Evidence for European presence in the Americas in AD 1021.

Transatlantic exploration took place centuries before the crossing of Columbus. Physical evidence for early European presence in the Americas can be found in Newfoundland, Canada1,2. However, it has thus far not been possible to determine when this activity took place3-5. Here we provide evidence that the Vikings were present in Newfoundland in AD 1021. We overcome the imprecision of previous age estimates by making use of the cosmic-ray-induced upsurge in atmospheric radiocarbon concentrations in AD 993 (ref. 6). Our new date lays down a marker for European cognisance of the Americas, and represents the first known point at which humans encircled the globe. It also provides a definitive tie point for future research into the initial consequences of transatlantic activity, such as the transference of knowledge, and the potential exchange of genetic information, biota and pathologies7,8.

Exploring social-based discrimination among nursing home certified nursing assistants.

Certified nursing assistants (CNAs) provide the majority of direct care to nursing home residents in the United States and, therefore, are keys to ensuring optimal health outcomes for this frail older adult population. These diverse direct care workers, however, are often not recognized for their important contributions to older adult care and are subjected to poor working conditions. It is probable that social-based discrimination lies at the core of poor treatment toward CNAs. This review uses perspectives from critical social theory to explore the phenomenon of social-based discrimination toward CNAs that may originate from social order, power, and culture. Understanding manifestations of social-based discrimination in nursing homes is critical to creating solutions for severe disparity problems among perceived lower-class workers and subsequently improving resident care delivery.

Pharmacokinetics of fluticasone propionate and salmeterol delivered as a combination dry powder via a capsule-based inhaler and a multi-dose inhaler.

AIM: To compare salmeterol (SALM) and fluticasone propionate (FP) systemic exposure following inhaled salmeterol/fluticasone propionate combination (SFC) from a unit-dose capsule-based inhaler (Rotacaps(®)/Rotahaler(®)) and a multi-dose dry powder inhaler (Diskus(®)) in healthy volunteers. METHODS: An open-label, randomised, repeat-dose, cross-over, adaptive design study (n = 36 in each part) evaluated SFC 50/250 µg and SFC 50/100 µg in Rotacaps used with two types of Rotahaler inhalers (airflow resistance similar to (S) and lower than (L) Diskus) versus the Diskus. Primary endpoints were area under the concentration-time curve over the dosing interval [AUC0-τ] and maximum plasma concentration [Cmax]. RESULTS: SFC 50/250 µg Rotacaps/Rotahaler (S) showed 1.2-1.9-fold greater FP and SALM systemic exposure compared with Diskus. FP and SALM systemic exposure were comparable to DISKUS following SFC 50/250 µg Rotacaps/Rotahaler (L) (90% CI of ratio of Rotahaler to DISKUS within 0.8-1.25) for salmeterol (AUC0-τ and Cmax) and FP (AUC0-τ). Following SFC 50/100 µg Rotacaps/Rotahaler (L), FP and SALM systemic exposures were 1.2-1.4 fold higher in terms of FP (AUC0-τ and Cmax) and salmeterol (Cmax) compared with Diskus. SFC at both doses and via both inhalers was well tolerated. CONCLUSIONS: SFC 50/250 µg Rotacaps/Rotahaler (L) showed comparable systemic exposure to Diskus in terms of FP AUC and SALM AUC and Cmax. These results merit further progression of SFC 50/250 µg Rotacaps/Rotahaler (L) to phase 3 clinical evaluation in asthma and COPD patients. The lack of pharmacokinetic comparability between the inhalers for SFC 50/100 µg requires further evaluation.

Health Equity in the Veterans Health Administration From Veterans' Perspectives by Race and Sex.

Importance: Advancing equitable patient-centered care in the Veterans Health Administration (VHA) requires understanding the differential experiences of unique patient groups. Objective: To inform a comprehensive strategy for improving VHA health equity through the comparative qualitative analysis of care experiences at the VHA among veterans of Black and White race and male and female sex. Design, Setting, and Participants: This qualitative study used a technique termed freelisting, an anthropologic technique eliciting responses in list form, at an urban academic VHA medical center from August 2, 2021, to February 9, 2022. Participants included veterans with chronic hypertension. The length of individual lists, item order in those lists, and item frequency across lists were used to calculate a salience score for each item, allowing comparison of salient words and topics within and across different groups. Participants were asked about current perceptions of VHA care, challenges in the past year, virtual care, suggestions for change, and experiences of racism. Data were analyzed from February 10 through September 30, 2022. Main Outcomes and Measures: The Smith salience index, which measures the frequency and rank of each word or phrase, was calculated for each group. Results: Responses from 49 veterans (12 Black men, 12 Black women, 12 White men, and 13 White women) were compared by race (24 Black and 25 White) and sex (24 men and 25 women). The mean (SD) age was 64.5 (9.2) years. Some positive items were salient across race and sex, including "good medical care" and telehealth as a "comfortable/great option," as were some negative items, including "long waits/delays in getting care," "transportation/traffic challenges," and "anxiety/stress/fear." Reporting "no impact" of racism on experiences of VHA health care was salient across race and sex; however, reports of race-related unprofessional treatment and active avoidance of race-related conflict differed by race (present among Black and not White participants). Experiences of interpersonal interactions also diverged. "Impersonal/cursory" telehealth experiences and the need for "more personal/attentive" care were salient among women and Black participants, but not men or White participants, who associated VHA care with courtesy and respect. Conclusions and Relevance: In this qualitative freelist study of veteran experiences, divergent experiences of interpersonal care by race and sex provided insights for improving equitable, patient-centered VHA care. Future research and interventions could focus on identifying differences across broader categories both within and beyond race and sex and bolstering efforts to improve respect and personalized care to diverse veteran populations.

Toward Part-per-Million Precision in the Determination of an Ion's Collision Cross Section Using Multipass Cyclic Ion Mobility.

In cyclic ion mobility (cIMS), ions are permitted to travel multiple passes around the drift cell, increasing the distance traveled and the relative separation between ions. This study tests the hypothesis that multiple passes around the cell can also result in improved precision when measuring an ion's mobility and the collision cross section (TWCCS) derived therefrom. Experiments were performed with a diverse set of compounds, including 16 polycyclic aromatic hydrocarbons using gas chromatographic atmospheric pressure chemical ionization and a set of drug molecules by direct infusion electrospray ionization. The average periodic drift time, viz., the average time required for the ion to travel around the cIMS cell once, shifts dramatically, approaching part-per-million (ppm) precision as the number of passes increases to ∼100. Extrapolation of the precision of the CCS values with respect to the number of passes led to the prediction that the precision will reach 1000 ppm after 50 passes, 100 ppm after 100 passes, and <10 ppm after 150 passes. Experiments wherein the number of passes exceeded 100 produced TWCCS values having within-run precisions ranging between 15 and 117 ppm. The improved precision with an increasing number of passes may be a consequence of mitigating space-charge effects by allowing the ions to occupy a larger region of the cIMS cell. A method is proposed to enable practical measurements of TWCCS with ppm precision and is demonstrated to characterize an unknown drug mixture.

The relationship between perceived discrimination and patient experiences with health care.

BACKGROUND: Prior studies have shown that racial/ethnic minorities have lower Consumer Assessments of Healthcare Providers and Systems (CAHPS) scores. Perceived discrimination may mediate the relationship between race/ethnicity and patient experiences with care. OBJECTIVE: To examine the relationship between perceived discrimination based on race/ethnicity and Medicaid insurance and CAHPS reports and ratings of care. METHODS: The study analyzed 2007 survey data from 1509 Florida Medicaid beneficiaries. CAHPS reports (getting needed care, timeliness of care, communication with doctor, and health plan customer service) and ratings (personal doctor, specialist care, overall health care, and health plan) of care were the primary outcome variables. Patient perceptions of discrimination based on their race/ethnicity and having Medicaid insurance were the primary independent variables. Regression analysis modeled the effect of perceptions of discrimination on CAHPS reports and ratings controlling for age, sex, education, self-rated health status, race/ethnicity, survey language, and fee-for-service enrollment. SEs were corrected for correlation within plans. RESULTS: Medicaid beneficiaries reporting discrimination based on race/ethnicity had lower CAHPS scores, ranging from 15 points lower (on a 0-100 scale) for getting needed care to 6 points lower for specialist rating, compared with those who never experienced discrimination. Similar results were obtained for perceived discrimination based on Medicaid insurance. CONCLUSIONS: Perceptions of discrimination based on race/ethnicity and Medicaid insurance are prevalent and are associated with substantially lower CAHPS reports and ratings of care. Practices must develop and implement strategies to reduce perceived discrimination among patients.

Perspectives on Racism in Health Care Among Black Veterans With Chronic Kidney Disease.

Importance: The burden of chronic kidney disease (CKD) and end-stage kidney disease falls disproportionately on Black individuals in the US, with Black veterans experiencing substantial consequences, and only a portion of the disparities in health conditions and health care can be explained by nonbiological factors. Among Black individuals, racism is likely one of those factors, suggesting the need to examine the consequences of racism and the resulting social structures that establish and perpetuate these racial disparities. Objective: To investigate the health care experiences of Black veterans with CKD and identify and explore the racial discrimination encountered by this vulnerable population. Design, Setting, and Participants: This qualitative study used semistructured interview guides to investigate the health care experiences of 36 Black veterans with CKD who received care at the Corporal Michael Crescenz Veterans Affairs Medical Center in Philadelphia, Pennsylvania, from October 2018 to September 2019. Interview transcripts were analyzed using applied thematic analysis. Results: Among 36 Black veterans with CKD who characterized racism in the context of their care at a Veterans Affairs medical center, the mean (SD) age was 66.0 (7.8) years; 35 participants (97.2%) were male, 1 participant (2.8%) was female, and 19 participants (52.8%) were married. The mean (SD) duration of military service was 8.0 (7.0) years. Overall, 15 participants (41.7%) were not dependent on dialysis, and hypertension was the most common comorbidity (9 participants [25.0%]). Veterans described the ways in which racism produced emotional and physical stress, including psychological symptoms (eg, anger and hurt) and physiological symptoms (eg, headaches). Veterans described a strong sense of distrust in the health care system coupled with a need to be hypervigilant during clinical encounters. When encountering racism, veterans described bottling up their feelings, which sometimes led to maladaptive behavior (eg, substance use). Veterans also described individual and collective positive strategies (eg, faith) for coping with the stress of racism. Conclusions and Relevance: In this study, Black veterans with CKD experienced racism in the clinical setting that produced physical and emotional stress and a strong sense of distrust in the health care system. These findings highlight an important opportunity for education and training of health care professionals in the implementation of trauma-informed approaches to care as a means of addressing race-based stress and trauma.

Imaging and analytical methods as applied to the evaluation of vasculature and hypoxia in human brain tumors.

Tissue hypoxia results from the interaction of cellular respiration, vascular oxygen carrying capacity, and vessel distribution. We studied the relationship between tumor vasculature and regions of low pO(2) using quantitative analysis of binding of the 2-nitroimidazole EF5 given to patients intravenously (21 mg/kg) approximately 24 h preceding surgery. We describe new computer algorithms for determining EF5 binding as a function of radial distance from individual blood vessels and converting this value to tissue pO(2). Tissues from six human brain tumors were assessed. In a hemangiopericytoma, a WHO Grade 2 and WHO Grade 3 glial brain tumor, all tissue pO(2) values calculated by EF5 binding were >20 mmHg (described as "physiologically oxygenated"). In these three tumors, EF5 binding gradients (measured as a function of distance from each observed vessel) were low, with small positive and negative values averaging close to zero. Much lower tissue oxygen levels were found, including near some vessels, in glioblastomas. Gradients of EF5 binding away from vessels were larger in glioblastomas than in the low-grade tumors, but positive and negative values again averaged to near zero. Based on these preliminary data, we hypothesize a new paradigm for tumor blood flow in human brain tumors whereby in-flowing and out-flowing blood patterns may have contrasting effects on average tissue EF5 (and by inference, oxygen) gradients. Our studies also imply that neither distance to the nearest blood vessel nor distance from each observed blood vessel provide reliable estimates of tissue pO(2).

A Novel ABCA4 Mutation Associated with a Late-Onset Stargardt Disease Phenotype: A Hypomorphic Allele?

BACKGROUND: Stargardt disease (STGD) is the most common juvenile hereditary macular dystrophy. In the majority of cases, the diagnosis is made prior to 20 years of age and usually leads to loss of central vision. Late-onset STGD affects a smaller number of patients. Identifying genetic changes which could be associated with clinically important differences in severity or presentation of the disease is important for understanding the mechanisms of visual loss and for planning future therapeutic approaches. METHODS: We report a patient with the classic phenotype of STGD with late-onset mild disease exhibiting a slow clinical progression over 14 months of follow-up. RESULTS: A 37-year-old man presented with STGD and good vision of 6/24 in the right eye and of 6/6 in the left eye as well as typical electrophysiology findings. Objective and subjective visual deterioration was not noted over a period of 14 months. Macular genetic testing revealed a novel missense mutation in ABCA4 (Thr829Met) combined with Gly1961Glu, a classic STGD mutation usually associated with a moderately severe phenotype. CONCLUSIONS: It is suggested that the Thr829Met mutation could give rise to a hypomorphic allele of the ABC transporter with a resultant phenotype of comparatively mild STGD.

Molecular analysis of idiopathic subglottic stenosis for Mycobacterium species.

OBJECTIVES/HYPOTHESIS: Idiopathic subglottic stenosis (iSGS) is an unexplained obstruction involving the lower laryngeal and upper tracheal airway. Persistent mucosal inflammation is a hallmark of the disease. Epithelial microbiota dysbiosis is found in other chronic inflammatory mucosal diseases; however, the relationship between tracheal microbiota composition and iSGS is unknown. Given the critical role for host defense at mucosal barriers, we analyzed tissue specimens from iSGS patients for the presence of microbial pathogens. METHODS: Utilizing 30 human iSGS, 20 intubation-related tracheal stenosis (iLTS), and 20 healthy control specimens, we applied molecular, immunohistochemical, electron microscopic, immunologic, and Sanger-sequencing techniques. RESULTS: With unbiased culture-independent nucleic acid, protein, and immunologic approaches, we demonstrate that Mycobacterium species are uniquely associated with iSGS. Phylogenetic analysis of the mycobacterial virulence factor rpoB suggests that, rather than Mycobacterium tuberculosis, a variant member of the Mycobacterium tuberculosis complex or a closely related novel mycobacterium is present in iSGS specimens. CONCLUSION: These studies identify a novel pathogenic role for established large airway bacteria and provide new targets for future therapeutic intervention. LEVEL OF EVIDENCE: NA Laryngoscope, 127:179-185, 2017.

EF5 binding and clinical outcome in human soft tissue sarcomas.

PURPOSE: To study the 2-nitroimidazole agent EF5 as a surrogate for measuring hypoxia in a series of patients with soft tissue sarcomas, and to determine whether hypoxia measured with this technique was associated with patient outcome. METHODS AND MATERIALS: Patients with soft tissue sarcomas of the head and neck, extremity, trunk, or retroperitoneum for whom surgical excision was the initial treatment of choice, were given 21 mg/kg EF5 24-48 hours before surgery. Biopsy specimens were stained for EF5 binding with fluorescence-labeled monoclonal antibodies, and the images were analyzed quantitatively. Endpoints included the relationship between EF5 binding, clinically important prognostic factors, and patient outcome. RESULTS: Two patients with recurrent and 14 patients with de novo sarcomas were studied. There were seven low-grade, one intermediate-grade, and eight high-grade tumors. No relationship was found between EF5 binding and patient age, sex, hemoglobin level, or tumor size. In de novo tumors, the presence of mitoses and histologic grade were positively correlated with hypoxia. High-grade and -stage de novo tumors had higher levels of EF5 binding compared with low-grade and -stage tumors. Patients with de novo tumors containing moderate to severe hypoxia (> or = 20% EF5 binding), high grade, or > or = 7% mitoses were more likely to develop metastases. CONCLUSIONS: Further studies in a larger cohort of patients are necessary to determine whether hypoxia, as measured by EF5 binding, is an independent prognostic factor for outcome in high-grade sarcomas. Such data should be useful to identify high-risk patients for clinical trials to determine whether early chemotherapy will influence the occurrence of metastasis.

Comparative measurements of hypoxia in human brain tumors using needle electrodes and EF5 binding.

Hypoxia is known to be an important prognostic marker in many human cancers. We report the use of two oxygen measurement techniques in human brain tumors and compare these data with semiquantitative histological end points. Oxygenation was measured using the Eppendorf needle electrode and/or EF5 binding in 28 brain tumors. These data were compared with necrosis, mitosis, and endothelial proliferation. In some tumors, absolute EF5 binding was converted to tissue pO(2) based on in vitro calibrations. Eppendorf electrode readings could not be used to identify WHO grade 1/2 versus WHO grade 3/4 tumors, they could not differentiate grade 3 versus grade 4 glial-derived neoplasms, nor did they correlate with necrosis or endothelial proliferation scores. EF5 binding increased as the tumor grade increased and was significantly associated with necrosis and endothelial proliferation. There was no statistically significant correlation between the two hypoxia detection techniques, although both methods indicated similar absolute ranges of tissue pO(2). There was substantial inter- and intratumoral heterogeneity of EF5 binding in WHO grade 4 glial neoplasms. The majority of cells in glial-derived tumor had levels of hypoxia that were mild to moderate (defined herein as 10% to 0.5% pO(2)) rather than severe (defined as approximately 0.1% pO(2)). Immunohistochemical detection of EF5 binding tracks histological parameters in adult brain tumors, with increased binding associated with increasing necrosis and endothelial proliferation. The proportion of moderately to severely hypoxic cells is relatively low, even in the high-grade tumors. Human brain tumors are dominated by oxic to moderately hypoxic cells.

Idiopathic subglottic stenosis is associated with activation of the inflammatory IL-17A/IL-23 axis.

OBJECTIVES/HYPOTHESIS: Idiopathic subglottic stenosis (iSGS) is a rare and devastating extrathoracic obstruction involving the lower laryngeal and upper tracheal airway. It arises without known antecedent injury or associated disease process. Persistent mucosal inflammation and a localized fibrotic response are hallmarks of the disease. Despite the initial clinical description of iSGS more than 40 year ago, there have been no substantive investigations into the pathogenesis of this enigmatic and progressive airway obstruction. In these studies, we present the initial characterization of the molecular pathogenesis underlying the fibrosing phenotype of iSGS. METHODS: Utilizing 20 human iSGS and healthy control specimens, we applied histologic, immunohistochemical, molecular, and immunologic techniques. RESULTS: We demonstrate significant activation of the canonical IL-23/IL-17A pathway in the tracheal mucosa of iSGS patients, as well as identify γδ T cells as the primary cellular source of IL-17A. CONCLUSION: Our results suggest that aberrant mucosal immune activation is a component in of the pathogenesis of iSGS. Most critically, our work offers new targets for future therapeutic intervention. LEVEL OF EVIDENCE: NA Laryngoscope, 126:E356-E361, 2016.

Hypoxia and Photofrin uptake in the intraperitoneal carcinomatosis and sarcomatosis of photodynamic therapy patients.

PURPOSE: Response to photodynamic therapy depends on adequate tumor oxygenation as well as sufficient accumulation of photosensitizer in the tumor. The goal of this study was to investigate the presence of hypoxia and retention of the photosensitizer Photofrin in the tumors of patients with intra-abdominal carcinomatosis or sarcomatosis. EXPERIMENTAL DESIGN: Tumor nodules from 10 patients were studied. In nine of these patients, hypoxia was identified in histological sections of biopsied tumor after administration of the hypoxia marker 2-(2-nitroimidazol-1[H]-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide (EF5). In separate tumor nodules from 10 patients, Photofrin uptake was measured by fluorescence after tissue solubilization. RESULTS: Hypoxia existed in the tumors of five patients, with three of these patients demonstrating at least one severely hypoxic nodule. Physiological levels of oxygen were present in the tumors of four patients. An association between tumor size and hypoxia was not evident because some tumor nodules as small as approximately 2 mm in diameter were severely hypoxic. However, even these tumor nodules contained vascular networks. Three patients with severely hypoxic tumor nodules exhibited moderate levels of Photofrin uptake of 3.9 +/- 0.4 to 3.9 +/- 0.5 ng/mg (mean +/- SE). The four patients with tumors of physiological oxygenation did not consistently exhibit high tumor concentrations of Photofrin: mean +/- SE drug uptake among these patients ranged from 0.6 +/- 0.8 to 5.8 +/- 0.5 ng/mg. CONCLUSIONS: Carcinomatosis or sarcomatosis of the i.p. cavity may exhibit severe tumor hypoxia. Photofrin accumulation in tumors varied by a factor of approximately 10x among all patients, and, on average, those with severe hypoxia in at least one nodule did not demonstrate poor Photofrin uptake in separate tumor samples. These data emphasize the need for reconsideration of the generally accepted paradigm of small tumor size, good oxygenation, and good drug delivery because this may vary on an individual tumor basis.

Hypoxia is important in the biology and aggression of human glial brain tumors.

We investigated whether increasing levels of tissue hypoxia, measured by the binding of EF5 [2-(2-nitro-1-H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl) acetamide] or by Eppendorf needle electrodes, were associated with tumor aggressiveness in patients with previously untreated glial brain tumors. We hypothesized that more extensive and severe hypoxia would be present in tumor cells from patients bearing more clinically aggressive tumors. Hypoxia was measured with the 2-nitroimidazole imaging agent EF5 in 18 patients with supratentorial glial neoplasms. In 12 patients, needle electrode measurements were made intraoperatively. Time to recurrence was used as an indicator of tumor aggression and was analyzed as a function of EF5 binding, electrode values and recursive partitioning analysis (RPA) classification. On the basis of EF5 binding, WHO grade 2 tumors were characterized by modest cellular hypoxia (pO2s approximately 10%) and grade 3 tumors by modest-to-moderate hypoxia (pO2s approximately 10%- 2.5%). Severe hypoxia (approximately 0.1% oxygen) was present in 5 of 12 grade 4 tumors. A correlation between more rapid tumor recurrence and hypoxia was demonstrated with EF5 binding, but this relationship was not predicted by Eppendorf measurements.

Quantitative analysis of multiple illicit drugs in preserved oral fluid by solid-phase extraction and liquid chromatography-tandem mass spectrometry.

We present a validated method for the simultaneous analysis of basic drugs which comprises a sample clean-up step, using mixed-mode solid-phase extraction (SPE), followed by LC-MS/MS analysis. Deuterated analogues for all of the analytes of interest were used for quantitation. The applied HPLC gradient ensured the elution of all the drugs examined within 14 min and produced chromatographic peaks of acceptable symmetry. Selectivity of the method was achieved by a combination of retention time, and two precursor-product ion transitions for the non-deuterated analogues. Oral fluid was collected with the Intercept, a FDA approved sampling device that is used on a large scale in the US for workplace drug testing. However, this collection system contains some ingredients (stabilizers and preservatives) that can cause substantial interferences, e.g. ion suppression or enhancement during LC-MS/MS analysis, in the absence of suitable sample pre-treatment. The use of the SPE was demonstrated to be highly effective and led to significant decreases in the interferences. Extraction was found to be both reproducible and efficient with recoveries >76% for all of the analytes. Furthermore, the processed samples were demonstrated to be stable for 48 h, except for cocaine and benzoylecgonine, where a slight negative trend was observed, but did not compromise the quantitation. In all cases the method was linear over the range investigated (2-200 microg/L) with an excellent intra-assay and inter-assay precision (coefficients of variation <10% in most cases) for QC samples spiked at a concentration of 4, 12 and 100 microg/L. Limits of quantitation were estimated to be at 2 microg/L with limits of detection ranging from 0.2 to 0.5 microg/L, which meets the requirements of SAMHSA for oral fluid testing in the workplace. The method was subsequently applied to the analysis of Intercept samples collected at the roadside by the police, and to determine MDMA and MDA levels in oral fluid samples from a controlled study.

Mixed-mode solid-phase extraction procedures for the determination of MDMA and metabolites in urine using LC-MS, LC-UV, or GC-NPD.

A solid-phase extraction (SPE) procedure was developed for the liquid chromatographic-mass spectrometric (LC-MS) analysis of 3,4-methylenedioxymethamphetamine (MDMA) and its metabolites 3,4-methylenedioxyamphetamine (MDA) and N,alpha-dimethyl-(3-methoxy-4-hydroxybenzene) (HMMA) ethanamine in urine. The procedure, with modifications, was also demonstrated using LC-UV and GC-nitrogen-phosphorus detection (NPD). A mixed-mode cation exchange SPE cartridge was effective for both reducing matrix impurities and for preconcentrating the analytes for the analysis. The concentration range investigated spanned from 0.10 to 20 microg/mL with recoveries ranging from 88% to 108% for all analytes using LC-MS. Both LC methods and the GC method allow for complete resolution of MDMA, MDA, and HMMA, as well as the internal standards (MDMA-d5 and 3,4-methylenedioxypropylamphetamine) in less than 10 min. Lower limits of quantitation (LLOQ) for the LC-MS method were 0.1 microg/mL for MDMA and MDA and 0.04 microg/mL for HMMA. Compared with LC-UV and GC-NPD, the LC-MS method requires the least amount of sample manipulation and provided the highest sample throughput. GC-NPD gave comparable selectivity, but the extra sample manipulation steps required contributed to lower recovery, lower precision, and increased time for the analysis. LC-MS analysis demonstrated better selectivity for the determination of HMMA in incurred samples compared with LC-UV analysis.