole blood by UHPLC-MS/MS for therapeutic drug monitoring.

The aim of this study was to develop and validate a UHPLC-MS/MS assay to quantify cyclosporin (CYC), tacrolimus (TAC), sirolimus (SIR) and everolimus (EVE) in human whole blood for therapeutic drug monitoring. Analytes were extracted from 50 µL human whole blood by protein precipitation. The separation of the drugs was performed on an Acquity UPLC BEH C18 column. Analytes were eluted with a mobile phase consisting of 2 mM ammonium acetate with 0.1% formic acid (v/v) in deionised water and 2 mM ammonium acetate with 0.1% formic acid (v/v) in methanol at a flow rate of 300 µL/min in gradient elution. The method performance was evaluated by analysing patient blood samples and/or external quality control samples [proficiency testing (PT) scheme]. The method was linear from 23.75 to 1094.0, 1.3 to 42.4, 1.3 to 47.0 and 1.2-41.6 µg/mL for CYC, TAC, SIR and EVE, respectively. The within- and between-assay reproducibility results were ˂ 11%. Results from PT and patient sample quantification were comparable to those obtained previously by an in-house validated method using protein precipitation and liquid-liquid extraction. This method showed good analytical performance for quantifying CYC, TAC, SIR and EVE in whole blood over their respective calibration ranges.

Alternative Input for Perfusion Management Devices: Voice Recognition for Data Input and the Effects on Charting and Perioperative Calculation Use.

Technology in healthcare has become increasingly prevalent and user friendly. In the last decade, advances in hands-free methods of data input have become more viable in a variety of medical professions. The aim of this study was to assess the advantages or disadvantages of hands-free charting through a voice-to-text app designed for perfusionists. Twelve clinical perfusion students using two different simulated bypass cases were recorded and assessed for the number of events noticed and charted, as well as the speed at which they accomplished these steps. Paper charts were compared with a custom app with voice-to-text charting capability. Data was analyzed using linear mixed models to detect differences in length of time until a chartable event was noticed, and how long after noticing an event it took to record the event. Timeliness of recording an event was made by assessing log-transformed time data. There was significantly more information recorded when charting on paper, while charting with voice-to-text resulted in significantly faster mean time from noticing an event to the recording of it. There was no significant difference between how many events were noticed and recorded. When using paper charting, a higher percentage of events that were missed were drug administration events, while voice charting had a higher percentage of missed events that were associated with cardioplegia delivery or bypass timing. With a decreased time interval between noticing an event and charting the event, speech-to-text for perfusion could be of benefit in situations where many events occur at once, such as emergency situations or highly active portions of bypass such as initiation and termination. While efforts were made to make the app as intuitive as possible, there is room for improvement.

Discussion: can upper extremity (deltoid) near infrared spectroscopy be used to assess cerebral tissue bed saturation on femorally cannulated veno-arterial extracorporeal membrane oxygenation patients?

Continuous cerebral tissue saturation monitoring with near infrared spectroscopy may help clinicians identify cerebral desaturation early; however, patients have reported discomfort from near infrared spectroscopy monitoring pads on the forehead. This study aims to compare upper extremity near infrared spectroscopy monitoring to cerebral near infrared spectroscopy monitoring to assess its viability as a surrogate for cerebral saturation. A retrospective analysis of 10 femorally cannulated veno-arterial extracorporeal membrane oxygenation patients was performed comparing left (L) and right (R) upper extremity (deltoid) near infrared spectroscopy monitoring to cerebral near infrared spectroscopy monitoring (n = 20 data sets, 10 left and 10 right) and right radial blood gasses. Deltoid and cerebral near infrared spectroscopy values were recorded every 15 minutes for at least 24 hours when possible, were plotted on scatter grams, and were analyzed using Pearson product-moment coefficient (r). Based on the concept of covariance, a moderate-good relationship r = 0.50-0.75 was noted in 10% (n = 2) of the study group. A fair relationship r = 0.25-0.50 was noted in 50% (n = 10), and little or no relationship was noted in 40% (n = 8). None of the study group displayed a good to excellent relationship (r = 0.75 or above). In addition, coefficient of multiple determination for multiple regression R2 was calculated and strong fit of the regression line was not noted. Although cerebral near infrared spectroscopy monitoring has been extremely helpful in identifying low cerebral tissue saturation on veno-arterial extracorporeal membrane oxygenation patients, the use of upper extremity (peripheral deltoid) tissue monitoring does not provide adequate correlation and should not be used as a surrogate to cerebral monitoring.

Quantification of mycophenolic acid in human plasma by liquid chromatography with time-of-flight mass spectrometry for therapeutic drug monitoring.

This study presents, for the first time, the development and validation of a liquid chromatography and time-of-flight mass-spectrometry (LC-TOF-MS) based assay to quantify mycophenolic acid (MPA) in patient samples as part of a routine therapeutic drug monitoring service. MPA was extracted from 50 µl human plasma by protein precipitation, using sulindac as internal standard (IS). Separation was obtained on a Luna™ Omega polar C18 column kept at 40°C. The mobile phase consisted of a mixture of acetonitrile-deionized water (50:50, v/v) with 0.1% formic acid at a flow rate of 350 µl/min. Analyte and IS were monitored on a TOF-MS using a Jet-Stream™ (electrospray) interface running in positive mode. Assay performance was evaluated by analysing patient plasma (N = 69) and external quality assessment (N = 6) samples. The retention times were 2.66 and 2.18 min for MPA and IS, respectively. The lower limit of quantification of MPA was 0.1 µg/ml. The within- and between-assay reproducibility results ranged from 1.81 to 10.72%. Patient and external quality assessment sample results were comparable with those obtained previously by an in-house validated LC-MS/MS method. This method showed satisfactory analytical performance for the determination of MPA in plasma over the calibration range of 0.1-15.0 µg/ml.

Perfusion Recruitment Strategies Using Choice-Based Conjoint Analysis.

Perfusionists are leaving the workforce in higher rates as baby-boomers retire, yet the volume of cases involving perfusion continues to increase. This change has resulted in a high-demand market with organizations competing for the same candidates. The purpose of this study was to use a choice-based conjoint (CBC) analysis to determine what attributes of employment opportunities are most important to perfusionists so hiring organizations are better equipped to produce employment offers specific to each candidate. The Sawtooth Software Discover program was used to create a CBC survey that was distributed to perfusionists. After a series of demographic questions, respondents chose between three employment offers that comprised eight attributes, each with three varying options. Completed surveys were analyzed using the Discover software, producing importance scores. Two hundred forty surveys were completed, 154 (64%) male and 86 (36%) female. Extracorporeal membrane oxygenation (ECMO) impact to the perfusionist and hours worked per week ranked most important, at 19.4% and 17.5%, respectively. A significant difference exists between ECMO impact to perfusionists and all other attributes. The least important attributes were annual continuing education funds and sign-on bonus/relocation, at 6.8% and 4.7%, respectively. Respondents in their 20s and 30s find the importance of ECMO impact to be significantly less important than those in their 50s. Perfusionists with a master's degree believe salary to be significantly more important than perfusionists with other types of perfusion education. Based on the respondents who completed this survey, the impact ECMO has on the perfusionist was the most significant job attribute. Offering large sign-on bonuses and relocation packages to entice new employees is not indicated as a viable way to satisfy perfusionists. Appropriate management of hours worked, days on call, and how ECMO impacts the perfusionists will be most impactful in recruitment of new employees.

Comparison of three infant venous reservoirs with vacuum-assisted venous drainage during varying levels of cardiotomy suction.

BACKGROUND: Vacuum-assisted venous drainage has gained widespread use within the pediatric perfusion community for use during cardiopulmonary bypass. It is questioned whether its efficiency may be compromised with application of excessive cardiotomy suction to the infant hard-shell venous reservoir. An in vitro simulation circuit was used to research this phenomenon. A comparison of three different infant hard-shell venous reservoirs also took place to determine if one reservoir type was more advantageous when handling cardiotomy suction. The reservoirs tested were the Maquet VHK 11000, Medtronic Affinity Pixie, and Terumo Capiox FX05. METHODS: The in vitro simulation circuit consisted of a 1 L reservoir bag that was cannulated at one access point with an Edwards Lifesciences 10Fr aortic cannula and the other access area with an Edwards Lifesciences 10Fr right angle venous cannula and 12Fr right angle venous cannula that were joined together. Key points of measurement and response variables were the pressures on the connection of the venous cannulas, inlet of the venous reservoir, and flow through the venous line. Vacuum was applied and manipulated with a Maquet VAVD Controller to settings of -20 mmHg, -30 mmHg, -40 mmHg, -50 mmHg, and -60 mmHg. Cardiotomy suction was added at settings of 1 LPM, 2 LPM, 3 LPM, and 4 LPM. Values from each response variable were monitored and recorded. These data were utilized to compare the reservoirs with a random coefficient model for each response variable. CONCLUSIONS: There is an adverse effect of excessive cardiotomy suction on the efficacy of vacuum-assisted venous drainage in infant hard-shell venous reservoirs. There is no significant difference between the VHK 11000, Pixie, and FX05 regarding their ability to handle this occurrence. An important discovery was that the FX05 showed a greater transfer of vacuum to the venous cannulas and reservoir inlet.

Improving Decreased Heater-Cooler Efficiency as a Result of Heater-Cooler Infection Control Strategy.

Heater-cooler units (HCUs) play a vital role in temperature management during cardiopulmonary bypass. In recent years, HCUs have been shown to play a significant role in the propagation of bacteria causing patient infection and significant harm. As a result, various institutions across the world have begun moving the HCU either far away or outside of the operative theater entirely. The purpose of this study was to examine the effect that the increased length of HCU water lines have on the ability of the device to heat and cool. We hypothesized that the increase in water line distance leads to a decrease in HCU efficiency and that insulating the water lines would blunt the effect of this increase in distance. Five water line conditions were compared under two cooling and two warming ranges. Short water lines, long water lines, and long water lines with foam, rubber, or tape insulation were compared. Cooling from an arterial line temperature of 26.7-19.7°C showed no difference between conditions with the exception that every long line condition takes significantly longer to cool than short water lines. Cooling from 35.6 to 28.6°C revealed that all insulations reduce the cooling time compared with long water lines without insulation, but only foam insulation reduces to the level of the short water lines. During warming conditions, all insulations reduced the warming time compared with long uninsulated water lines, but none were comparable with short water lines. Increased water line length leads to a decrease in HCU efficiency. Insulation is effective at increasing efficiency of long water lines, but only at warmer temperatures and not to the level of short water lines. Only foam-insulated long water lines were able to match the efficiency of short water lines, but only across a single temperature range.

Effects of ECMO Simulations and Protocols on Patient Safety.

The use of extracorporeal membrane oxygenation (ECMO) has greatly increased over the years; however, the survival rate is only above 56%. There has been a drastic increase in ECMO centers and cases. ECMO has become a popular therapy route for patients with respiratory and cardiac complications; however, patient safety is a major concern. Perfusion and non-perfusion students from the University of Nebraska Medical Center were recruited to participate in three simulation trials. The trials consisted of five different tasks that are required for managing or preventing catastrophic events on ECMO. Students were evaluated for the time it took to complete each task, number of errors made, and protocol referencing. The data indicated that there was a decrease in time for the 1st vs. 2nd trial (p = .02) for perfusion students and a decrease from the 1st to 3rd trial (p = .001) for the circuit set-up simulation. There was a decrease in priming time from the 1st to 3rd trial (p = .02) and for the pump change (p = .0098) for the perfusion students as well. The non-perfusion students had a significant decrease in time for the circuit set-up in the 1st vs. 2nd (p = .004) and 1st vs. 3rd trial (p = .002). There was a decrease in time for priming (.004), pump change (p = .002), tubing change (p = .0098), and errors during the tubing change (p = .02) in the non-perfusion students. Both groups felt more confident after the simulations and the non-perfusion students specifically felt like they were more familiar with the purpose of ECMO after the simulation. ECMO simulations and protocols may improve patient safety by strengthening the skills needed for rapid management, fewer errors, and higher levels of confidence during the management of ECMO and catastrophic events.

Flow Awareness as a New Safety Device for Cardiopulmonary Bypass.

Safety devices such as bubble detectors and level detectors have been in use for more than 40 years and initially reduced the risks associated with gaseous emboli during cardiopulmonary bypass; however, the risks have not been eliminated. This research explored a new safety device designed to further reduce these risks: Flow Awareness Technology. This device visually alerts the perfusionist when the ratio of venous return does not equal the amount of arterial flow. To determine the efficacy of this device, 33 participants with no perfusion background were randomly assigned to Group A (flow awareness only), Group B (level detector only), or Group C (control). These participants were instructed to turn off the arterial pump when they noticed that the fluid in the venous reservoir had begun to drop or if their assigned safety device was triggered. The venous line was fully occluded at times unknown to the participants. These times coincided with before, during, or after times that the participants were expected to paper-chart known values. Each participant's amount of fluid lost (in milliliters) from the reservoir and reaction time (in seconds) to shut off the arterial roller pump were measured. Group A lost an average of 80.8 mL, Group B lost an average of 173.6 mL, and Group C lost an average of 140.3 mL. Average measured time for each group is as follows: Group A took 2.16 seconds, Group B took 4.31 seconds, and Group C took 4.09 seconds to shut off the arterial pump. Statistics support the hypothesis that Flow Awareness Technology significantly reduces the reaction time to an adverse event such as a sudden occlusion of the venous line, thus reducing the amount of fluid lost during such an event.

Colloid Oncotic Pressure, Monitoring its Effects in Cardiac Surgery.

Hemodilution is a common perioperative practice. The deleterious effects of excessive hemodilution and subsequent edema formation have been well documented by numerous authors. Colloid oncotic pressure (COP) is a reliable clinical indicator of hemodilution in cardiac surgery. The intent of this study is to determine if a correlation exists between COP and various patient outcome variables. It would also be helpful to know if there is a particular COP value to avoid preventing or limiting patient morbidity. Blood samples from 61 adult patients (mean age = 70 years old) undergoing cardiopulmonary bypass surgery were collected for COP calculation and comparison. Sample collection was performed before heparinization, during cardiopulmonary bypass, at the conclusion of cardiopulmonary bypass, and in the intensive care unit. The resultant values obtained were used to generate a calculated COP. The lowest sustained COP was then compared with various patient outcome variables such as fluid balance, post-operative weight gain, post-operative blood loss, extubation time, length of stay, and blood products administered. A statistically significant difference (p < .05) was found between the COP and each of the monitored continuous variables. The data also suggest that maintaining a patient's COP at or above 15 mmHg could be desirable. Frequent monitoring of a patient's COP can provide a potential benefit to clinical decision making.

Removal of Gross Air Embolization from Cardiopulmonary Bypass Circuits with Integrated Arterial Line Filters: A Comparison of Circuit Designs.

Advances in technology, the desire to minimize blood product transfusions, and concerns relating to inflammatory mediators have lead many practitioners and manufacturers to minimize cardiopulmonary bypass (CBP) circuit designs. The oxygenator and arterial line filter (ALF) have been integrated into one device as a method of attaining a reduction in prime volume and surface area. The instructions for use of a currently available oxygenator with integrated ALF recommends incorporating a recirculation line distal to the oxygenator. However, according to an unscientific survey, 70% of respondents utilize CPB circuits incorporating integrated ALFs without a path of recirculation distal to the oxygenator outlet. Considering this circuit design, the ability to quickly remove a gross air bolus in the blood path distal to the oxygenator may be compromised. This in vitro study was designed to determine if the time required to remove a gross air bolus from a CPB circuit without a path of recirculation distal to the oxygenator will be significantly longer than that of a circuit with a path of recirculation distal to the oxygenator. A significant difference was found in the mean time required to remove a gross air bolus between the circuit designs (p = .0003). Additionally, There was found to be a statistically significant difference in the mean time required to remove a gross air bolus between Trial 1 and Trials 4 (p = .015) and 5 (p =.014) irrespective of the circuit design. Under the parameters of this study, a recirculation line distal to an oxygenator with an integrated ALF significantly decreases the time it takes to remove an air bolus from the CPB circuit and may be safer for clinical use than the same circuit without a recirculation line.

Therapeutic Drug Monitoring of Everolimus: A Consensus Report.

In 2014, the Immunosuppressive Drugs Scientific Committee of the International Association of Therapeutic Drug Monitoring and Clinical Toxicology called a meeting of international experts to provide recommendations to guide therapeutic drug monitoring (TDM) of everolimus (EVR) and its optimal use in clinical practice. EVR is a potent inhibitor of the mammalian target of rapamycin, approved for the prevention of organ transplant rejection and for the treatment of various types of cancer and tuberous sclerosis complex. EVR fulfills the prerequisites for TDM, having a narrow therapeutic range, high interindividual pharmacokinetic variability, and established drug exposure-response relationships. EVR trough concentrations (C0) demonstrate a good relationship with overall exposure, providing a simple and reliable index for TDM. Whole-blood samples should be used for measurement of EVR C0, and sampling times should be standardized to occur within 1 hour before the next dose, which should be taken at the same time everyday and preferably without food. In transplantation settings, EVR should be generally targeted to a C0 of 3-8 ng/mL when used in combination with other immunosuppressive drugs (calcineurin inhibitors and glucocorticoids); in calcineurin inhibitor-free regimens, the EVR target C0 range should be 6-10 ng/mL. Further studies are required to determine the clinical utility of TDM in nontransplantation settings. The choice of analytical method and differences between methods should be carefully considered when determining EVR concentrations, and when comparing and interpreting clinical trial outcomes. At present, a fully validated liquid chromatography tandem mass spectrometry assay is the preferred method for determination of EVR C0, with a lower limit of quantification close to 1 ng/mL. Use of certified commercially available whole-blood calibrators to avoid calibration bias and participation in external proficiency-testing programs to allow continuous cross-validation and proof of analytical quality are highly recommended. Development of alternative assays to facilitate on-site measurement of EVR C0 is encouraged.

Assuring the Proper Analytical Performance of Measurement Procedures for Immunosuppressive Drug Concentrations in Clinical Practice: Recommendations of the International Association of Therapeutic Drug Monitoring and Clinical Toxicology Immunosuppressive Drug Scientific Committee.

Monitoring immunosuppressive drugs (ISDs) in blood or plasma is still a key therapeutic drug monitoring (TDM) application in clinical settings. Narrow target ranges and severe side effects at drug underexposure or overexposure make accurate and precise measurements a must. This overview prepared by the Immunosuppressive Drugs Scientific Committee of the International Association of Therapeutic Drug Monitoring and Clinical Toxicology is intended to serve as a summary and guidance document describing the current state-of-the-art in the TDM of ISDs.

Long-term evaluation of analytical methods used in sirolimus therapeutic drug monitoring.

Results of therapeutic monitoring of sirolimus blood concentrations are assay and laboratory dependent. This study compared performance over time of the IMx microparticle enzyme immunoassay (MEIA), Architect chemiluminescent microparticle immunoassay (CMIA), and liquid chromatography with mass spectrometric detection (LC/MS/MS) as part of a proficiency testing scheme. Pooled samples from sirolimus-treated patients and whole-blood samples spiked with known quantities of sirolimus were assayed monthly between 2004 and 2012. When results of pooled patient samples were compared with LC/MS/MS, the MEIA assay showed an overall mean percent bias of -2.3% ± 11.2% that, although initially positive, became increasingly negative from 2007 through 2009. The CMIA, which replaced the MEIA assay, had a mean percent bias of 21.9% ± 12.3%, remaining stable from 2007 through 2012. Similarly, for spiked samples, the MEIA showed an increasingly negative bias over time vs. LC/MS/MS, whereas CMIA maintained a stable positive bias. Based on comparison of immunoassay measurements on individual patient samples, CMIA values were more than 25% higher than MEIA values. These results highlight the importance of continued proficiency testing and regular monitoring of sirolimus assay performance. Clinicians must be aware of the methodology used and adjust target levels accordingly to avoid potential effects on efficacy and toxicity.

Substandard drugs: a potential crisis for public health.

Poor-quality medicines present a serious public health problem, particularly in emerging economies and developing countries, and may have a significant impact on the national clinical and economic burden. Attention has largely focused on the increasing availability of deliberately falsified drugs, but substandard medicines are also reaching patients because of poor manufacturing and quality-control practices in the production of genuine drugs (either branded or generic). Substandard medicines are widespread and represent a threat to health because they can inadvertently lead to healthcare failures, such as antibiotic resistance and the spread of disease within a community, as well as death or additional illness in individuals. This article reviews the different aspects of substandard drug formulation that can occur (for example, pharmacological variability between drug batches or between generic and originator drugs, incorrect drug quantity and presence of impurities). The possible means of addressing substandard manufacturing practices are also discussed. A concerted effort is required on the part of governments, drug manufacturers, charities and healthcare providers to ensure that only drugs of acceptable quality reach the patient.

Standardization of LC-MS for therapeutic drug monitoring of tacrolimus.

BACKGROUND: LC-MS is increasingly used for therapeutic drug monitoring of tacrolimus. A recent summary from an international proficiency-testing scheme demonstrated that the mass spectrometry respondents were the largest method group. However, these methods lack standardization, which may explain the relatively poor interlaboratory agreement for such methods. This study aimed to provide one path toward the standardization of tacrolimus quantification by use of LC-MS. METHODS: A 40-member whole blood tacrolimus proficiency panel was circulated to 7 laboratories, 4 in the US and 3 in Europe, offering routine LC-MS-based quantification of tacrolimus. All laboratories used a common LC-MS platform and followed the manufacturer's instructions that accompanied an LC-MS reagent kit intended for tacrolimus quantification in whole blood samples. Four patient pools were prepared that had sufficient volume to allow comparison with a tacrolimus reference measurement procedure. RESULTS: For the 40-member panel, the standardized MassTrak LC-MS assay demonstrated excellent agreement with a validated LC-MS method used by Analytical Services International (y = 1.02x - 0.02; r = 0.99). The CVs for the pooled patient samples ranged from 2.0% to 5.4%. The mean difference from the reference measurement procedure ranged from 0.4% to 4.4%. CONCLUSIONS: Tacrolimus assay standardization, which must include all facets of the analysis, is necessary to compare patient results between laboratories and to interpret consensus guidelines. LC-MS can provide accurate and precise measurement of tacrolimus between laboratories.

Has the new USP assay for heparin affected dosage for patients undergoing cardiopulmonary bypass?

In October 2009, the U.S. Pharmacopoeia (USP) changed the monograph for heparin to bring USP units in line with international units for heparin. The result was a 10% decrease in potency as measured by in vitro laboratory tests. This decrease led to questions regarding dosing guidelines. There existed a need for an in vivo study to determine the practical changes that may need to be implemented in regard to heparin administration for cardiopulmonary bypass in the clinical setting. A retrospective study was conducted to determine the heparin dose administered and the corresponding effect on patients undergoing coronary artery bypass grafting surgery using cardiopulmonary bypass. The study compared the heparin dose requirements and activated clotting time (ACT) results using the heparin before and after the USP changes. An analysis of the data was performed to determine the increased heparin dose required to achieve the same effect as before the USP change. This new heparin dosing protocol was instituted at Concord Hospital, Concord, NH. A prospective study was then preformed to verify the effects of the dosing change. In the new heparin group, the postheparin ACT fell by 9.1% (p = .028) and the patients achieving an ACT > 479 seconds fell by 12.8% as compared with the old heparin group. After adjustment of the loading dose calculation for heparin, the prospective study demonstrated the postheparin ACT (p = .684) and the percentage of patients achieving an ACT > 479 seconds (p = 1.000) to be similar to the values obtained before the USP change. An increase of the loading dose of approximately 12% is needed to achieve the patient effects seen before the UPS change.

Simultaneous determination of tramadol, O-desmethyltramadol and N-desmethyltramadol in human urine by gas chromatography-mass spectrometry.

Analytical procedures for the determination of tramadol (T), O-desmethyltramadol (ODT), and N-desmethyltramadol (NDT) in human urine have been developed and validated using gas chromatography-mass spectrometry (GC/MS). Sample preparation involved liquid-liquid extraction with methyl-tert-butyl ether (MTBE) and followed by back extraction with 0.1 M hydrochloric acid. Proadifen (SKF525A) was selected as internal standard (IS). Extraction efficiencies of T, ODT and NDT were 102.12, 101.30, and 98.21%, respectively. The calibration curves were linear (r(2)>0.99) in the concentration range 10-1000 ng/mL for all compounds. Limits of quantification (LOQ) were 10, 10 and 20 ng/mL for T, ODT and NDT, respectively. Intra-assay precision was within 1.29-6.48% and inter-assay precision was within 1.28-6.84% for T, ODT and NDT. Intra-assay accuracy was within 91.79-106.89% for all analytes. This method detected urine concentrations of T, ODT and NDT in six healthy volunteers for 7 days after administration of 50 mg oral doses of tramadol.