Polypharmacy and Drug Interactions in the COVID-19 Pandemic.

The COVID-19 pandemic generated a great impact on health systems. We compared evolution, polypharmacy, and potential drug-drug interactions (P-DDIs) in COVID-19 and non-COVID-19 hospitalizations during first wave of pandemic. Prescriptions for hospitalized patients ≥ 18 years (COVID-19 and non-COVID-19 rooms) between April and September 2020 were included. The computerized medical decision support system SIMDA and the physician order entry system Hdc.DrApp.la were used. Patients in COVID-19 rooms were divided into detectable and non-detectable, according to real-time reverse transcription polymerase chain reaction (RT-PCR). Number of drugs, prescribed on day 1, after day 1, and total; polypharmacy, excessive polypharmacy, and P-DDIs were compared. 1,623 admissions were evaluated: 881 COVID-19, 538 detectable and 343 non-detectable, and 742 non-COVID-19. Mortality was 15% in COVID-19 and 13% in non-COVID-19 (RR [non-COVID-19 vs. COVID-19]: 0.84 [95% CI] [0.66-1.07]). In COVID-19, mortality was 19% in detectable and 9% in non-detectable (RR: 2.07 [1.42-3.00]). Average number of drugs was 4.54/patient (SD ± 3.06) in COVID-19 and 5.92/patient (±3.24) in non-COVID-19 (p<0.001) on day 1 and 5.57/patient (±3.93) in COVID-19 and 9.17/patient (±5.27) in non-COVID-19 (p<0.001) throughout the hospitalization. 45% received polypharmacy in COVID-19 and 62% in non-COVID-19 (RR: 1.38 [1.25-1.51]) and excessive polypharmacy 7% in COVID-19 and 14% in non-COVID-19 (RR: 2.09 [1.54-2.83]). The frequency of total P-DDIs was 0.31/patient (±0.67) in COVID-19 and 0.40/patient (±0.94) in non-COVID-19 (p=0.022). Hospitalizations in the COVID-19 setting are associated with less use of drugs, less polypharmacy and less P-DDIs. Detectable patients had higher mortality.

Development and Implementation of the Hdc.DrApp.la and SIMDA Programs to Reduce Polypharmacy and Drug-drug Interactions in Patients Hospitalized in Internal Medicine.

OBJECTIVES: We evaluated polypharmacy and possible drug-drug interactions (p-DDIs) in hospitalized patients before and after using the SIMDA Computerized Medical Decision Support System (CMDSS). MATERIALS AND METHODS: We included the prescriptions of ≥ 18 years hospitalized patients in the internal medicine department. We developed and implemented the Hdc.DrApp Physician Order Entry System and the CMDSS SIMDA, which detects p-DDIs and signals dosage adjustment based on renal function. To evaluate the impact of the CMDSS, we made a comparison Before (Survey) / After (Intervention): Survey between Oct/22/2019, and Mar/21/2020, and Intervention between Apr/4/2020 and Sep/3/2020. We analyze prescriptions from the first day and after the first day. We compared the number of drugs, polypharmacy (≥ 5 drugs), excessive polypharmacy (≥ 10 drugs), and p-DDIs. We evaluated differences with the X2 test, Yates correction, Fisher's exact test, ANOVA, and post hoc tests according to their characteristics. RESULTS: We evaluated 2,834 admissions: Survey 1,211 and Intervention 1,623. The number of drugs per patient was 6.02 (± 3.20) in Survey and 5.17 (± 3.22) in Intervention (p < 0.001) on the first day and 9.68 (± 5.60) in Survey and 7.22 (± 4.93) in Intervention (p < 0.001) throughout the hospitalization. Polypharmacy was present in 64% of the Survey and 53% of Interventions (RR: 0.83 (0.78-0.88); and excessive polypharmacy in 14% of the Survey and 10% of Intervention (RR: 0.73, 0.60-0.90). The frequency of total p-DDIs was 1.91/patient (± 4.11) in Survey and 0.35 (± 0.81) in the Intervention (p < 0.001). CONCLUSIONS: We developed and implemented the Hdc.DrApp and SIMDA systems that were easy to use and allowed us to quantify and reduce polypharmacy and p-DDIs.

Pharmacokinetic and pharmacodynamic study of three products of epoetin alfa as single subcutaneous dose in healthy volunteers.

Hemax® is an epoetin alfa product developed by Biosidus S.A. in Argentina at the end of the 1980s and has been present in that market since 1991. The initial presentation was a lyophilized powder containing albumin as stabilizer, to best adapt to environmental conditions in developing countries; more recently, a prefilled syringe, albumin-free presentation was developed, since this presentation has become the preferred standard in many markets. The primary objective was to compare the pharmacokinetic profile of different formulations of epoetin alfa after a single subcutaneous administration to healthy volunteers of 40 000 IU of Eprex/Erypo® and Hemax® PFS. This clinical trial was conceived following an open-label, randomized, three-way three-period cross-over balanced, and sequential design. The study was conducted on 24 healthy volunteers. To analyze similarity between Hemax® PFS and the innovator product, Eprex®, area under the curve (AUC) and Cmax of both products have been compared. The 90% CI lower limit for the geometric mean ratios was higher than 80% for any comparisons, and the 90% CI upper limit for these geometric ratios was below 125% for all the comparisons made, thus demonstrating equivalence between both products. The comparison between Hemax® PFS and Eprex® resulted in similar 90% CI for Cmax , AUC(0-120 h) and AUC(0-inf) ratios, all of them within the 80-125% interval, with a power above 95% for each ratio. These findings suggest biosimilar patterns for absorption velocity (with Tmax close to 15 h), absorption extent, and elimination (with an elimination half-life close to 25-30 h for each formulation).

Dextrophropoxyphene effects on QTc-interval prolongation: Frequency and characteristics in relation to plasma levels.

OBJECTIVE: To evaluate frequency and risk factors for dextropropoxypheneinduced QT-interval prolongation in the clinical setting. DESIGN: Prospective, noninterventional, observational, longitudinal cohort approach. Electrocardiograms were blindly evaluated by independent professionals. SETTING: General ward of a public hospital of metropolitan Buenos Aires. PATIENTS, PARTICIPANTS: Ninety-two patients with indication of receiving dextropropoxyphene for analgesic purposes were included consecutively. All patients finished the study. INTERVENTIONS: All patients were monitored with electrocardiographic controls (previous to drug administration and during steady state) to diagnose and quantify changes in the duration of the QTc interval. MAIN OUTCOME MEASURE: Frequency of drug-induced QTc interval prolongation, QTc interval correlation with plasma drug, and metabolite levels. RESULTS: Ninety-two patients were studied (50 percent males). All patients received a (mean ± SD [range]) dextropropoxyphene dose of 125 ± 25[100-150] mg/d. Dextropropoxyphene and norpropoxyphene concentrations were 112 ± 38[45-199] and 65 ± 33[13-129] ng/mL, respectively. The intra-treatment QTc interval was >450 ms in only one patient (only with the Hodge correction). There were no cases of QTc > 500 ms, and there were no significant differences in the results considering different correction formulas (Bazzet, Fridericia, Framingham, Hodges). Dextropropoxyphene concentrations correlated with QTc (R > 0.45) interval and ΔQTc (R 0.52-0.87), whereas norpropoxyphene correlation was even greater for QTc (R > 0.40-0.64) and ΔQTc (R > 0.47-0.92). Depending on the QTc correction formula, eight patients presented ΔQTc > 30 ms and one patient with ΔQTc > 60 ms. No patient presented arrhythmia during the study. CONCLUSIONS: The authors did not observe a relationship between dextropropoxyphene and QTc interval prolongation at the therapeutic doses used in Argentina.

A Critical View of "On TB Vaccines, Patients' Demands, and Modern Printed Media in Times of Biomedical Uncertainties: Buenos Aires, 1920-1950".

The putative Pueyo's vaccine was a commercial venture that obtained marketing authorization in 1946, a turbulent period of Argentine history. After a few months, health authorities withdrew financial support from the state to buy the vaccine and required patients to sign a written consent to receive that product. An independent investigation did not find any evidence of benefit in non-clinical and clinical evaluation of the putative vaccine.

Interactions of a prenylated flavonoid from Dalea elegans with fluconazole against azole- resistant Candida albicans.

BACKGROUND: The prenylated flavonoid 2', 4'-dihydroxy-5'-(1'″, 1'″-dimethylallyl)-8-prenylpinocembrin (8PP, formerly 6PP) shows antifungal activity, inhibits rhodamine 6G efflux and reverses fluconazole (FCZ) resistance in azole-resistant Candida albicans overexpressing cdr1, cdr2 and mdr1 transporters. PURPOSE AND DESIGN: In this paper, we tried to characterize 8PP in vitro interactions on the cell growth and lethality of C. albicans. We also initiated preliminary in vivo toxicological studies on mice. METHODS: The effects of 8PP and FCZ on cell growth and viability of C. albicans were evaluated by CLSI guidelines. The checkerboard assay was used to search for interactions on cell growth. The time-kill assay was used to study fungicidal effects. Acute toxicity was evaluated at a single dose schedules. RESULTS: From the checkerboard design, and using a starting inoculum of 103CFU/ml, the fractional inhibitory concentration (FIC) of FCZ and 8PP could be determined as 0.11 and 0.50, respectively, with a FIC index value (FICI) of 0.61. This FICI and the isobologram showing a concave shape suggests an additive interaction between them. At a higher starting inoculum (105CFU/ml), C. albicans growth and viability were decreased by FCZ, 8PP and their combination in a concentration-dependent way. For FCZ, minimum fungicidal concentration (MFC) and FC50 (the concentration that kills 50% of the fungal cells) were 4-fold reduced (280-70µM) in combination with 125µM 8PP. A decrease of 3 log units in viable counts with respect to control was reached (3.65 ± 1.05 ‰, p< 0.0001). Thus, both fungistatic compounds when combined achieved an almost complete fungicidal effect at lower concentrations respecting of each of them alone. In preliminary toxicological assessment, lethal dose 50% (LD50) for 8PP by the i.p. route was 357 and 245mg/kg, for female and male adult albino mice, respectively. FCZ LD50 was 785 and 650mg/kg for female and male animals, respectively CONCLUSIONS: In vitro results suggest additive interactions between 8PP and FCZ with respect to C. albicans cell growth. Besides killing per se, 8PP helps FCZ to achieve an almost complete fungicidal effect, which would be crucial to eradicate fungal infections.

Simultaneous quantitation of meperidine, normeperidine, tramadol, propoxyphene and norpropoxyphene in human plasma using solid-phase extraction and gas chromatography/mass spectrometry: Method validation and application to cardiovascular safety of therapeutic doses.

RATIONALE: Several opioid analgesics have been related to the prolongation of cardiac repolarization, a condition which can be fatal. In order to establish a correct estimation of the risk/benefit balance of therapeutic doses of meperidine, normeperidine, tramadol, propoxyphene and norpropoxyphene, it was necessary to develop an analytical method to determinate plasma concentrations of these opioids. METHODS: Here we describe a method which incorporates strong alkaline treatment to obtain norpropoxyphene amide followed by a one-elution step solid-phase extraction, and without further derivatization. Separation and quantification were achieved by gas chromatography/electron ionization mass spectrometry (GC/EI-MS) in selected-ion monitoring mode. Quantification was performed with 500 µL of plasma by the addition of deuterated analogues as internal standards. RESULTS: The proposed method has been validated in the linearity range of 25-1000 ng/mL for all the analytes, with correlation coefficients higher than 0.990. The lower limit of quantification was 25 ng/mL. The intra- and inter-day precision, calculated in terms of relative standard deviation, were 2.0-12.0% and 6.0-15.0%, respectively. The accuracy, in terms of relative error, was within a ± 10% interval. The absolute recovery and extraction efficiency ranged from 81.0 to 111.0% and 81.0 to 105.0%, respectively. CONCLUSIONS: A GC/MS method for the rapid and simultaneous determination of meperidine, normeperidine, tramadol, propoxyphene and norpropoxyphene in human plasma was developed, optimized and validated. This procedure was shown to be sensitive and specific using small specimen amounts, suitable for application in routine analysis for forensic purposes and therapeutic monitoring. To our knowledge, this is the first full validation of the simultaneous determination of these opioids and their metabolites in plasma samples.

In vivo Phenotyping Methods: Cytochrome P450 Probes with Emphasis on the Cocktail Approach.

BACKGROUND: Differences in drug response among patients are common. Most major drugs are effective in only 25 to 60 percent of the patients, in part due to the CYP enzymes, whose activity vary up to 50-fold between individuals for some index metabolic reactions. Several factors affect CYP activity, among which genetic polymorphisms have been studied as the major cause for long time. Age, gender, disease states, and environmental influences such as smoking, concomitant drug treatment or exposure to environmental chemicals are also important. METHODS: This article reviews the available literature on multiple phenotypes assessment as an important tool to predict possible therapeutic failures or toxic reactions to conventional drug doses during patient evaluation. RESULTS: Probe drugs can be used in various combinations allowing for the in vivo assessment of multiple pathways of drug metabolism in a single experiment, configuring a new tool known as phenotyping "cocktails". There are several drug cocktails with different advantages and disadvantages. Most of them have sufficient clinical evidence and data validation to support their use in clinical setting as a surrogate for the risk of adverse reaction in the course of therapy, leading to a better balance between efficacy and safety. CONCLUSION: Probes characteristics and metabolic ratio measurements are important in the evaluation of phenotyping cocktails as near-future applications.

Meperidine-induced QTc-interval prolongation: prevalence, risk factors, and correlation to plasma drug and metabolite concentrations
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A prolongation of the QTc-interval has been described for several opioids, including pethidine (meperidine). OBJECTIVE: To evaluate in the clinical setting the frequency and risk factors associated with the QT-interval prolongation induced by meperidine. RESEARCH DESIGN AND METHODS: We recruited patients requiring meperidine administration and recorded their medical history and comorbidities predisposing to QT-interval prolongation. Ionograms and electrocardiograms (ECGs) were performed at baseline and during treatment; QT was corrected using the Bazzet, Fridericia, Framinghan, and Hogdes formulas. We measured meperidine and normeperidine by gas chromatography. Values are expressed as mean ± SD (range). RESULTS: 58 patients were studied (43.1% males). All patients received meperidine at a dose of 304 ± 133 (120 - 480) mg/day. Meperidine and normeperidine concentrations were 369 ± 60 (265 - 519) and 49 ± 17 (15 - 78) ng/mL, respectively. Intratreatment control found QTcB 370 ± 30 (305 - 433), QTcFri 353 ± 35 (281 - 429), QTcFra 360 ± 30 (299 - 429), QTcH 359 ± 27 (304 - 427), ΔQTcB +9 ± 42 (-90 to +136), ΔQTcFri +4 ± 45 (-86 to +137), ΔQTcFra +5 ± 40 (-77 to +129), and ΔQTcH +7 ± 40 (-76 to +129) ms. Meperidine concentration correlated with QTc-interval (R > 0.36) and ΔQTc (R > 0.69) but the correlation was even better for normeperidine concentration, QTc (R > 0.52) and ΔQTc (R > 0.81). Depending on the QTc correction formula used, 13 - 15 patients (22.41 - 25.86%) presented ΔQTc values > 30 ms, and 7 - 8 patients (12.07- 13.79%) showed ΔQTc values > 60 ms. Renal failure was associated with risk for ΔQTc > 30 ms of 3.74 (IC95% 1.73 - 8.10) and for ΔQTc > 60 ms of 4.27 (IC 95% 1.26 - 14.48). No patient developed arrhythmias during the study. CONCLUSIONS: Meperidine treatment causes ECG changes (QTc-interval prolongation) in high correlation with normeperidine plasma concentration. Renal failure increases the risk.
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Relative bioavailability between two teriparatide formulations in healthy volunteers.

OBJECTIVE: To compare the pharmacokinetics, relative bioavailability (RB), immunogenicity, and safety after a single dose of test or reference formulation of teriparatide in healthy human volunteers in order to demonstrate whether both products are similar. RESEARCH DESIGN AND METHODS: We compared pharmacokinetic parameters, immunogenicity, and safety after a single dose of two formulations (Osteofortil® and Forteo®) of teriparatide in a randomizedsequence, open-label, two-period crossover study in 24 healthy volunteers. The washout period between formulations was 7 days. Blood samples were collected at baseline and 0, 5, 10, 15, 20, 25, 30, 45, 60, 75, 90, 120, 150 minutes, and 3 and 4 hours after administration. Teriparatide concentrations were determined using ELISA. Adverse events were monitored. RESULTS: Geometric mean (90% CI) Cmax for test and reference formulations were 165.86 (153.35 - 212.13) and 175.37 (164.04 - 221.04) pg/mL, the AUC0-t was 14,932 (5,275 - 15,752) and 14,153 (1,861 - 16,875) pg×min/mL, and the AUC0-∞ was 16,147 (15,047 - 18,799) and 15,467 (14,473 - 18,126) pg×min/mL, respectively. The test/reference ratios (90% CI) for Cmax, AUC0-t, and AUC0-∞ were 94.58% (85.29 - 104.87), 105.5% (97.77 - 113.84), and 104.4% (96.97 - 112.39), respectively No subject reported adverse events. CONCLUSION: Test formulation met pharmacokinetic criteria for bioequivalence.

Tramadol Induced QTc-Interval Prolongation: Prevalence, Clinical Factors and Correlation to Plasma Concentrations.

UNLABELLED: In recent years, several cases of torsade de pointes have been associated with many opioids. However, to present no cases have been reported with tramadol. OBJECTIVE: To evaluate the effect of tramadol on QT-interval in the clinical setting. RESEARCH DESIGN AND METHODS: Medical history and comorbidities predisposing to QT interval prolongation were registered for patients requiring medical assistance that involved tramadol administration. Ionograms and ECGs were performed at baseline and intratreatment; QT interval was analyzed after correction with Bazzet, Fridericia, Framinghan and Hogdes formula. RESULTS: 115 patients were studied (50.4% males) All patients had received tramadol 150-400 mg/day during 3.0-5.0 days at the moment of intratreatment control. Plasma concentrations of tramadol were 201-1613 ng/mL. Intratreatment electrocardiographic control, as mean ± SD (range), showed QTcB 372±32 (305 to 433), QTcFri 356±37 (281 to 429), QTcFra 363±33 (299 to 429), QTcH 362±30 (304 to 427), ΔQTcB 26±40 (-73 to 110), ΔQTcFri 24±48 (-97 to 121), ΔQTcFra 22±42 (-81 to 109) and .QTcH 22±38 (-68 to 110) ms. QTc interval presents high correlation with plasma tramadol concentrations (for .QTc, R>0.77). Renal failure was associated with a relative risk for ΔQTc > 30 ms of 1.90 (IC95% 1.31-2.74) and for ΔQTc > 60 ms of 4.74 (IC95% 2.57-8.74). No patient had evidence of arrhythmia during the present study. CONCLUSION: Tramadol produces QTc interval prolongation in good correlation with plasma drug concentrations; renal failure is a risk factor for higher concentration and QT prolongation by tramadol.

Drug-Induced QTc Interval Prolongation: A Multicenter Study to Detect Drugs and Clinical Factors Involved in Every Day Practice.

OBJECTIVE: The actual prevalence of drug induced QTc prolongation in clinical practice is unknown. Our objective was to determine the occurrence and characteristics of drug-induced QT prolongation in several common clinical practices. Additionally, a subgroup of patients treated with dextropropoxyphene of particular interest for the regulatory authority was analysed. RESEARCH DESIGN AND METHODS: Medical history and comorbidities predisposing to QT interval prolongation were registered for 1270 patient requiring medical assistance that involved drug administration. Three ionograms and ECGs were performed: baseline, intra- and after treatment; QT interval was corrected with Bazzet formula. RESULTS: Among patients, 9.9% presented QTc >450/470 ms, 3% QTc > 500 ms, 12.7% ΔQTc >30 ms and 5.2% ΔQTc >60 ms. QTc prolongation associated with congestive heart failure, ischemic cardiopathy, diabetes, renal failure, arrhythmias, hypothyroidism, and bradycardia. At univariate analysis, clarithromycin, haloperidol, tramadol, amiodarone, glyceryl trinitrate, amoxicillin + clavulanic acid, amoxicillin + sulbactam, ampicillin + sulbactam, fentanyl, piperacillin + tazobactam, and diazepam prolonged QTc. Prolongation remained significantly associated with furosemide, clarithromycin, glyceryl trinitrate and betalactamase inhibitors after multivariate analysis. CONCLUSION: QT interval prolongation in everyday practice is frequent, in association to clinical factors and drugs that can be easily identified for monitoring and prevention strategies.

Age-distribution and genotype-phenotype correlation for N-acetyltransferase in Argentine children under isoniazid treatment.

INTRODUCTION: Metabolic clearance of isoniazid (INH) may be up to 10 times faster in individuals who are rapid acetylators compared with slow acetylators. In addition, the acetylation phenotype has been suggested to change with age. A better knowledge of the age distribution of the acetylation genotype and phenotype in children requiring INH for tuberculosis treatment or prevention could be important to optimize safety and efficacy of INH use. OBJECTIVES: The aim of the present study was to evaluate the genotype and phenotype of NAT2 in an Argentinean pediatric population rom Buenos Aires. In addition, we wanted to describe genotype-phenotype correlation, as well as its distribution at different ages. METHODOLOGY: NAT2 genotyping was performed by RFLP technique, searching for common polymorphisms. Acetylisoniazid and isoniazid concentrations were measured by HPLC and NAT2 phenotype was defined from the ratio of both concentrations (Metabolic Ratio, MR). RESULTS: Almost half of the patients (46.02%) possessed wild-type haplotype, with 17.05% of individuals having two fully functional alleles, 57.95% one fully functional allele and 25% with no fully functional allele. According to phenotype, most children (96.59%) were classified as fast acetylators, whereas 1.14% of the cases were intermediate and 2.27% slow acetylators. There was a positive association between age and MR (R = 0.52985, p < 0.000001) with a significant MR difference between age categories (p < 0.001). CONCLUSIONS: We found a high proportion of rapid acetylators compared with other populations. Acetylator phenotype showed a positive correlation with age, with a significant change around the 4th year of life.

A prenylated flavanone from Dalea elegans inhibits rhodamine 6 G efflux and reverses fluconazole-resistance in Candida albicans.

In previous studies, 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin, a prenylated flavonoid isolated from Dalea elegans roots, showed activity against multiresistant Staphylococcus aureus and Candida albicans, as well as an uncoupling effect on mitochondria and antioxidant activity. The aim of this study was to evaluate the inhibitory effects of 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin and fluconazole on the efflux of rhodamine 6 G in azole-resistant C. albicans 12-99 that expresses multidrug transporters Cdr1p, Cdr2p, and Mdr1p. The effect of fluconazole and 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin on rhodamine 6 G efflux was assessed in both azole-sensitive and azole-resistant C. albicans. Between 1 and 1000 µM, 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin inhibited rhodamine 6 G efflux only in azole-resistant C. albicans 12-99 in a concentration-dependent manner (IC50 = 119 µM); a competitive effect was observed. It also showed selectivity of action in comparison with other flavanones (6-prenylpinocembrin, isolated from aerial parts of D. elegans, pinocembrin, naringenin, and hesperetin, all at 250 µM). To check the possible implications of the inhibition of azole efflux on cell growth, antifungal assays were conducted. Minimal inhibitory concentration values were 150 µM for 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin and higher than 400 µM for fluconazole. The combination of both compounds at either inhibitory or subinhibitory concentrations was significantly more effective than each compound separately. Minimal inhibitory concentration for fluconazole decreased by more than 400 times in the presence of 100 µM 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin, reversing azole resistance and giving values similar to those of azole-sensitive C. albicans. These data are consistent with a dual action of 2',4'-dihydroxy-5'-(1''',1'''-dimethylallyl)-6-prenylpinocembrin: direct antifungal effect on azole-resistant C. albicans 12-99 and inhibition of azole transporters, which results in reversion of fluconazole resistance.

Effect of bitter compounds on amylase secretion in murine submandibular glands: Signaling pathway mechanisms.

BACKGROUND: Amylase is synthesized in submandibular glands (SMG) and released into the oral cavity to degrade carbohydrates in the mouth. Bitter taste receptors (T2R) belong to the G-protein coupled receptor (GPCR) family and are expressed in the taste cells and also in the digestive tract. METHODS: The activity of amylase secreted by murine SMG was measured, detecting maltose by Bernfeld's method. Amylase and T2R6 were detected by imunohistochemistry and Western blot. The expression of Ggustducin, Gi, and phospholipase Cß2 was also studied by Western blot. cAMP levels were measured by radioimmunoassay and inositol monophosphate production was quantified by ELISA. RESULTS: Theophylline, denatonium and cycloheximide exerted a dose-dependent inhibition on amylase secretion. This effect was reverted by preincubating SMG with an anti-Gαi antibody. cAMP production was increased by the same compounds, an effect that was also abrogated by an anti-Gαi antibody. Bitter compounds reduced inositol monophosphate formation in SMG and H-89, a protein kinase A inhibitor, reverted this action, revealing that this protein kinase down regulates phospholipase C activity. GENERAL SIGNIFICANCE: We demonstrated that theophylline, denatonium and cycloheximide inhibit salivary amylase secretion, activating an intracellular signaling pathway that involves cAMP and phospholipase C, that cross talks via protein kinase A.