A critical assessment of bioactive compounds databases.

Compound databases (DBs) are essential tools for drug discovery. The number of DBs in public domain is increasing, so it is important to analyze these DBs. In this article, the main characteristics of 64 DBs will be presented. The methodological strategy used was a literature search. To analyze the characteristics obtained in the review, the DBs were categorized into two subsections: Open Access and Commercial DBs. Open access includes generalist DBs (containing compounds of diverse origins), DBs with specific applicability, DBs exclusive to natural products and those containing compounds with specific pharmacological action. The literature review showed that there are challenges to making these repositories available, such as standardizing information curation practices and funding to maintain and sustain them.

[Box: see text].

Chemical characterization, antinociceptive and anti-inflammatory effect of Lippia lacunosa, a species used by the Bandeirantes.

ETHNOPHARMACOLOGICAL RELEVANCE: Lippia lacunosa Mart. & Schauer is an endemic plant from the Serra do Espinhaço mountain range located on the Atlantic plateau, Brazil. It is known as "chá de pedestre" and "rosmaninho" in folk medicine. This species has a characteristic mango aroma and is widely used by the population for flu, colds, sinus infections, coughing, relaxing baths, and foot baths after long walks. It is often confused with and, therefore, used interchangeably with L. rotundifolia and L. pseudothea. AIM OF THE STUDY: This study aimed to increase scientific knowledge on the ethnopharmacological use of Lippia lacunosa through the evaluation of the micromolecular composition and anti-inflammatory and antinociceptive activities of the hexane and ethanolic extracts, essential oil, and fractions in mice. MATERIALS AND METHODS: The chemical profile of L. lacunosa extracts and fractions were obtained by chromatographic methods such as Ultra-Performance Liquid Chromatography (UPLC), Gas Chromatography (GC), Column Chromatography (CC), and Thin Layer Chromatography (TLC). Carrageenan-induced paw edema was used to investigate the anti-inflammatory activity in mice. Mechanical allodynia induced by carrageenan and hot plate tests were employed to evaluate the antinociceptive activity. RESULTS: The main constituents found in the essential oil were the monoterpenes myrcene (13.81%), linalool (6.84%), ipsenone (21.2%), and myrcenone (25.44%); and sesquiterpenes elemol (7.30%) and spathulenol (3.15%). The chromatograph fractionation of essential oil yielded a fraction rich in the main compounds (F33), ipsenone and mircenone. In experimental models of paw edema and mechanical allodynia induced by carrageenan (600 µg, 30 µL, i.pl.), the administration of hexane extract, essential oil (50 or 100 mg/kg, p.o.) or majority fraction (10 mg/kg, p.o.) reduced paw edema. The ethanolic extract (100 mg/kg) reduced mechanical allodynia only in the 2 nd h of evaluation. On the other hand, the hexane extract (50 or 100 mg/kg) and essential oil (100 mg/kg), as well as the majority fraction (10 mg/kg), reduced mechanical allodynia throughout the evaluation period. The hexane extract, essential oil, and majority fraction F33 also reduced the heat-induced nociceptive response. Also, majority fraction F33 did not affect the time mice spent in the rota-rod apparatus. CONCLUSIONS: The elucidation of the composition of the essential oil and the demonstration of the activity of L. lacunosa in experimental models of acute inflammation and also in models of nociceptive and inflammatory pain can help to increase knowledge on the ancient ethnopharmacological use by the Bandeirantes, aiming at the evaluation of the species as a candidate for herbal medicine or phytopharmaceutical in the treatment of patients with inflammatory and painful conditions.

Campomanesia lineatifolia Ruiz & Pavón (Myrtaceae): Isolation of major and minor compounds of phenolic-rich extract by high-speed countercurrent chromatography and anti-inflammatory evaluation.

ETHNOPHARMACOLOGICAL RELEVANCE: Campomanesia lineatifolia Ruiz & Pavón (Myrtaceae), an edible species found in Brazilian Forest, possesses leaves that are traditionally used for the treatment of gastrointestinal disorders in Brazil. Extracts of C. lineatifolia are rich in phenolics and exhibit antioxidant, and gastric antiulcer properties. Furthermore, Campomanesia spp. have been described to possess anti-inflammatory properties, but studies related to chemical constituents of C. lineatifolia are scarce in the literature. AIM OF THE STUDY: This work aims to identify the chemical composition of the phenolic-rich ethanol extract (PEE) from C. lineatifolia leaves and evaluate the anti-inflammatory activity that could be related to its ethnopharmacological use. MATERIALS AND METHODS: The high-speed countercurrent chromatography (HSCCC), using an isocratic and a step gradient elution method, and NMR, HPLC-ESI-QTOF-MS/MS were used to isolate and identify the chemicals of PEE, respectively. Lipopolysaccharide-(LPS)-stimulated THP-1 cells were used to evaluate the anti-inflammatory activities from PEE and the two majority flavonoids isolated by measure TNF-α and NF-κB inhibition assays. RESULTS: Fourteen compounds were isolated from the PEE, further identified by NMR and HPLC-ESI-QTOF-MS/MS, twelve of them are new compounds, and two others are already known for the species. The PEE, quercitrin and myricitrin promoted a concentration-dependent inhibition of TNF-α, and PEE promoted an inhibition of NF-κB pathway. CONCLUSIONS: PEE from C. lineatifolia leaves demonstrated significant anti-inflammatory activity that may be related to the traditional use to treat gastrointestinal disorders.

Antithrombotic potential of Lippia alba: A mechanistic approach.

ETHNOPHARMACOLOGICAL RELEVANCE: Lippia alba (Mill.) N.E.Br. ex Britton & P. Wilson is traditionally used in Brazil as an adjunct in the relief of mild anxiety, as an antispasmodic, and as an antidyspeptic. This medicinal species was included in the Phytotherapeutic Form of the Brazilian Pharmacopeia 2nd edition (2021) and has already been described as the most used medicinal plant in a study with patients from an Anticoagulation Clinic in Brazil. Meanwhile, no studies were found that support the safety of the use of L. alba in patients using anticoagulants, a drug with several safety limitations. AIM OF THE STUDY: Provide scientific evidence to ensure the safety of the concomitant use of L. alba and warfarin and support the management of these patients by evaluating its in vitro anticoagulant effect and chemical composition. And, as a timely complementation, evaluate the potential of this medicinal species in the development of new antithrombotics. METHODS: The chemical profile of L. alba derivatives was analyzed by chromatographic methods such as Ultra-Performance Liquid Chromatography (UPLC) coupled with electrospray ionization mass spectrometry (ESI-MS), qualitative UPLC using Diode-Array Detection, and Thin Layer Chromatography. The anticoagulant activity was evaluated by the innovative Thrombin Generation Assay by Calibrated Automated Thrombogram method and using traditional coagulometric tests: prothrombin time, activated partial thromboplastin time, and plasma fibrinogen measurement. RESULTS: Extracts and fractions prolonged the coagulation time in all the tests and reduced thrombin formation in thrombin generation assay. Coagulation times with the addition of ethanloic extract (2.26 mg/mL) was 17.78s, 46.43s and 14.25s respectively in prothrombin time, activated partial thromboplastin time and fibrinogren plasma measurement. In thrombin generation test, this same extract showed ETP as 323 nM/min compared to control (815 nM/min) with high tissue factor and 582 nM/min compared to control (1147 nM/min) using low tissue factor. Presence of flavonoids, phenylpropanoids, and triterpenes were confirmed by chromatographic methods and 13 compounds were identified by UPLC-ESI-MS. Based on these results and on the scientific literature, it is possible to propose that phenylpropanoids and flavonoids are related to the anticoagulant activity observed. CONCLUSION: The results demonstrate the in vitro anticoagulant activity of L. alba, probably due to the activation of intrinsic and extrinsic pathways. It is concluded, then, that there is a potential for interaction, which needs to be further studied, between L. alba and warfarin. Also, this medicinal species shows a great potential for use in the development of new antithrombotics.

Anticoagulant and antiplatelet activity of aqueous extracts of Citrus sinensis and Lippia alba : interactions and potential for the development of antithrombotics.

Citrus sinensis and Lippia alba are herbal medicines widely used in the form of tea (infusion, decoction), which ethanolic extracts have already shown great anticoagulant activity in vitro . For this reason, they seem to be excellent candidates for the development of new antithrombotics and also have the potential to interact with them. The aim of this study was to evaluate the activity of aqueous extracts in blood coagulation and platelet aggregation, in addition to analysing the micromolecular composition of these species. Thrombin generation test (TGT) by the Calibrated Automated Thrombogram method and Platelet Aggregation Test by turbidimetry were performed to evaluate the biological activities, while the chemical composition was qualitatively evaluated using high-performance liquid chromatography. Aqueous extracts were elaborated according to the folk use. All extracts were effective in reducing thrombin formation in TGT. Infusion of L. alba and infusion and decoction of C. sinensis at a concentration of 0.6 mg/ml significantly reduced platelet aggregation induced by ADP, and only the decoction of L. alba at the same concentration was able to significantly reduce collagen-induced platelet aggregation. The presence of phenylpropanoids and flavonoids in C. sinensis and L. alba extracts was verified. Furthermore, hesperidin was identified in C. sinensis through coinjection. C. sinensis and L. alba are rich in phenolics and demonstrated an in-vitro effect on important processes of haemostasis (blood coagulation, platelet agreggation), corroborating the potential of C. sinensis and L. alba for the development of antithrombotics and interact with them.

Changes in antiparasitical activity of gold nanorods according to the chosen synthesis.

Gold nanorods (GNRs) are increasingly being studied for diagnostic and therapeutic purposes. Green synthesis based methods with natural compounds as additives stand out as a hope in terms of better synthesis methodology, with advantages of producing potentially less toxic and, perhaps, biologically active GNRs due to influence of natural additives used during synthesis. Exploring green chemistry using different natural phenolic compounds, the present work reveals different in vitro activity of GNRs evaluated against different parasites that causes skin infectious diseases compared to GNRs produced by convencional seed mediated method. This approach brings advantages in producing active GNRs, with ease calling, less cytotoxic and with a better selectivity index (SI) than GNRs synthesized by conventional seed mediated synthesis, opening new possibilities for therapies. Natural compounds used in green syntheses were gallic acid (GA), resveratrol (RSV) and a purified fraction of the hydroalcoholic extract of Stryphnodendron obovatum. GNRs exhibited great activity against Leishmania braziliensis, and the dermatophytes Tricophyton rubrum, T. interdigitale and Microsporum gypseum. The anti-Leishmania and antidermatophytic activity of GNRs reinforce the applicability of GNRs in biomedical field and the influence of synthesis method in biological activity, showing benefits related to the seedless synthesis with natural compounds. In addition, these preliminary results indicate the possibility of exploring at maximum the physical and chemical properties of GNRs in addition to the biological activity itself, such as the development of topical antiparasitic formulations for association with phototherapy.

Chemical Composition and In Vitro Anti-Helicobacter pylori Activity of Campomanesia lineatifolia Ruiz & Pavón (Myrtaceae) Essential Oil.

Helicobacter pylori is the most common cause of gastritis and peptic ulcers, and the number of resistant strains to multiple conventional antimicrobial agents has been increasing in different parts of the world. Several studies have shown that some essential oils (EO) have bioactive compounds, which can be attributed to antimicrobial activity. Therefore, EOs have been proposed as a natural alternative to antibiotics, or for use in combination with conventional treatment for H. pylori infection. Campomanesia lineatifolia is an edible species found in the Brazilian forests, and their leaves are traditionally used for the treatment of gastrointestinal disorders. Anti-inflammatory, gastroprotective, and antioxidant properties are attributed to C. lineatifolia leaf extracts; however, studies related to the chemical constituents of the essential oil and anti-H. pylori activity is not described. This work aims to identify the chemical composition of the EO from C. lineatifolia leaves and evaluate the anti-H. pylori activity. The EO was obtained by hydrodistillation from C. lineatifolia leaves and characterized by gas chromatography-mass spectrometry analyses. To assess the in vitro anti-H. pylori activity of the C. lineatifolia leaf's EO (6 µL/mL-25 µL/mL), we performed broth microdilution assays by using type cultures (ATCC 49503, NCTC 11638, both clarithromycin-sensitive) and clinical isolate strains (SSR359, clarithromycin-sensitive, and SSR366, clarithromycin-resistant). A total of eight new compounds were identified from the EO (3-hexen-1-ol (46.15%), α-cadinol (20.35%), 1,1-diethoxyethane (13.08%), 2,3-dicyano-7,7-dimethyl-5,6-benzonorbornadiene (10.78%), aromadendrene 2 (3.0%), [3-S-(3α, 3aα, 6α, 8aα)]-4,5,6,7,8,8a-hexahydro-3,7,7-trimethyl-8-methylene-3H-3a,6-methanoazulene (2.99%), α-bisabolol (0.94%), and ß-curcumene (0.8%)), corresponding to 98.09% of the total oil composition. The EO inhibited the growth of all H. pylori strains tested (MIC 6 µL/mL). To our knowledge, the current study investigates the relation between the chemical composition and the anti-H. pylori activity of the C. lineatifolia EO for the first time. Our findings show the potential use of the C. lineatifolia leaf EO against sensitive and resistant clarithromycin H. pylori strains and suggest that this antimicrobial activity could be related to its ethnopharmacological use.

Costus spiralis extract restores kidney function in cisplatin-induced nephrotoxicity model: Ethnopharmacological use, chemical and toxicological investigation.

ETHNOPHARMACOLOGICAL RELEVANCE: Costus spiralis (Jacq.). Roscoe (Costaceae) is traditionally used in Brazil for the treatment of kidney diseases such as pyelonephritis, urethra inflammation, kidney stones, and inflammatory conditions. There are reports of its use by Brazilian Indians since the 17th century when it was known as "pacocatinga." Currently, the use of the Costus species in Brazil is widespread, which was evidenced by the inclusion of the genus in the Brazilian National List of Medicinal Plants of Interest to the Unified Health System (RENISUS). AIM OF THE STUDY: This study aimed to confirm the ethnopharmacological use of Costus spiralis in the treatment of kidney diseases, toxicity study using animal models, and the phytochemistry of the species. MATERIALS AND METHODS: The chemical profile of Costus spiralis leaves extract (CSLE) was obtained for the hydroethanolic extract by ultra-performance liquid chromatography coupled to a mass spectrometer and ultraviolet detector with diode array (UPLC-UV/DAD-ESI-MS). The acute oral toxicity of the extract was predicted using the neutral red uptake cytotoxicity assay. Wistar rats were used in a model in vivo for confirmation of acute oral toxicity (2000 mg/kg p.o. for 14 days.) and determination of the effect on a cisplatin-induced nephrotoxicity model. RESULTS: The analysis by UPLC-UV/DAD-ESI-MS showed that the chemical composition of the extract is mostly di-glycosylated flavones of apigenin. In the extract were identified the flavones vicenin II and schaftoside. The quantification of total flavonoids by spectrometry showed 0.880%. CSLE proved to be safe for acute oral administration (2000 mg/kg) with an IC50 value of 222.9 µg/mL and predicted oral toxic dose of 523.82 µg/mL in a neutral red uptake cytotoxicity assay. The absence of death allows the classification of the extract in class 5 according to OECD 423 guidelines and therefore it can be considered as a high acute safety product, which is highly relevant, considering the wide popular use of the species. In the cisplatin-induced nephrotoxicity model, C. spiralis extract (5, 15, and 30 mg/kg) significantly improved renal function, reversing almost completely the effects on plasma creatinine levels and creatinine clearance (p < 0.001). CONCLUSIONS: This study demonstrates that oral administration of Costus spiralis extract leaves is safe and effective in restoring the renal function in rats in a cisplatin-induced nephrotoxicity. It is suggested that the observed activity is related to the flavonoids present. This hypothesis should be confirmed, and the participation of other secondary metabolites should be investigated in the future.

Antifungal effect of hydroethanolic extract of Fridericia chica (Bonpl.) L. G. Lohmann leaves and its therapeutic use in a vulvovaginal candidosis model.

Vulvovaginal candidosis (VVC) is one of the most frequent causes of gynecological consultations. Therefore, the development of new antifungal therapies against VVC is relevant. In this context, the leaves of Fridericia chica (Bonpl.) L. G. Lohmann are considered a therapeutic alternative since they are traditionally used in VVC therapy. However, no scientific evidence has supported this use against fungal vaginal infections. Then, we aimed to characterize the antifungal effect of a hydroethanolic extract of F. chica leaves (HEFc) and evaluate the therapeutic potential of this extract in a VVC model. HEFc inhibited the growth of C. albicans (256-1,204 µg/mL) and C. krusei (512 µg/mL) in vitro. HEFc inhibited yeast-to-hypha transition in C. albicans and has a low ability to induce resistance in this species. Intravaginal use of the HEFc at 50 mg/mL causes mycological cure in animals with VVC after 6 days of treatment, similar to the effect observed for the commercial antifungal nystatin. These results support the traditional use of F. chica leaves as a topical therapeutic option to treat VVC.

The rationality involved in the popular use of coffee (Coffea sp) and sunflower seed (Helianthus annuus) for the treatment of vitiligo: a case report.

INTRODUCTION: Vitiligo is a dermatological disease that affects about 0.38% to 2.9% of the world population. Currently, the main treatments used for vitiligo involve the use of topical drugs such as corticosteroids and calcineurin inhibitors, phototherapy, systemic treatment with steroids and even surgical grafts and, in acute cases, depigmenting treatments. Natural products are an alternative for the treatment of vitiligo: mamacadela (Brosimum gaudichaudii), a plant rich in furanocoumarins, and sowthistle (Sonchus oleraceus), rich in phenolic substances, are already used to treat vitiligo. There are also popular reports of the use of a preparation containing coffee (Coffea sp) and sunflower seed (Helianthus annuus) to treat vitiligo. CASE REPORT: A female patient, 28 years old, diagnosed with vitiligo, reported having obtained a positive result in the repigmentation of the pale white patches after the daily use of a preparation containing coffee and sunflower seed for about one year. DISCUSSION: Data from the scientific literature demonstrated that chemical constituents of these plants, such as chlorogenic acid and its isomers, which have antioxidant and anti-inflammatory action, and substances such as linoleic acid and vitamins E and B, which help in the process of melanin formation on the skin, may be responsible for the observed repigmentation of the patches. Further research on this case report is important for scientific validation and the development of new therapeutic options, especially with less adverse effects, in the treatment of vitiligo.

Mechanisms and interactions in concomitant use of herbs and warfarin therapy: An updated review.

This review is an updated and expanded version published in this journal in 2016. Warfarin pharmacotherapy is extremely complex, since in addition to being a low therapeutic index drug, it does not follow the dose-response pattern and has characteristics that predispose the occurrence of interactions, such as high binding rate to plasma proteins, metabolization by cytochrome P450 enzymes, further to acting in the complex process of blood coagulation, platelet activation, and inflammation. For these reasons, warfarin has great potential for interaction with drugs, foods, and herbal medicines. Herb-warfarin interactions, however, are still not very well studied; thus, the objective of this update is to present new information on the subject aiming to provide a scientific basis to help health professionals in the clinical management of these interactions. A literature review was performed from May to June 2021 in multiple databases and articles published in 2016 to 2021 were included. A total of 59 articles describing 114 herbal medicines were reported to interact with warfarin. Of the plants mentioned, 84% had the potential to increase warfarin effect and the risk of bleeding. Targets possibly involved in these interactions include the processes of blood coagulation, platelet activation, and inflammation, in addition to the pharmacokinetics and pharmacodynamics of warfarin. Despite these alarming numbers, however, the clinical management of interactions is known to be effective. Thus, it is important that the use of these herbal medicines be done with caution in anticoagulated patients and that studies of herb-drug interactions be encouraged in order to generate information to support the clinical management of patients.

In vitro anticoagulant activity of selected medicinal plants: potential interactions with warfarin and development of new anticoagulants.

OBJECTIVES: Warfarin is the most widely used anticoagulant in the world, but it has several limitations including its narrow therapeutic range, need for dose adjustment and high potential for interactions. The simultaneous use of other drugs or even medicinal plants and certain foods could interfere with its therapeutic activity. In this context, this study aims to investigate the in vitro anticoagulant potential and phytochemical constitution of 17 plants selected from a previous clinical cross-sectional study (2014), that investigated the habits of plant utilization among patients taking warfarin. METHODS: Ethanol extracts and essential oils were evaluated, in vitro, as to their effect in the prothrombin time (PT) and activated partial thromboplastin time (aPTT) tests. Four species that presented aPTT >50 s were selected for phytochemical evaluation. RESULTS: Thirteen of the 17 plants selected demonstrated a significant anticoagulant effect in at least one of the evaluated parameters. Citrus sinensis (PT=14.75 and aPTT=53.15), Mentha crispa (aPTT=51.25), Mikania laevigata (PT=14.90 and aPTT=52.10), and Nasturtium officinale (aPTT=50.55) showed greater anticoagulant potential compared to normal plasma pool (PT=12.25 and aPTT=37.73). Chemical profiles of these four species were obtained, and certain compounds were identified: rosmarinic acid from M. crispa and isoorientin from N. officinale. CONCLUSIONS: Thus, the results of this study could be a useful indicator for clinical practice towards the possibility of interaction between these plants and anticoagulants, although further clinical research is needed taking into consideration the limitations of in vitro studies. These findings also suggest that further research into the action of these plants could be of real clinical value in identifying potential alternative anticoagulant therapies.

TNF-α inhibition, antioxidant effects and chemical analysis of extracts and fraction from Brazilian guaraná seed powder.

Paullinia cupana Kunth., commonly named Guaraná, is a plant from Brazil used as stimulant. The aim of this study was to evaluate the potential of extracts and tannins-rich and methylxanthines-free fraction from guaraná in the anti-inflammatory and antioxidant effect in vitro. Extract 1 obtained good yields of tannins and methylxanthines and was used to identify a type-A procyanidin trimer by LC-ESI-MS. Fraction 4 was rich in tannins and absent of methylxanthines. The extracts and fraction exhibited strong capacity for scavenging DPPH radical with IC50 between 5.88 and 42.75-µg/mL and inhibited TNF-α release by LPS-activated THP-1 cells when compared with control cells and did not present toxicity to THP-1 cells. The fraction 4, rich in tannins, was highly active, with IC50 5.88 µg/mL by DPPH method and inhibited TNF-α release in 83.50% at 90 µg/mL. These results reinforced potential anti-inflammatory of guaraná and data for new therapeutic approaches.

New antiglaucomatous agent for the treatment of open angle glaucoma: Polymeric inserts for drug release and in vitro and in vivo study.

A benzamidine derivative from diminazene was tested for a novel activity: treatment of primary open-angle glaucoma. This drug was incorporated into mucoadhesive polymeric inserts prepared using chitosan (Chs) and chondroitin sulfate (CS). Of current interest is the mucoadhesion, which increases the contact time with the ocular surface, resulting in improved bioavailability; also, the inserts are made to act as a prolonged release system. In the present work the inserts were prepared by the solvent casting method using different polymeric proportions (30:70, 50:50, 75:25% w/w Chs:CS and 100% Chs). Thermal analysis and infrared spectroscopy both demonstrated physical dispersion of the active drug. The most promising was the 50:50% Chs:CS which demonstrated that it was not fragile and has an in vitro release profile of up to 180 minutes. In addition, it presented greater adhesion strength in relation to the other formulations. These physicochemical results corroborate the in vivo tests performed. In this sense, we also demonstrated that the treatment with the 50:50% insert can control the intraocular pressure (IOP) for at least 3 weeks and prevents damage to the retinal ganglion cells (RGCs) compared to the placebo insert. Thus, this indicates thus that the new drug is quite viable and promising in glaucoma treatment.

Use of chitosan as pharmaceutical excipient in ocular drug delivery systems: Sterilization and pharmacokinetics.

The use of chitosan as a pharmaceutical excipient in the ocular field is already established. Nevertheless, some aspects related to its ocular administration, such as sterilization and excipient's pharmacokinetics, remain unclear. So, in this study, we evaluated those two relevant aspects, related to chitosan administration in eye. We used chitosan-based ocular inserts (CI) as formulation model. CI were produced by solvent/casting method and sterilized by saturated steam. Sterilization was confirmed by direct inoculation of inserts in suitable microbiological growth media. Physicochemical characterization of inserts before and after sterilization was performed. Results suggested that, although steam sterilization changed the arrangement of the matrix, the heat and the humidity did not modify the structure of the main polymeric chain. Pharmacokinetics of CI radiolabeled with technetium-99m (99m Tc) was assessed by scintigraphic images and ex vivo biodistribution study, after ocular administration in male Wistar rats. Scintigraphic and images analysis and ex vivo biodistribution study showed that the insert remained mainly in the eye until 6 hr after administration and its degradation products began to migrate to the abdominal cavity after 18 hr. Together, these data represent an important step forward the manufacturing and the clinical application of CI in the ophthalmic field.

Development and validation of analytical method by HPLC-DAD for determination of vasodilator active in pharmaceutical ophthalmic forms

The substance 4-Aminobenzamidine dihydrochloride (4-AD) is one of the degradation products of diminazene aceturate and has demonstrated antiglaucomatous potential. Glaucoma is the second leading cause of blindness worldwide; thus, new therapeutic alternatives must be studied, for example, the molecule 4-AD vehiculated into polymeric inserts for prolonged release. The present work aims to develop and validate an analytical method to quantify 4-AD in pharmaceutical ophthalmic forms. A HPLC was used with UV-Vis detector, at 290 ƞm and ACE® C18 column (125 × 4.6 mm, 5 µm), in which the mobile phase consists of phosphate buffer (pH 7.4) and triethylamine (30 mmol/L), under an isocratic flow of 1.0 mL/min. The retention time of 3.2 minutes was observed. The method was developed and validated in accordance with ANVISA recommendations and ICH guides. The linearity range was established between the concentrations 5 and 25 µg/mL (correlation coefficient r = 0.993). The accuracy, repeatability, and intermediate precision tests obtained a relative standard deviation less than or equal to 5%. In addition, the method was considered selective, exact. and robust, with pH being its critical factor. Therefore, the HPLC analysis method is robust and can be used to quantify 4-AD in pharmaceutical forms for ocular application.(AU)

Thrombin generation test with the calibrated automated thrombogram and anticoagulant activity of Mentha crispa.

: Evaluate the in-vitro effect of Mentha crispa extract on blood coagulation, compare the conventional coagulometric tests with thrombin generation test (TGT), and study the qualitative micromolecular composition of M. crispa. Extract of M. crispa was incubated with plasma and used in the coagulometric tests: prothrombin and activated partial thromboplastin times, fibrinogen, and TGT. A phytochemical prospection was performed to evaluate the chemical composition of this extract. The extract was efficient in prolonging prothrombin time and activated partial thromboplastin time, and reducing fibrinogen levels and TGT parameters, indicating that the extract of M. crispa inhibited the intrinsic and extrinsic pathways of blood coagulation. The results obtained in TGT are in agreement with the results of conventional coagulometric tests and the in-vitro anticoagulant activity of M. crispa suggests that its use by patients using oral anticoagulants deserves caution.

In Vitro Anticoagulant Activity of Mikania laevigata: Deepening the Study of the Possible Interaction Between Guaco and Anticoagulants.

Mikania laevigata, popularly known in Brazil as guaco, is widely used for respiratory disorders. As this plant is rich in coumarins, there is evidence of indications that it may cause bleeding and therefore should not be used concomitantly with anticoagulants. The basis of this information is very theoretical, with no clinical evidence of such contraindication. Thus, the aim of this study was to evaluate the in vitro effect of M. laevigata extract on blood coagulation through prothrombin time (PT) and activated partial thromboplastin time (aPTT) tests, fibrinogen plasma concentration, and the new thrombin generation test, which investigate, with high sensibility, hemostatic changes (CAAE 60904316.6.0000.5149), besides evaluating its qualitative micromolecular composition, providing scientific evidence to support the management of patients taking warfarin. Ethanolic extracts of guaco leaves were incubated with a plasma pool of healthy individuals at concentrations of 1.67, 2.26, and 2.86 mg/mL. The presence of flavonoids, tannins, coumarins, and triterpenes was demonstrated by selective reagents in thin layer chromatography. Benzoylgrandifloric acid, cinnamoylgrandifloric acid, o-coumaric acid, coumarin, and quercetin-3-ß-glucoside were identified by coinjection in ultraperformance liquid chromatography. The extract at all concentrations prolonged TP and aPPT and reduced the potential for endogenous thrombin potential by the thrombin generation test. The control plasma had endogenous thrombin potential = 1465 nM/min, and after the addition of M. laevigata extract (2.26 mg/mL), this value was reduced to 1087 nM/min, indicating a lower generation of thrombin. Related to fibrinogen plasma concentration, concentrations of 2.26 and 2.86 mg/mL were effective in reducing plasma fibrinogen levels. These results allow us to conclude that the guaco extract demonstrated an anticoagulant effect in vitro, possibly interfering with intrinsic, extrinsic, and common coagulation pathways. A discussion on the contribution of the identified substances to the activity is also present.

Chitosan/hydroxyethyl cellulose inserts for sustained-release of dorzolamide for glaucoma treatment: In vitro and in vivo evaluation.

Eye drops containing hydrophilic drugs are commonly used to reduce intraocular pressure (IOP) in glaucoma patients, but compliance to the treatement is commonly reduced by frequent dosing and eventual systemic side effects. Sustained-release drug delivery systems, such as ocular inserts, can reduce dosing, limit systemic exposure, reduce side effects, and, then, improve patient adherence to therapy. Here, we developed and evaluated chitosan/hydroxyethyl cellulose-based ocular inserts for sustained release of dorzolamide, a hydrophilic drug. Dorzolamide inserts (DI) were produced by solvent/casting method and characterized by various physicochemical techniques. Pharmacokinetics studies were performed using scintigraphic images and ex vivo biodistribution. The effectiveness of inserts was tested in glaucomatous rats. Characterization studies showed that the drug strongly interacted with the polymeric matrix, but in vitro results showed that DI took only 3 h to release 75% of dorzolamide entraped. However, scintigraphic images and ex vivo biodistribution studies revealed that more than 50% of 99mTc-dorzolamide remained in the eye after 18 h of DI administration, while only about 30% of the drug remained in the eye after drops instilation. DI exerted significant hypotensive effect for two weeks, after single administration, while IOP values remained high in placebo and untreated groups. Eye drops were effective only during the treatment period. Only DI treatment prevented retinal ganglion cells death. Altogether, these findings evidenced the potential application of polymeric-based inserts for sustained release of dorzolamide in glaucoma management.

Maltodextrin and Gum Arabic-Based Microencapsulation Methods for Anthocyanin Preservation in Juçara Palm (Euterpe edulis Martius) Fruit Pulp.

The juçara fruits (Euterpe edulis Martius), native to the Atlantic Forest, are rich in anthocyanins. To preserve the anthocyanins in juçara fruit pulp, this study aimed to evaluate the effectiveness of microencapsulation by spray drying and freeze drying with maltodextrin (dextrose equivalent 16.5 to 19.5) and gum arabic in different proportions. The obtained microparticles were characterized by quantifying the total polyphenol and anthocyanin contents, by performing differential scanning calorimetry, thermogravimetry, and infrared spectroscopy and by using scanning electron microscopy to analyze the morphology of the particles. The total amount of polyphenols in the fruit pulp was 750 ± 16.7 mg GAE/100 g of the freeze-dried sample. The total anthocyanins in the fruit pulp was 181.25 ± 5.36 (mg/100 g). The microparticles were formed by employing maltodextrin and gum arabic in a 1:1 proportion as the polymeric matrix; the mixtures of pulp and polymeric matrix were prepared in proportions of 2:3 and 2:1, preserving up to 83.69% of the anthocyanin content. Lyophilization of the 2:1 mixture resulted in an anthocyanin content of 116.89 ± 4.43 (mg/100 g), whereas lyophilization of the 2:3 mixture resulted in 151.68 ± 1.39 (mg/100 g) anthocyanin content, which did not differ from the value obtained by spray drying the 2:3 mixture (150.76 ± 5.79 (mg/100 g)). Thermal analyses showed that the microparticles obtained by freeze drying at a ratio of 2:3 presented greater resistance to degradation with increasing temperature. The incorporation of the pulp in the polymeric matrix was demonstrated by IR analyses. Microparticles obtained by freeze drying showed the formation of various-sized flakes, whereas those obtained by spray drying were spherical in shape. Microencapsulation is a possible alternative for improving the stability of the anthocyanins in this fruit.