Interprofessional Education to Address Substance Use among Adults with Persistent Pain: A Pre-Post Program Evaluation.

BACKGROUND: Substance use disorders (SUDs) are highly prevalent among adults with persistent pain. Yet, standard competencies for integrating pain and SUD content are lacking across health science student curricula. Additionally, pharmacotherapies to treat SUDs are underutilized. AIM: To address these gaps, a team of health science faculty created an interprofessional simulation activity using a standardized patient and evaluated learner outcomes related to assessment and treatment of comorbid persistent pain and substance use. METHODS: A total of 304 health science students representing nursing, medicine, pharmacy, and social work programs attended virtual learning sessions. Interprofessional student teams developed a team-based care plan for an adult with musculoskeletal pain who takes prescribed opioids while using alcohol. Pre- and post-activity surveys assessing knowledge and confidence were matched for 198 students. Descriptive statistics summarized survey data with inferential analysis of paired data. RESULTS: The largest significant improvements between pre- and post-activity knowledge were observed in items specific to pharmacotherapy options for alcohol and opioid use disorders. Similar gains were noted in students' confidence regarding pharmacotherapies. No significant differences were noted on pre-post-activity knowledge scores between the three main profession groups (medicine, nursing, and pharmacy). CONCLUSIONS: Students attending this interprofessional simulation demonstrated improved knowledge and confidence, particularly in pharmacotherapies for alcohol and opioid use disorders. Replication of such programs can be used to provide consistent content across health science disciplines to heighten awareness and receptivity to medications available to treat SUDs in people treated for persistent pain. The curriculum is freely available from the corresponding author.

Examination of Urinary Excretion of Unchanged Drug in Humans and Preclinical Animal Models: Increasing the Predictability of Poor Metabolism in Humans.

PURPOSE: A dataset of fraction excreted unchanged in the urine (fe) values was developed and used to evaluate the ability of preclinical animal species to predict high urinary excretion, and corresponding poor metabolism, in humans. METHODS: A literature review of fe values in rats, dogs, and monkeys was conducted for all Biopharmaceutics Drug Disposition Classification System (BDDCS) Class 3 and 4 drugs (n=352) and a set of Class 1 and 2 drugs (n=80). The final dataset consisted of 202 total fe values for 135 unique drugs. Human and animal data were compared through correlations, two-fold analysis, and binary classifications of high (fe ≥30%) versus low (<30%) urinary excretion in humans. Receiver Operating Characteristic curves were plotted to optimize animal fe thresholds. RESULTS: Significant correlations were found between fe values for each animal species and human fe (p<0.05). Sixty-five percent of all fe values were within two-fold of human fe with animals more likely to underpredict human urinary excretion as opposed to overpredict. Dogs were the most reliable predictors of human fe of the three animal species examined with 72% of fe values within two-fold of human fe and the greatest accuracy in predicting human fe ≥30%. ROC determined thresholds of ≥25% in rats, ≥19% in dogs, and ≥10% in monkeys had improved accuracies in predicting human fe of ≥30%. CONCLUSIONS: Drugs with high urinary excretion in animals are likely to have high urinary excretion in humans. Animal models tend to underpredict the urinary excretion of unchanged drug in humans.

Pharmacologic Activities of 3'-Hydroxypterostilbene: Cytotoxic, Anti-Oxidant, Anti-Adipogenic, Anti-Inflammatory, Histone Deacetylase and Sirtuin 1 Inhibitory Activity.

PURPOSE: Delineate the selected pharmacodynamics of a naturally occurring stilbene 3'-Hydroxypterostilbene. OBJECTIVE: Characterize for the first time the pharmacodynamics bioactivity in several in-vitro assays with relevant roles in heart disease, inflammation, cancer, and diabetes etiology and pathophysiology. METHODS: 3'-Hydroxypterostilbene was studied in in-vitro assays to identify possible bioactivity. RESULTS: 3'-Hydroxypterostilbene demonstrated anti-oxidant, anti-inflammatory, cytotoxic, anti-adipogenic, histone deacetylase, and sirtuin-1 inhibitory activity. CONCLUSIONS: The importance of understanding individual stilbene pharmacologic activities were delineated. Small changes in chemical structure of stilbene compounds result in significant pharmacodynamic differences. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Preclinical Pharmacokinetics and Pharmacodynamics and Content Analysis of Gnetol in Foodstuffs.

Studies were undertaken to evaluate the bioavailability in rats and content analysis of gnetol in Gnetum gnemon products reported to contain gnetol and to examine the pharmacological properties of gnetol in in vitro models including anti-inflammatory/analgesic, antidiabetic, anti-adipogenesis, and anticancer activity. Male Sprague-Dawley rats were cannulated and dosed either intravenously with gnetol (10 mg/kg) or orally (100 mg/kg). Various methanolic extractions of G. gnemon products were quantified. Gnetol's effect on cell viability in selected cell lines with or without inflammatory stimulus was assessed. α-Amylase and α-glucosidase inhibition was evaluated. Cyclooxygenase (COX)-1, COX-2, and histone deacetylase inhibition and adipogenesis inhibition were examined. After oral and intravenous administration, gnetol was detected in both serum and urine as the parent compound and as a glucuronidated metabolite. The bioavailability of gnetol was determined to be 6%. Gnetol is rapidly glucuronidated and is excreted in urine and via nonrenal routes. Gnetol was found to exist as an aglycone and as a glycoside in G. gnemon products. Gnetol showed concentration-dependent reductions in cell viability in cancer cell lines with greatest activity in colorectal cancer and potent COX-1, histone deacetylase, and weak COX-2 activities along with limited reduction in inflammation. Gnetol also possessed concentration-dependent alpha-amylase, alpha-glucosidase, and adipogenesis activities. Pretreatment of mice with gnetol was able to increase the latency period to response in analgesia models.

An Interactive Online Interprofessional Opioid Education Training Using Standardized Patients.

Introduction: Opioid pain management is complex and requires a collaborative approach. To prepare health professions students to care for patients who have chronic pain, we developed an interprofessional education (IPE) session for delivery using a virtual platform that featured a standardized patient (SP) interaction. Methods: The SP case highlighted a patient on opioids for chronic low back pain resulting from a car accident. Despite no improvement in pain or function, the patient continued taking opioids and developed behaviors that could represent opioid misuse. During the synchronous, online session, interprofessional teams of students interviewed an SP and collaborated to develop a holistic care plan to address the patient's pain and potential opioid misuse. The session evaluation included pre- and postsession surveys. Results: Over 750 students from medicine, pharmacy, nursing, and social work programs participated in the virtual IPE sessions during a single year. Students rated the session positively. Matched survey responses suggested improved confidence in knowledge and skills, and learning through Zoom was rated favorably. Discussion: We successfully implemented a synchronous online IPE session involving SP interactions that allowed students to practice team-based care of a patient with chronic pain who was taking opioids. Based on the success of this IPE session, including the success of the online delivery model, future IPE sessions will continue virtually.

High-performance liquid chromatographic analysis of lacosamide in canine serum using ultraviolet detection: application to pre-clinical pharmacokinetics in dogs.

A method for analysis of lacosamide [(R)-2-acetamido-N-benzyl-3-methoxypropionamide] is needed for both human and veterinary pharmacokinetic investigations. While lacosamide is currently used to manage partial-onset seizures in humans suffering from epilepsy, it is also presently being investigated for use in the treatment of canine epilepsy in veterinary medicine. Currently, no dosing regimen for the drug exists in dogs. A novel and simple high-performance liquid chromatography method was developed for determination of lacosamide in dog serum. Serum proteins (0.1 mL) were precipitated with -20.0°C acetonitrile after addition of the internal standard, daidzein. Separation was achieved with a Phenomenex® Luna® C18 (2) (5 µm, 250 × 4.60 mm) column with ultraviolet detection at 210 nm. The calibration curves were linear ranging from 0.5 to 25 µg/mL. Precision of the assay was <13% (RSD) and was within 12% for all points in the calibration curve. The limit of quantitation for this method was 0.5 µg/mL. The assay was applied successfully to a pre-clinical study of lacosamide pharmacokinetics in dogs.

A Mixed-Methods Exploration of the Effect of Disabling Backward Navigation on Calculations-Focused Computerized Examinations.

Objective. The aim of this mixed-methods study was to examine the effect of disabled backward navigation on computerized calculation examinations in multiple courses.Methods. Student performance on comprehensive pharmacy calculation examinations before and after implementation of disabled backward navigation were compared. Deidentified data from ExamSoft were used to determine median examination scores, passing rates, and time to completion for all three attempts given on comprehensive calculation exams held in a pharmacy calculations course (PharDSci 504) and in three applied patient care laboratory courses (Pharm 531, 541, and 551). An anonymous, voluntary student survey gathered student perceptions of disabled backward navigation. Qualitative data were evaluated for thematic findings.Results. The impact of disabled backward navigation on test scores and passing rates varied by course and test attempt. Students in Pharm 541 and 551 performed significantly worse on the initial test attempt after backward navigation was disabled compared to the previous year, with no significant differences in student performance seen on the retakes. Performance in PharDSci 504 and Pharm 531 followed the opposite pattern, with no significant difference in performance for the initial tests but significantly increased performance on the retakes. The amount of time spent on examinations either significantly decreased or remained the same. Student perceptions were generally consistent across all cohorts, with at least 74% agreeing that disabling backward navigation increased examination difficulty.Conclusion. Disabling backward navigation had a mixed effect on student examination performance. This may highlight how student behaviors change as backward navigation is disabled.

Stereospecific pharmacokinetics of racemic homoeriodictyol, isosakuranetin, and taxifolin in rats and their disposition in fruit.

The chirality of flavonoids has been overlooked in the majority of pharmacokinetic studies of homoeriodictyol, isosakuranetin, and taxifolin. The stereospecific pharmacokinetic disposition of these xenobiotics in male Sprague-Dawley rats is described for the first time. Validated HPLC methods were used to analyze serum and urine samples of rats following intravenous administration of each flavonoid via jugular vein cannulation and to determine their content in selected fruits. The characterization and interpretation of the pharmacokinetic disposition profiles of homoeriodictyol, isosakuranetin, and taxifolin are described. A discrepancy exists between half-lives in serum and urine which may be attributed to low assay sensitivity in serum for the three compounds; thus, a more accurate estimation of the pharmacokinetic parameters was obtained from urine. The pharmacokinetics of homoeriodictyol, isosakuranetin, and taxifolin revealed distribution, metabolism, and elimination that were dependent on the stereochemistry of the stereoisomers. The (-)-(S)-enantiomers of homoeriodictyol and isosakuranetin and the (+)-(2S; 3R)-stereoisomer of taxifolin were predominant in lemon, grapefruit, and tomato. These findings were achieved using chiral methods of analysis; the utility and necessity of developing chiral methods of analysis for chiral xenobiotics are discussed.

Interprofessional education on opioid use and pain identifies team-based learning needs.

BACKGROUND: An interprofessional education (IPE) activity was designed for health professional students in pharmacy, medicine, nursing, social work, and addiction studies. The goals were to practice team-based collaboration for patients who are prescribed opioids for chronic pain and to evaluate student responses to the activity. INTERPROFESSIONAL EDUCATION ACTIVITY: Student teams were guided through an unfolding patient case that included evaluating the patient's history, screening tool results, morphine equivalent dose, prescription monitoring program report, and videos of a patient-provider interaction. The two-hour, in-person IPE activity culminated in creation of a patient-centered treatment plan. Surveys were administered to compare pre- and post-course opioid knowledge and post-course IPE attitudes among the healthcare professions. DISCUSSION: Pharmacy students' baseline opioid knowledge scores were similar to nursing students, significantly lower than medical students, and significantly higher than social work students. Pharmacy students reported significantly higher gains in opioid knowledge than medical students. Nursing and social work students showed significantly higher levels of agreement that the course enhanced attitudes toward interprofessional collaboration compared to medicine and pharmacy students. Students most frequently noted working with other professions as the most valuable aspect of the IPE activity. IMPLICATIONS: Training gaps can be met using novel IPE activities specific to chronic pain and opioid use. Depending on profession, students demonstrated varied baseline knowledge regarding opioid use for chronic pain. Comparing knowledge gains and attitudes on IPE collaboration among professions can detect areas for program refinement to address each professions' unique needs.

High-performance liquid chromatographic analysis: applications to nutraceutical content and urinary disposition of oxyresveratrol in rats.

A high-performance liquid chromatographic (HPLC) method was developed for the analysis of the stilbene, oxyresveratrol. This method involves the use of a Luna C(18) column with ultraviolet detection at 320 nm. The mobile phase consisted of acetonitrile, water and formic acid (30 : 70 : 0.04 v/v) with a flow rate of 0.6 mL/min. The calibration curves were linear over the range of 0.5-100.0 microg/mL. The mean extraction efficiency was between 98.9 and 109%. The precision of the assay was 0.069-18.4% (RSD%), and within 20% at the limit of quantitation (0.5 microg/mL). The bias of the assay was <15% and within 15% at the limit of quantitation. This assay was successfully applied to pre-clinical pharmacokinetic samples from rat urine and to nutraceutical product analysis.

Development and validation of a rapid and sensitive HPLC method for the quantification of 5-fluorocytosine and its metabolites.

To study the intracellular metabolism of the prodrug 5-fluorocytosine (5FC), we developed a novel reverse-phase high-performance liquid chromatography method to simultaneously detect 5FC and its four major anabolic metabolites: 5-fluorouracil, 5-fluorouridine, 5-fluorouridine-monophosphate and 5-fluoro-2'deoxyuridine-5'-monophosphate. Separation of each compound was accomplished under isocratic conditions using a C(18) column and mobile phase of formic acid-water (1 : 99 v/v). The method was validated for both accuracy and reproducibility in cell culture media. Additionally, metabolites were assessed for stability at ambient temperatures and following freeze-thaw cycles. Calibration curves were linear over a range of 1-200 microg/mL. Limit of quantification for four of the five compounds was 1 microg/mL in cell culture media (RSD < 11%). This method was successfully used to monitor intracellular conversion of 5FC to its metabolic products over a 24h period.

The Effect of Pharmacy Students' Attendance on Examination Performance in Two Sequential Active-Learning Pharmacotherapy Courses.

Objective. To examine the relationship between class attendance by Doctor of Pharmacy students and their performance on pharmacotherapy examinations within an active-learning classroom model. Methods. Second-year pharmacy students enrolled in a pharmacotherapy course series were included in the study (N=160). Class attendance was taken manually by members of the study team over a one-year study period (fall 2017 and spring 2018 semesters). Course attendance was not required and had no direct impact on student grades. Scores from the six competency-based examinations and overall course grades for each semester course, respectively, were then linked to class attendance records. Two additional examination attempts (retake and extended learning experience) were administered to students who did not receive a score of at least 80% on the initial exam or retake exam, respectively. Results. Class attendance was documented during 48 class sessions. Of the six examinations given each semester, students required an average of 1 retake of the examination during the fall semester and 1.5 retakes in the spring semester. A significant negative correlation was found in both courses between students missing more classes and receiving a lower final course grade. For each missed class session, there was a reduction in overall course grade of 0.18% and 0.14% in the fall and spring courses, respectively. Conclusion. Regular class attendance by pharmacy students enrolled in an active-learning pharmacotherapy curriculum was associated with higher scores on examinations. The results of this study illustrate the importance of attending active-learning sessions to attain higher examination scores. Further research is needed to determine whether class attendance is associated with students' improved ability to apply pharmacotherapy concepts.

Study Behaviors Associated with Student Pharmacists' Academic Success in an Active Classroom Pharmacy Curriculum.

Objective. To identify the specific study behaviors that promoted student pharmacists' success in an active-learning pharmacy curriculum. Methods. The Washington State University College of Pharmacy and Pharmaceutical Sciences implemented an active-learning, flipped classroom model for instruction to equitably deliver course content to Doctor of Pharmacy students on both its main and extended campuses. Students' ability to adapt to the new model and its impact on their study behaviors were unknown. A qualitative descriptive design that included semi-structured interviews was applied to evaluate the study behaviors of high-performing students. The study sample included 13 third and fourth professional year pharmacy students in the top 20% of their respective classes. Results. Interview responses were unaffected by baseline demographics such as gender and year of graduation. Content analysis generated five primary themes related to the behavioral strategies used by high performers: preparing for class, preparing for testing, seeking help, knowing yourself, and building on strengths. These were mapped to the four tenants of Wenger's social learning theory in the representation of findings: learning as doing, learning as belonging, learning as becoming, and learning as experience. Conclusion. High-performing students demonstrated a refined ability to select and modify study behaviors that aided in their academic success, demonstrating a high degree of metacognition. The results of this research may assist pharmacy faculty members in identifying critical elements for success of students enrolled in pharmacy programs using an active learning model.

A student initiative to implement peer-led study groups for a pharmacogenomics course: Evaluation of student performance and perceptions.

INTRODUCTION: To better elucidate the impact of cooperative learning outside the classroom, a student-initiated research project was conducted to explore the effects of participating in peer-led study groups (PLSGs) on student examination scores and perceptions. METHODS: First-year pharmacy students were given the opportunity to participate in weekly PLSGs for a pharmacogenomics course during spring 2016 and spring 2017. Student exam performance was stratified by those who attended vs. those who did not. Optional pre- and post-course surveys examined student perceptions of PLSGs. RESULTS: No significant differences were seen between the attendance groups in spring 2016. In spring 2017, student attendees were significantly more likely to pass two of their six exams (p = .04, p = .0029) and to have higher exam scores on one exam (p = .02) in comparison to non-attendees. Overall exam score averages were significantly different between attendees and non-attendees during spring 2017 (p = .03) but not during spring 2016 (p = .38). Perception surveys indicated students believed participation helped them to demonstrate competency and build confidence. Additionally, students reported they felt more comfortable clarifying questions during the study groups vs. during class time. CONCLUSIONS: The impact of study group participation on student exam performance was minimal over the two years of data collection, but there were instances where exam scores were positively impacted. Students perceived value in study group participation even if it did not translate directly to improved exam performance on all exams.

Stereospecific high-performance liquid chromatography of taxifolin, applications in pharmacokinetics, and determination in tu fu ling (Rhizoma smilacis glabrae) and apple (Malus x domestica).

A stereospecific method of analysis of racemic taxifolin (+/-3,5,7,3',4'-pentahydroxyflavanone) in biological fluids is necessary to study pharmacokinetics and disposition in fruit and herbs. A simple high-performance liquid chromatographic method was developed for the determination of all four taxifolin enantiomers. Separation was achieved on a Chiralcel(R) OJ-RH column with UV detection at 288 nm. The standard curves in serum were linear over a range of 0.5-100.0 microg/mL for each enantiomer. The mean extraction efficiency was >88.0%. Precision of the assay was <15% (CV), and was within 12% at the limit of quantitation (0.5 microg/mL). The bias of the assay was <15%, and was within 6% at the limit of quantitation. The assay was successfully applied to stereospecific disposition of taxifolin enantiomers in rats and to the quantification of taxifolin enantiomers in tu fu ling (Rhizoma smilacis glabrae) and apple (Malus x domestica).

Clinical toxicities of nanocarrier systems.

Toxicity of nanocarrier systems involves physiological, physicochemical, and molecular considerations. Nanoparticle exposures through the skin, the respiratory tract, the gastrointestinal tract and the lymphatics have been described. Nanocarrier systems may induce cytotoxicity and/or genotoxicity, whereas their antigenicity is still not well understood. Nanocarrier may alter the physicochemical properties of xenobiotics resulting in pharmaceutical changes in stability, solubility, and pharmacokinetic disposition. In particular, nanocarriers may reduce toxicity of hydrophobic cancer drugs that are solubilized. Nano regulation is still undergoing major changes to encompass environmental, health, and safety issues. The rapid commercialization of nanotechnology requires thoughtful environmental, health and safety research, meaningful, and an open discussion of broader societal impacts, and urgent toxicological oversight action.

Differential effects of resveratrol on androgen-responsive LNCaP human prostate cancer cells in vitro and in vivo.

Resveratrol is a phytochemical that has been under consideration for use as a prostate cancer chemopreventive agent. However, the efficacy, as well as the mechanisms of action of resveratrol on prostate cancer prevention, remains largely unknown. This study seeks to address these questions and examine the cancer preventive effects of resveratrol using complementary human LNCaP prostate cancer cell culture and xenograft models. In cultured LNCaP cells, we found that resveratrol inhibited cell growth. The growth inhibitory effects of resveratrol appeared to be through modulation of both androgen- and estrogen-mediated events. Global gene expression analysis using microarrays identified androgen-responsive genes as a group of genes universally affected by resveratrol in LNCaP cells in vitro. The effect of resveratrol on expression of these genes appeared to be through inhibition of both androgen- and estrogen-mediated transcription. In a xenograft model, resveratrol delayed LNCaP tumor growth and inhibited expression of a marker for steroid hormone responses. However, exposure to resveratrol also led to increased angiogenesis and inhibition of apoptosis in the xenograft. In summary, resveratrol may act through modulation of steroid hormone-dependent pathways to inhibit prostate cancer cell growth in both culture and xenografts, but exposure in vivo may be of concern.

Clinical pharmacokinetic and pharmacodynamic profile of etoricoxib.

The NSAID etoricoxib is a selective inhibitor of cyclo-oxygenase 2 (COX-2), approved for treatment of patients with chronic arthropathies and musculoskeletal and dental pain. The rate of absorption of etoricoxib is moderate when given orally (the maximum plasma drug concentration occurs after approximately 1 hour), and the extent of absorption is similar with oral and intravenous doses. Etoricoxib is extensively protein bound, primarily to plasma albumin, and has an apparent volume of distribution of 120 L in humans. The area under the plasma concentration-time curve (AUC) of etoricoxib increases in proportion to increasing oral doses between 5 and 120 mg. The elimination half-life of approximately 20 hours in healthy subjects enables once-daily dosing. Etoricoxib is eliminated following biotransformation to carboxylic acid and glucuronide metabolites that are excreted in urine and faeces, with little of the drug (<1%) being eliminated unchanged in the urine. Etoricoxib is metabolized primarily by the cytochrome P450 (CYP) 3A4 isoenzyme. Plasma concentrations (AUC) of etoricoxib appear not to be different in patients with chronic renal insufficiency compared with individuals who have normal renal function. Compared with healthy subjects, it has been reported that the AUC is increased by approximately 40% in patients with moderate hepatic impairment. No inhibitory effects on CYP2C9, 2C19, 2D6, 2E1 or 3A4 are expected to occur with etoricoxib. Coadministration of etoricoxib with other drugs has been examined only to a limited extent, thus further assessment is necessary. Etoricoxib has been assessed for the management of several specific disease states, including pain, osteoarthritis, and rheumatoid arthritis, and has shown similar efficacy in comparison with traditional NSAIDs (including naproxen, diclofenac and ibuprofen) in these conditions. Etoricoxib has demonstrated a significant reduction in gastrointestinal toxicity compared with many traditional NSAIDs. The renal adverse effects of etoricoxib appear to be similar to those of other NSAIDs, and the cardiovascular adverse effects of this selective COX-2 inhibitor require further clinical scrutiny. Further study is necessary to delineate the relevance of the pharmacokinetic disposition in terms of the clinical benefits and risks of etoricoxib compared with other options in the clinical arsenal.

Stereospecific high-performance liquid chromatographic assay of isosakuranetin in rat urine.

A stereospecific method of analysis of racemic isosakuranetin (5,7-dihydroxy-4'-methoxyflavanone) in biological fluids is necessary to study pharmacokinetics. A simple high-performance liquid chromatographic method was developed for the determination of isosakuranetin enantiomers. Separation was achieved on a Chiralpak AD-RH column with ultraviolet (UV)-detection at 286 nm. The standard curves in urine were linear ranging from 0.5 to 100.0 microg/ml for each enantiomer. The mean extraction efficiency was >88.0%. Precision of the assay was <15% (CV) and was within 12% at the limit of quantitation (0.5 microg/ml). Bias of the assay was <15% and was within 6% at the limit of quantitation. The assay was applied successfully to stereospecific disposition of isosakuranetin enantiomers in rat urine.

Stereospecific analysis of sakuranetin by high-performance liquid chromatography: pharmacokinetic and botanical applications.

A stereospecific method for analysis of sakuranetin was developed. Separation was accomplished using a Chiralpak AD-RH column with UV (ultraviolet) detection at 288 nm. The stereospecific linear calibration curves ranged from 0.5 to 100 microg/mL. The mean extraction efficiency was >98%. Precision of the assay was <12% (relative standard deviation (R.S.D.)%), and within 10% at the limit of quantitation (0.5 microg/mL). Bias of the assay was lower than 10%, and within 5% at the limit of quantitation. The assay was applied successfully to pharmacokinetic quantification in rats, and the stereospecific quantification in oranges, grapefruit juice, and matico (Piper aduncum L.).