Pharmacy

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Investigation of polyphenol diversity among lentil species (Lens spp.) using mass spectrometry-based metabolomics guided by photodiode array detection
(ScienceDirect, 2025-03) Purves, Randy W.; Khazaei, Hamid; Elessawy, Fatma M.; Munro, Roger; Shurmer, Bryn O.; Vandenberg, Albert
Polyphenol diversity was investigated among seven lentil species, including Lens culinaris (cultivated lentil), L. orientalis, L. tomentosus, L. odemensis, L. lamottei, L. ervoides, and L. nigricans, using photodiode array detection coupled with liquid chromatography - mass spectrometry (LC-MS). Principal component analysis showed that most species grouped individually, except L. tomentosus and L. odemensis, which overlapped. The LC-MS data from both negative and positive electrospray ionization modes were used to identify 85 polyphenols observed in the UV–vis spectra, which included 27 proanthocyanidins, 17 flavonols, 15 flavones, and 12 hydroxybenzoic acids. An untargeted (comprehensive) analysis of the LC-MS data using Compound Discoverer software identified additional polyphenols (231 total), including numerous overlapping proanthocyanidins that contribute to a broad peak in the UV–vis spectra. The software analysis uncovered some notable differences among polyphenol profiles and intensities within the flavones, flavonols, and phenolic acids present in the species. This result indicates natural variation among the lentil wild relatives, which in part, is attributed to structurally isomeric compounds. A hierarchical clustering analysis, and a differential analysis using volcano plots used to look for statistically significant differences in polyphenols, illustrated significantly lower relative levels of polyphenols in L. culinaris compared with the wild types, especially within the proanthocyanidins and flavones. Our results highlight the potential of lentil wild relatives to enhance lentil seed quality.
Caffeine and Nicotine with N-Substituted Diazirine Photoaffinity Labels Form Adducts at Tyrosine-39 of α-Synuclein
(ACS Publications, 0002-04) Mejia-Gutierrez, Melissa; Moser, Brigitte; Pirlot, Marissa; Zhang, Haixia; Chumala, Paulos; Katselis, George S.; Palmer, David R.; Krol, Ed S.
Aggregates of the protein α-synuclein are found in Lewy bodies in the brains of Parkinson’s disease (PD) patients. Small molecules that can attenuate or halt α-synuclein aggregation have been studied as potential therapeutics for PD. However, we have a limited understanding of how these molecules bind to α-synuclein. We previously identified that caffeine, nicotine, and 1-aminoindan all bind to both the N- and C-terminus of α-synuclein, although the binding location remains unknown. In an effort to identify these binding regions on α-synuclein, we synthesized diazirine photoaffinity probes attached to caffeine (C-Dz), nicotine (N-Dz), and 1-aminoindan (I-Dz) and allowed each to react with α-synuclein in vitro. We then treated the incubation mixture with trypsin and employed time-of-flight mass spectrometry to analyze the resulting peptides. Our findings reveal a distinctive binding pattern among the probes: C–Dz forms covalent bonds with Tyr-39 and Glu-20, while N-Dz selectively forms a covalent bond with Tyr-39. Intriguingly, we could not detect the labeling of I-Dz to any specific amino acids. All of the diazirine-bound peptides were found near the N-terminus. Our results suggest that the N-terminal region near Tyr-39 bears further study to elucidate the binding interactions of small molecules with α-synuclein and may be a target for anti-PD agents.
Melanin from the fungus Gliocephalotrichum simplex protects seeds from the effects of exposure to gamma radiation
(Nature, 2025) Kunal, Swaraj; Bhanushali, Sushrut; Raghukumar, Seshagiri; Dadachova, Ekaterina
Protection of seeds from ionizing radiation is an important future need in space travel. We examined the usefulness of eumelanin from the fungus Gliocephalotrichum MTCC 5489 for this purpose. Seeds of Oryza sativa, Brassica nigra, Vigna radiata and V. aconitifolia were enclosed in Petri dishes coated with paint containing various concentrations of nanomelanin and exposed to doses of 0.1 to 2.0 kGy Cs-137 or 0.1 to 0.5 kGy Co-60 radiation. While Cs-137 radiation severely affected the rate of germination (germination index, GI) and germination percentage of Oryza sativa, whereas other seeds were less affected, Co-60 markedly diminished the GI of all the seeds. Increasing concentrations of melanin coating afforded significant protection to the GI of O. sativa seeds exposed to Cs-137 and to all seeds exposed to Co-60 radiation. The germination percentage of seeds was dose- and radiation dependent, with no effect observed for all doses of Cs-137 or 0.1 kGy Co-60 radiation, whereas 0.5 kGy Co-60 radiation killed 50 to 60% of the unprotected seeds. Melanin paint offered nearly 100% survival. This study demonstrated that melanin-containing materials can provide effective shielding from gamma radiation for diverse types of seeds that are important for human consumption, which has implications for space agriculture and agriculture in extreme environments.
Effect of nutrition education integrating the health belief model and theory of planned behavior during pregnancy on gestational weight gain and birth weight in Southeast Ethiopia using complex analyses
(Springer Nature, 2025) Beressa, Girma; Whiting, Susan; Belachew, Tefera
Background The incidence of inadequate or excessive gestational weight gain (GWG) is an indicator of reproductive health problems. However, scientific evidence for the effect of nutrition education during pregnancy on GWG and neonatal birth weight (BW) in urban settings in Ethiopia is sparse. This study aimed to assess the effect of nutrition education during pregnancy on GWG and neonatal birth weight (BW) in urban settings in Southeast Ethiopia. Methods A community-based two-arm parallel cluster randomized controlled trial was conducted among 447 randomly selected pregnant women attending antenatal care (224 intervention and 223 control) from February to December 2021. Study participants were selected by multistage cluster sampling followed by systematic sampling. Women receiving the intervention received six nutrition education sessions, while women in the control group received standard care. GWG was the difference between the last recorded weight before delivery and the weight recorded during the first trimester. BW was measured within the first hour of delivery. The generalized structural equations model (GSEM) and structural equations model (SEM) were used to examine the direct, indirect, and total effects of nutrition education on GWG and BW via the dietary diversity score (DDS), food security (FS), and knowledge. Results The GSEM revealed that receiving intervention during pregnancy had a total effect on GWG [(AOR = 2.056, 95% CI: 1.705, 2.695)]. Having dietary diversity had direct and total effects on GWG [(AOR = 1.105, 95% CI: 1.022, 1.196)]. Having food security had a total effect on GWG [(AOR = 1.928, 95% CI: 1.817, 2.052)]. Having fruit and vegetable knowledge had a total effect on GWG [(AOR = 1.971, 95% CI: 1.856, 2.105)]. The SEM revealed that receiving intervention during pregnancy had a direct effect on BW (unstandardized β = 0.144, 95% CI: 0.034, 0.252). Similarly, it revealed that receiving intervention during pregnancy had a direct effect on DDS (β = 0.580, 95% CI: 0.024, 1.038). Likewise, it indicated that receiving intervention during pregnancy had a total effect on increasing BW (β = 0.137, 95% CI: 0.029, 0.243). Nevertheless, there was no statistically observed indirect effect of nutrition education during pregnancy on GWG and BW via mediators. Conclusion The SEM revealed that receiving nutrition education interventions during pregnancy had a total effect on GWG and direct and total effects on BW. The generalized structural equation modelling (GSEM) and structural equation modelling (SEM) findings show that integrating theory-based nutrition education during pregnancy will improve gestational weight gain (GWG) and birth weight (BW) in Ethiopia.
Prescription for change: Unveiling burnout perspectives among pharmacy leaders
(Canadian Pharmacists Journal, 2024-11) Kiriazopoulos, Stefanie; Perepelkin, Jason; Alford, Heather
Background: Burnout among pharmacists is increasingly pertinent, with growing demand for effective interventions. Burnout can lead to reduced productivity, increased job turnover, medical errors, poor patient satisfaction, and other negative outcomes for patients and providers. Growing attention to burnout in the pharmacy profession highlights the need for personal, organizational, and systemic solutions. However, the uptake and relative efficacy of different approaches remain unclear, particularly within community pharmacy practice. This study sought the viewpoint of community pharmacy leaders (i.e., community pharmacy managers, district managers, franchisees, owners, and executives from various pharmacy organizations) to characterize burnout from their perspectives. Methods: This qualitative study followed a grounded theory approach. Community pharmacy leaders were interviewed using a semistructured format to gather in-depth insights into their experiences and perspectives on burnout and engagement. Results: Sixteen people were interviewed; interviews lasted 30 to 65 minutes, averaging 51 minutes long. Six themes were identified: perceived disconnection between front-line staff and pharmacy decision-makers, overwhelming work demands, cautious optimism toward the expanding scope of pharmacy practice, the importance of employee recognition and appreciation, appropriateness and use of existing work resources, and multimodal, systemic responsibility and solutions to burnout. Conclusion: Addressing burnout requires a multifaceted approach involving personal, organizational, and systemic interventions. Evidence from this study provides valuable insights into the feasibility and efficacy of specific interventions, informing future strategies to enhance workplace well-being and engagement. The study highlights the importance of managing job demands and maximizing resources, emphasizing that personal approaches alone are insufficient and that organizational and systemic interventions are crucial.
Inside the journey: A qualitative study of intravenous ketamine therapy for treatment-resistant depression
(Journal of Affective Disorders Reports, 2025-02) Perepelkin, Jason; Hantjidis, Polixeni; Robison, Kirsten; Halpape, Katelyn
Introduction Treatment-resistant depression (TRD) affects over 30 % of patients with depression and is associated with high morbidity and mortality. Intravenous (IV) ketamine has shown rapid and robust antidepressant effects for TRD. However, limited data exist on long-term maintenance and patient experiences, especially in North America. This study aims to explore the lived experiences of individuals receiving IV ketamine for TRD in Saskatchewan, Canada. Methods This qualitative study involved semi-structured interviews with 19 individuals receiving IV ketamine at the Linden Medical Centre. Participants were included if they were over 18 years of age and received IV ketamine for TRD. Data were analyzed using grounded theory methodology to identify emerging themes. Results Key themes included the acute effects of ketamine during treatment, which varied among patients but generally involved psychological relief and altered perceptions. Participants noted significant improvements in mood, outlook on life, and daily functioning. However, barriers such as treatment cost, accessibility, and stigma were prevalent. Psychosocial factors and the clinic environment also substantially influenced treatment outcomes. Interpretation The findings highlight the profound impact of IV ketamine on patients with TRD, emphasizing the importance of a supportive clinic environment and addressing barriers to accessibility. Despite the financial burden and limited accessibility, ketamine treatments resulted in meaningful improvements, including reduced suicidality. Conclusion This study underscores the need for further research on individual predictors of ketamine response, cost-effectiveness, and educational materials to set realistic patient expectations. Understanding patient experiences can help optimize ketamine therapy for TRD, ultimately enhancing treatment outcomes.
Integration of medicinal chemistry in therapeutic decision-making: A way forward?
(Currents in Pharmacy Teaching and Learning, 2024) Krol, Ed; Albon, Simon P
Many attempts have been made to integrate medicinal chemistry knowledge into therapeutic decision-making in pharmacy programs across North America. Examples include the use of Structure-Based Therapeutic Evaluations, alignment of medicinal chemistry content with courses in pharmacology, pharmaceutics and pharmacotherapeutics, and team-based or problem-based learning methods. The majority of these approaches indicate that students have greater confidence or comfort with medicinal chemistry, but there remain few cases where an improvement in performance has been measured. This is especially challenging for assessing a student’s ability to implement medicinal chemistry learning in pharmacotherapeutic decision-making. Building upon our national special interest group’s recent environmental scan of medicinal chemistry instruction in Canadian Pharmacy programs, we are investigating strategies for integration of medicinal chemistry learning in therapeutic decision-making. This commentary will discuss methods and evidence to support medicinal chemistry integration, relevant assessment strategies, and potential paths forward.
Medical cannabis in schools: The experiences of caregivers
(Oxford University Press, 2023-05) Mansell, Holly; Zaslawski, Zina; Mbabaali, Sophia; King, Patricia M.; Kelly, Lauren E.; Lougheed, Taylor; Anderson, Jennifer; Huntsman, Richard J.; Alcorn, Jane
Abstract Objectives Implementing medical cannabis (MC) into a child’s daily routine can be challenging and there is a lack of guidance for its therapeutic use in schools in Canada. Our objective was to learn about the experiences of caregivers of school-aged children who require MC. Methods Qualitative description was used and caregivers were interviewed about MC in schools and in general. The transcripts were entered into Dedoose software for qualitative analysis and content analysis was performed. Sentences and statements were ascribed line by line into meaning units and labelled with codes, and organized according to categories and subcategories. Results Twelve caregivers of school-aged children who take MC participated. The most common reasons for treatment were drug-resistant epilepsy (DRE), autism, or other developmental disorders. Approximately half of the participants’ children (n = 6) took MC during the school day and most (5/6) perceived their experiences to be positive or neutral but reported a lack of knowledge about MC. While data saturation was not reached regarding MC in schools, rich dialogues were garnered about MC in general and three categories were identified: challenges (subcategories stigma, finding an authorizer, cost, dosing, and supply); parents as advocates (subcategories required knowledge, attitudes, skills, and sources of information); and caregiver relief for positive outcomes. Conclusions Caregivers demonstrate remarkable tenacity despite the many challenges associated with MC use. Education and practice change are needed to ensure that children using MC can benefit from or continue to experience its positive outcomes within the school environment and beyond.
Nutritional Status of Preschool Children (aged 24 TO 59 months) and Chickpea Production and Consumption in Ethiopia
(African Journal of Food, Agriculture, Nutrition and Development, 2024-10) Asrat, R. T; Girma, M; Tafesse, E; Whiting, S. J; Henry, Carol
Child under-nutrition is a major public health concern in developing countries including Ethiopia. Poor quality diet is the major determinant factor contributing to child undernutrition. Pulse crop production and consumption can improve children’s nutrient quality and dietary diversity. This study aimed to compare the nutritional status and child health, dietary intake, and wealth index of preschool children among pulse crop producer and non-producer families in Ethiopia. The study hypothesized that children living in families who grew pulses and consumed pulses (rather than selling their crop) would have better nutritional status due to availability of this protein-rich food source. A cross-sectional survey was conducted to determine demographic background, child feeding and caring practice and varieties and amount of pulse crop produced at the household level. A simple random sampling technique was used to select a sample of 432 children aged 24-59 months from central Ethiopia the Data was entered to SPSS v 20, WHO Anthro-plus software used to calculate the anthropometry indices, child dietary intake, level of chickpea production and consumption and wealth index, were computed using logistic regression. Chickpea (cicer arietinum L) was produced by 55.3% of the participants. Most (60.7%) of the children from chickpea producer families were high pulse consumers whereas only 15.5% of children from non-producers were high chickpea consumers (the difference in proportion is significant at p-value<0.001). Close to 60% of the children of the non-producer families had a low dietary diversity score (that is <3) compared with 18.8 % from the producer families (p-value <0.001). The study revealed that nutritional status of children in both producer and non-producer families were 41.0 % stunted, 16.1 % underweight and 6.1 % wasted. Stunting and underweight were significantly inversely associated with chickpea production at pvalue <0.001whereas for chickpea consumption, both stunting and underweight was significant at P < 0.05. However, wasting was not significantly associated with either production or consumption of chickpeas. In conclusion, chickpea production and consumption by families had a positive relationship with nutritional status of children. The authors recommend that engaging smallholder farmers in both chickpea production and consumption has potential in improving the dietary diversity and in turn benefit nutritional status of preschool children at the household level and therefore supporting smallholder farmer in chickpea production and consumption will help to contribute to decreasing the prevalence of under nutrition in the country.
Two-Spirit Peoples’ experiences accessing and receiving care from community pharmacies
(SAGE Publications, 2024) Marissa Pirlot; Swidrovich, Jaris
Background: Two-Spirit Peoples face unique challenges in accessing and receiving health care in Canada due to health services, including community pharmacy services, being built on hetero- and cis-normative models that impede appropriate care for this group. Currently, there is limited published information on Two-Spirit Peoples’ experiences accessing and receiving care in community pharmacy settings. Methods: To address the lack of published information, 21 Two-Spirit individuals shared their experiences in a focus group setting. Four different focus groups were held across Canada, including 1 in Saskatoon, Vancouver, Edmonton, and Toronto. Informed by Indigenous methodologies, data were recorded via audio recording and notetaking, and the audio was transcribed and then analyzed for themes using the Voice-Centred Relational Method. Results: Three major structural systems that affect the experiences of Two-Spirit Peoples in community pharmacies were identified: 1) white supremacy, 2) capitalism, and 3) heteronormativity. These 3 systemic issues presented themselves via racism, homophobia, transphobia, pharmacists’ lack of knowledge about Two-Spirit individuals and their health and lack of time spent educating or building relationships with Two-Spirit Peoples. Participants provided suggestions for how community pharmacists can better serve the Two-Spirit community, such as using inclusive language, adding pronouns and preferred names to patient files, increasing knowledge about Two-Spirit health and advocating for Two-Spirit Peoples. Discussion: The results suggest that dismantling current structures and ideologies in community pharmacy and society are required to overcome the identified issues. Conclusion: Two-Spirit Peoples face barriers when it comes to accessing and receiving care in community pharmacies, resulting in many Two-Spirit individuals avoiding health care to save themselves from unsafe and uncomfortable interactions. Pre- and postlicensure pharmacy education about Two-Spirit Peoples is required to improve Two-Spirit Peoples’ experiences accessing and receiving care in community pharmacies.
Cannabinoid Therapy in Athletics: A Review of Current Cannabis Research to Evaluate Potential Real-World Cannabinoid Applications in Sport
(Springer, 2024-08-21) Thompson, Elizabeth S.; Alcorn, Jane; Neary, J. Patrick
The increasing legalization of Cannabis sativa plant products has sparked growing interest in their therapeutic applications. Prohibition laws established in 1937 hindered formal research on cannabis, a plant with cultural and medicinal roots dating back to 2700 BC in Chinese history. Despite regulatory hurdles, published research on cannabis has emerged; yet elite athletes remain an underrepresented population in these studies. Athletes, known for exploring diverse substances to optimize performance, are drawn to the potential benefits of cannabinoid therapy, with anecdotal reports suggesting positive effects on issues ranging from anxiety to brain injuries. This review aims to evaluate empirical published cannabis research with a specific focus on its potential applications in athletics. The changing legal landscape, especially the removal of cannabis from drug testing programs in leagues such as the National Basketball Association (NBA), and endorsements by Major League Baseball (MLB) for cannabinoid products and the National Football League (NFL) for cannabis research, reflects a shift in the acceptability of such substances in sports. However, stigma, confusion, and a lack of education persist, hindering a cohesive understanding among sports organizations, including business professionals, policymakers, coaches, and medical/training staff, in addition to athletes themselves. Adding to the confusion is the lack of consistency with cannabinoid regulations from sport to sport, within or out of competition, and with cannabis bioactive compounds. The need for this review is underscored by the evolving attitudes toward cannabinoids in professional sports and the potential therapeutic benefits or harms they may offer. By synthesizing current cannabis research, this review aims to provide a comprehensive understanding of the applications and implications of cannabinoid use in the realm of athletics.
Development of a liquid chromatography-tandem mass spectrometry method for the analysis of docetaxel-loaded Poly(lactic-co-glycolic acid) nanoparticles
(2023) Khajavinia, Amir; El-Aneed, Anas
Docetaxel is among the most effective chemotherapeutic agents used for the treatment of solid tumors, such as breast cancer. Targeting docetaxel to the tumor site would increase the safety and efficacy of the treatment. The focus of this work was to develop an efficient liquid chromatography tandem mass spectrometry (LC-MS/MS) method to quantify docetaxel entrapped in optimized poly lactic-co-glycolic acid (PLGA) nanoparticles. Several nanoparticle formulations were prepared to optimize the nanoparticles based on their size and yield percentage using a modified solvent evaporation technique. The MS/MS fingerprints of docetaxel and paclitaxel (as internal standard) were used to identify diagnostic product ion for developing a multiple reaction monitoring (MRM) LC-MS/MS method for the quantification of docetaxel in the PLGA nanoparticles. A triple quadrupole linear ion trap instrument (AB Sciex 4000 QTRAP) equipped with electrospray ionization was used. The optimized nanoparticles had a zeta potential of -23.2 ± 1.4 mV and mean particle sizes of 202.2 ± 4.7 nm and 251.7 ± 8.2 nm before and after freeze-drying, respectively. Polydispersity index values of the nanoparticles confirmed their uniform size distribution. The developed LC-MS/MS method could quantify docetaxel in the PLGA matrix with accuracy and precision covering a broad linear range of 15.6 to 4000 ng/mL. Method validation was conducted using the regulatory guidelines of the Food and Drug Administration (FDA) and the European Medicines Agency (EMA) and showed acceptable values for all the tested criteria. The developed LC-MS/MS method with the novelty of using a phenyl column will be beneficial for future analysis of docetaxel loaded polymeric nano-delivery systems.
Carbon-based nanoparticles and their surface-modified counterparts as MALDI matrices
(2023) Khajavinia, Amir; El-Aneed, Anas
The Effect of Diphenylethane Side-chain Substituents on Dibenzocyclohexadiene Formation and their Inhibition of α-Synuclein Aggregation in vitro
(2023-01-15) Dalio Bernardes da Silva, Gabriel; Munir, Omer; Krol, Ed
The naturally-occurring di-catechol lignan nordihydroguaiaretic acid (NDGA) and an analog without methyl groups on the butyl linker both undergo intramolecular cyclization at pH 7.4 to form dibenzocyclooctadienes. Both NDGA and these dibenzocyclooctadienes have been shown to prevent in vitro aggregation of α-synuclein, an intrinsically disordered protein associated with Parkinson's disease. NDGA possesses two vicinal methyl groups on the butyl linker and the presence of these methyl groups attenuates the rate of intramolecular cyclization versus the unsubstituted analog, in opposition to the anticipated Thorpe-Ingold effect, likely due to steric repulsions during cyclization. Numerous 1,2-bis-ethane di-catechols are known to inhibit α-synuclein aggregation in vitro and we hypothesize that these compounds undergo a similar intramolecular cyclization and the cyclized products may be responsible for the activity. To test this hypothesis we prepared a series of 1,2-bis-ethane di-catechols with 0, 2 and 4 methyl substituents on the linker. We have confirmed that these compounds undergo intramolecular cyclization to form dibenzocyclohexadienes and that steric interactions between the methyl substituents leads to an increase in the rate of intramolecular cyclization, which is in contrast to what was observed for lignan di-catechols. The rate of cyclization to form six-membered rings is 10-30 times more rapid than formation of eight membered rings and the dibenzocyclohexadienes also prevent in vitro aggregation of α-synuclein.
Tandem mass spectrometric analysis of novel caffeine scaffold-based bifunctional compounds for Parkinson's disease
(2019) Nwabufo, Chukwunonso; El-Aneed, Anas; Krol, Ed
Rationale: Novel bifunctional compounds composed of a caffeine scaffold attached to nicotine (C8-6-N), 1-aminoindan (C8-6-I), or caffeine (C8-6-C8) were designed as therapeutics or diagnostics for Parkinson's disease (PD). In order to probe their pharmacological and toxicological profile, an appropriate analytical method is required. The goal of this study is to establish a tandem mass spectrometric fingerprint for the development of quantitative and qualitative methods that will aid future assessment of the in vitro and in vivo absorption, distribution, metabolism, excretion (ADME) and pharmacokinetic properties of these lead bifunctional compounds for PD. Methods: Accurate mass measurement was performed using a hybrid quadrupole orthogonal time-of-flight mass spectrometer while multistage MS/MS and MS3 analyses were conducted using a triple quadrupole linear ion trap mass spectrometer. Both instruments are equipped with an electrospray ionization (ESI) source and were operated in the positive ion mode. The source and compound parameters were optimized for all three tested bifunctional compounds. Results: The MS/MS analysis indicates that the fragmentation of C8-6-N and C8-6-I is driven by the dissociation of the nicotine and 1-aminoindan moieties, respectively, but not caffeine. A significant observation in the MS/MS fragmentation of C8-6-C8 suggests that a previously reported loss of acetaldehyde during caffeine dissociation is instead a loss of CO2. Conclusions: The collision-induced tandem mass spectrometry (CID-MS/MS) analysis of these novel bifunctional compounds revealed compound-specific diagnostic product ions and neutral losses for all three tested bifunctional compounds. The established MS/MS fingerprint will be applied to the future development of qualitative and quantitative methods.
Mass Spectrometric Detection and Characterization of Metabolites of Gemini Surfactants Used as Gene Delivery Vectors
(2020) Jin, Wei; Purves, Randy; Krol, Ed; Badea, ildiko; El-Aneed, Anas
Gemini surfactants are a class of lipid molecules that have been successfully used in vitro and in vivo as non-viral gene delivery vectors. However, the biological fate of gemini surfactants has not been well investigated. In particular, the metabolism of gemini surfactants after they enter cells as gene delivery vehicles is unknown. In this work, we used a high-resolution quadrupole-Orbitrap mass spectrometry (Q-Exactive®) instrument to detect the metabolites of three model gemini surfactants, namely a) unsubstituted (16-3-16), b) with pyridinium head groups (16(Py)-S-2-S-16(Py)), and c) substituted with a glycyl-lysine di-peptide (16-7N(GK)-16). The metabolites were characterized, and structures proposed, based on accurate masses and characteristic product ions. The metabolism of the three gemini surfactants was very different as 16-3-16 was not metabolized in PAM212 cells, whereas 16(Py)-S-2-S-16(Py) was metabolized primarily via phase I reactions, including oxidation and de-alkylation, producing metabolites that could be linked to its observed high toxicity. The third gemini surfactant 16-7N(GK)-16 was metabolized mainly via phase II reactions, including methylation, acetylation, glucose conjugation, palmityl conjugation, and stearyl conjugation. The metabolism of gemini surfactants provides insight for future directions in the design and development of more effective gemini surfactants with lower toxicity. The reported approach can also be applied to study the metabolism of other structurally related gemini surfactants.
Cellular Uptake and Distribution of Gemini Surfactant Nanoparticles Used as Gene Delivery Agents
(SpringerLink, 2019-08-06) Jin, Wei; Al-Dulaymi, Mays; Badea, Ildiko; Leary, Scot; Rehman, Jeveria; El-Aneed, Anas
Gemini surfactants are promising molecules utilized as non-viral gene delivery vectors. However, little is known about their cellular uptake and distribution after they release their therapeutic cargo. Therefore, we quantitatively evaluated the cellular uptake and distribution of three gemini surfactants: unsubstituted (16-3-16), with pyridinium head groups (16(Py)-S-2-S-16(Py)) and substituted with a glycyl-lysine di-peptide (16-7N(GK)-16). We also assessed the relationship between cellular uptake and distribution of each gemini surfactant and its overall efficiency and toxicity. Epidermal keratinocytes PAM 212 were treated with gemini surfactant nanoparticles formulated with plasmid DNA and harvested at various time points to collect the enriched nuclear, mitochondrial, plasma membrane, and cytosolic fractions. Gemini surfactants were then extracted from each subcellular fraction and quantified using a validated flow injection analysis-tandem mass spectrometry (FIA-MS/MS) method. Mass spectrometry is superior to the use of fluorescent tags that alter the physicochemical properties and pharmacokinetics of the nanoparticles and can be cleaved from the gemini surfactant molecules within biological systems. Overall, a significantly higher cellular uptake was observed for 16-7N(GK)-16 (17.0%) compared with 16-3-6 (3.6%) and 16(Py)-S-2-S-16(Py) (1.4%), which explained the relatively higher transfection efficiency of 16-7N(GK)-16. Gemini surfactants 16-3-16 and 16(Py)-S-2-S-16(Py) displayed similar subcellular distribution patterns, with major accumulation in the nucleus, followed by the mitochondrion, cytosol, and plasma membrane. In contrast, 16-7N(GK)-16 was relatively evenly distributed across all four subcellular fractions. However, accumulation within the nucleus after 5 h of treatment was the highest for 16(Py)-S-2-S-16(Py) (50.3%), followed by 16-3-16 (41.8%) and then 16-7N(GK)-16 (33.4%), possibly leading to its relatively higher toxicity.
Fast Quantification Without Conventional Chromatography, The Growing Power of Mass Spectrometry
(ACS Publications, 2020-06-08) gachumi, george; Purves, Randy; Hopf, Carston; El-Aneed, Anas
Mass spectrometry (MS) in hyphenated techniques is widely accepted as the gold standard quantitative tool in life sciences. However, MS possesses intrinsic analytical capabilities that allow it to be a stand-alone quantitative technique, particularly with current technological advancements. MS has a great potential for simplifying quantitative analysis without the need for tedious chromatographic separation. Its selectivity relies on multistage MS analysis (MSn), including tandem mass spectrometry (MS/MS), as well as the ever-growing advancements of high-resolution MS instruments. This perspective describes various analytical platforms that utilize MS as a stand-alone quantitative technique, namely, flow injection analysis (FIA), matrix assisted laser desorption ionization (MALDI), including MALDI-MS imaging and ion mobility, particularly high-field asymmetric waveform ion mobility spectrometry (FAIMS). When MS alone is not capable of providing reliable quantitative data, instead of conventional liquid chromatography (LC)-MS, the use of a guard column (i.e., fast chromatography) may be sufficient for quantification. Although the omission of chromatographic separation simplifies the analytical process, extra procedures may be needed during sample preparation and clean-up to address the issue of matrix effects. The discussion of this manuscript focuses on key parameters underlying the uniqueness of each technique for its application in quantitative analysis without the need for a chromatographic separation. In addition, the potential for each analytical strategy and its challenges are discussed as well as improvements needed to render them as mainstream quantitative analytical tools. Overcoming the hurdles for fully validating a quantitative method will allow MS alone to eventually become an indispensable quantitative tool for clinical and toxicological studies.
Employing in vitro metabolism to guide design of F-labelled PET probes of novel alpha-synuclein binding bifunctional compounds
(Taylor & Francis, 2021-06-30) Nwabufo, Chukwunonso; Aigbogun, Omozojie; Allen, Kevin; Madeline N. Owens; Jeremy S. Lee; Christopher P. Phenix; Krol, Ed
1. A challenge in the development of novel 18F-labelled positron emission tomography (PET) imaging probes is identification of metabolically stable sites to incorporate the 18F radioisotope. Metabolic loss of 18F from PET probes in vivo can lead to misleading biodistribution data as displaced 18F can accumulate in various tissues. 2. In this study we report on in vitro hepatic microsomal metabolism of novel caffeine containing bifunctional compounds (C8-6-I, C8-6-N, C8-6-C8) that can prevent in vitro aggregation of -synuclein, which is associated with the pathophysiology of Parkinson’s disease. The metabolic profile obtained guided us to synthesize stable isotope 19F-labelled analogues in which the fluorine was introduced at the metabolically stable N7 of the caffeine moiety. 3. An in vitro hepatic microsomal metabolism study of the 19F-labelled analogues resulted in similar metabolites to the unlabelled compounds and demonstrated that the fluorine was metabolically stable, suggesting that these analogues are appropriate PET imaging probes. This straightforward in vitro strategy is valuable for avoiding costly stability failures when designing radiolabelled compounds for PET imaging.
The Health-Care Provider’s Perspective of Education Before Kidney Transplantation
(Sage, 2016-08-22) Trivedi, Paraag; Rosaasen, Nicola; Mansell, Holly
CONTEXT: Adequate patient education is essential for preparing potential recipients for kidney transplantation. Health-care providers play a vital role in education and can identify gaps in patient understanding. OBJECTIVE: To identify deficits in patient knowledge from the perspective of a transplant multidisciplinary care team and determine whether their perceptions align with patients who have previously undergone a transplant. DESIGN: An open call was advertised for health-care providers to attend a focus group discussion regarding the educational needs of pretransplant patients in 1 Canadian center. A predetermined, semistructured set of questions was used to collect the views of transplant caregivers. A moderator, assistant moderator, and research assistant facilitated the discussion, which was transcribed verbatim. Paper surveys were distributed to collect opinions of those unable to attend or uncomfortable to voice their opinion in an open forum. Qualitative analysis software was used to identify any emergent themes. Results were compared to a previous study undertaken in transplant recipients. RESULTS: Despite pre- and posttransplant education, specific themes emerged including misconceptions about the assessment process and time on the wait list and the surgery, incongruency between patient expectations and outcome, and confusion regarding medications. Health-care provider perceptions were remarkably consistent with transplant recipients. CONCLUSION: Health-care providers identified gaps in patient understanding indicating that transplant candidates may not be internalizing what is taught. Innovative educational approaches may be needed to provide more successful patient education. Similarities between health-care provider and patient perceptions suggest that care providers are a valuable source of information.

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