Phytochemical analysis using GC-FID, FPLC fingerprinting, antioxidant, antimicrobial, anti- inflammatory activities analysis of traditionally used Eucalyptus globulus essential oil

Eucalyptus globules is an widely distributed in tropical and subtropical regions. It has been widely used as folk medicine, and folk cosmetic owing to its antioxidant values. Despite its importance, phytochemical and pharmacological studies remain infancy. This study was aimed at extraction of essential oil by steam-distillation and evaluation of bioactive components, antioxidant, antimicrobial, anti-inflammatory activities along with analysis by UV-VIS, FT-IR and Fluorescent techniques. Fast protein liquid chromatography (FPLC) was used to confirm the presence of polyphenols. Different antioxidant activities like DPPH., ABTS.+, .OH, superoxide, nitric oxide and reducing power of the essential oil. Essential oil was analyzed by UV-VIS, FT-IR and Fluorescent techniques. In vitro antimicrobial activity was also monitored. FT-IR fingerprint qualitative analysis was performed using commercial standards. Considerable amount of flavonoids were detected in essential oil. Oil exhibited considerable scavenging activities of ABTS.+, .OH, superoxide, nitric oxide and reducing power. UV-VIS, FT-IR analysis revealed the presence of polyphenolics in essential oil. Fluorescent spectroscopy revealed the presence of fluorophores in essential oil. FPLC and FT-IR fingerprint analysis revealed the presence of bioactive constituents like rutin, tannic acid, vanillic acid and ascorbic acid in the essential oil. A strong anti-inflammatory activity of oil was observed using fluorescent spectroscopy. An appreciable in vitro antibacterial activity against gram-negative bacteria like Acetobacter aceti and Pseudomonas aeruginosa was detected. The data provides the scientific support to the use of essential oil from Eucalyptus globules as a potent herbal source of bioactive compounds possessing natural antioxidant activities in food and pharmaceutical industries.

A novel sensitive colorimetric determination of catecholamine drug in dosage form via oxidative coupling reaction with MBTH and potassium ferricyanide

A new and direct colorimetric method has been established for the determination of catecholamine (methyldopa, MD) in both pure form and in pharmaceutical formulations. The method is based on the oxidative coupling reaction of MD with 3-methyl-2-benzothiazolinone hydrazone hydrochloride monohydrate (MBTH) and potassium ferricyanide at pH 10.4 in aqueous medium to form an orange product that has a maximum absorption at 460 nm. Beer's law plot showed good correlation in the concentration range of 1.0−56.0 µg mL-1, with detection limit of 0.31 µg mL-1. Molar absorptivity for the above method was found to be 6.56×103 L mol-1 cm-1. All the measurements were carried out at 25 ± 1.0 °C, the formation constant (logKf) value of colored species is 9.48 and the standard free energy (DG‡) is − 54.09 KJ mol-1. This method was applied successfully to determination of MD in tablets and the results were compared with the USP method. Common excipients used as additives in tablets do not interfere in the proposed method. The method is accurate, precise and highly reproducible, while being simple, cheap and less time consuming and hence can be suitably applied for routine analysis of MD in bulk and dosage forms.

A new and ecological miniaturized method by spectrophotometry for quantification of vancomycin in dosage form

Vancomycin, an important antibiotic, is marketed as lyophilized powder. In the context of routine analysis of this product, the existence of a more advantageous and effective method is interesting. Thus, the objective of this work is to develop and validate a new analytical method, faster, low-cost, ecological and miniaturized for quantification of vancomycin in lyophilized powder using spectrophotometry in ultraviolet region. Buffer solution pH 6.8, quartz cuvette with capacity of 700 µL and 280 nm were chosen. The method proved to be linear in the range of 50-150 µg/mL (0.9997). The selectivity of the method was proven in two ways: The standard-sample overlay aimed to identify vancomycin in the sample; The forced degradation test (sample solutions prepared in 0.01 M HCl, 0.01 M NaOH and aqueous conditions and kept at 60 ºC by 8 hours, and UV 254 nm at ambient temperature during 24 hours) aimed to show the susceptibility of the method to consequently indicate the stability of the sample. It was precise in intraday (RSD 1.27%), interday (RSD 1.18%) and between analysts (RSD 1.92%) levels. It was robust when small variations were performed in seven important parameters (wavelength, cuvette, filtration step, dibasic and monobasic phosphate brand, ultrasound time and source of water). The accuracy was proved by the standard recovery test and showed mean recovery of 101.10%. This method can be applied in routine analysis of quality control of vancomycin lyophilized powder and it is an effective, accessible and ecological alternative, which follows the Green Analytical Chemistry principles, presenting less waste generation, no use of toxic solvents, smaller sample volumes and required diluents, which impacts on the final cost of the analyzes.

Chlorine determination in medicinal plants by potentiometry with ion-selective electrode after microwave-induced combustion

A method based on microwave-induced combustion (MIC) was applied for medicinal plants digestion allowing further chlorine determination by potentiometry using ion-selective electrode (ISE). Sample masses ranging from 500 to 1000 mg were evaluated for MIC digestion. Water and 10, 25, 50, and 100 mmol/L NH4OH were investigated as absorbing solutions. The accuracy of the proposed method was evaluated by using certified reference materials (CRMs), by recovery tests (500 µg/g), and also by comparison with the results obtained by inductively coupled plasma optical emission spectrometry (ICP-OES) after microwave-assisted alkaline extraction (MAE). Using water or NH4OH solutions (10 to 100 mmol/L), recoveries close to 100% and relative standard deviation lower than 5% were obtained. Results were in agreement with CRMs values (better than 95%) and also with those values obtained by using the MAE method. The main advantage of the proposed method was the complete combustion of high sample mass (1000 mg) resulting in low quantification limit (12.5 µg/g) and chlorine determination at low concentration by ISE. Another advantage of the proposed method was the high chlorine stability in digests (up to 30 days of storage) even using water as absorbing solution, which is in agreement with green analytical chemistry recommendations. Finally, the proposed MIC method was applied for commercial medicinal plants and the chlorine concentration was in the range of 59.4 ± 1.4 to 2038 ± 70 µg/g. The proposed MIC method was considered suitable for quality control for chlorine determination in medicinal plants.

Use of Box–Behnken design for optimization of compounded medication: acyclovir capsules report

Campus compounding pharmacies play an important role in public health. Herpes simplex is one of the most common viral diseases in humans, which generates a great demand for acyclovir capsules in compounding pharmacy. It is well known that the formulation's components influence the effectiveness of the drug. The objective of this study is to show the applicability of Box-Behnken design in optimization of a compounded formulation and to evaluate the effect of excipients on dissolution and drug content in acyclovir 200 mg capsules produced at UFF´s University Pharmacy (FAU). The formulations were prepared and evaluated for average weight test, uniformity of dosage units and in vitro dissolution, while meeting pharmacopoeial specifications. A statistical analysis showed that sodium starch glycolate, Aerosil®, influences drug content and dissolution results. Magnesium stearate shows no influence on the dissolution at different concentrations but influences the assay results. A numerical optimization was applied to adjust the formulation variables based on the foresaid responses, accomplishing the best formulation that will be prepared and dispensed at FAU upon medical prescription.

Development of derivative spectrophotometric method for simultaneous determination of pyrazinamide and rifampicin in cubosome formulation

The ultraviolet spectrophotometry analysis for quantitative assay of drugs is a method accurate, sensitive, selective and reproductive with the advantage of being a simple and less expensive method. In this study, a derivative ultraviolet spectrophotometric method was developed for simultaneous determination of pyrazinamide (PYZ) and rifampicin (RIF). The spectrophotometric method was evaluated according to validation guidelines for specificity, linearity, limits of detection and quantification, precision, accuracy and robustness. The first-derivative spectra were obtained and by the zerocrossing point, the wavelength 247 nm and 365 nm were selected for PYZ and RIF quantification, respectively. No interference from cubosome excipients was detected in the proposed method. The results demonstrated linearity in a range of 4.0 – 12.0 µg/mL with an adequate correlation coefficient for both drugs. The intra and inter-day precision results (RSD < 5%) indicated the reproducibility of the method. The accuracy data showed satisfactory results (RSD < 5%) from recovery test. In addition, the robustness results showed that the PYZ and RIF content were unaffected by the solvent alteration of methanol to methanol:water (99:1, v/v). The derivative ultraviolet spectrophotometric method proved to be an excellent strategy for simultaneous determination of PYZ and RIF.

Post mortem changes: challenges in drug analysis in the diagnosis of deaths from intoxication

In forensic toxicology, alternative matrices and sampling sites are required for a correlation of antemortem and postmortem concentrations with the least possible error. Postmortem redistribution phenomena and biochemical changes inherent to these processes are possible, and represent interferences in these analyses. This study aimed to perform a bibliographic review through Pubmed database within a 10-year period of time, using the keywords: forensic analysis AND redistribution. We observed that for quantitative analyses the preferred matrix is blood from peripheral vessels, and when it is not available, vitreous humor is a great specimen for choice.

Ozonized oils: a review of its quality control, stability and effectiveness in the treatment of Acne vulgaris

Acne affects most young people and its topical treatment with antibacterials is associated with increased bacterial resistance to antibiotics and adverse effects. As an alternative, ozone therapy stands out through the application of ozonized oils. The objective of this work was to raise the scientific evidence about the effectiveness in the treatment of acne, in addition to the techniques of characterization and stability of ozonated oils. This is an exploratory, descriptive study with a quantitative approach, based on the analysis of scientific references in a bibliographic review of the expository type, of the last 20 years. Among the selected references, only four manuscripts reporting clinical studies of ozone therapy, with controversial results. Seven articles with the physicochemical characterization of ozonated oils were found. The major part of manuscripts reported the use of sunflower, sesame and olive oil. The more common techniques used to characterize the ozonation process are the peroxide value (PV) and the iodine index (Ii), which represents the proportion of unsaturated groups, whose values increase and decrease, respectively with ozonization progress. The viscosity of oils is increased by the formation of polymeric peroxides; the FTIR spectrum, which identifies the decrease in the stretch bands C = C, in addition to ozone formation, monitored by NMR, are also employed. Increased antimicrobial activity has been demonstrated with the ozone level of the oils, but the activity against Cutibacterium acne has not been reported. Only two article reported satisfactory stability for 6 months of refrigerated ozonized oil or kept at room temperature, showing the need for more specific research to support the application of ozonized oils in the treatment of acne and stability data of these products.

Analysis of seized stanozolol formulations in South Brazil by liquid chromatography coupled to quadrupole time-of-flight-mass spectrometry

Anabolic-androgenic steroids (AAS) are synthetic derivatives of testosterone which are used medically for several diseases. However, misuse is commonly observed by athletes to promote enhancement of strength and performance. AAS are frequently obtained through online black markets from clandestine drug manufacturing laboratories, without any quality standards, being potentially dangerous for users. The purpose of this work was the development and application of a fast and simple procedure for the quantitation of stanozolol by liquid chromatography coupled to quadrupole time-of-flight-mass spectrometry (LC-QTOF-MS) in tablets packs seized in Rio Grande do Sul state, Brazil. The samples of stanozolol were separated considering its dosage form. The internal standard (methyltestosterone) was added to the aliquots of the samples, dissolved in methanol and 5μL were injected into the analytical system. The newly developed method has been validated for lower limit of quantitation (LLOQ), linearity, accuracy, precision and selectivity. The LLOQ was 0.1 µg/mL. The developed method was successfully applied to 31 samples seized by the Secretaria da Receita Federal do Brasil (a Brazilian federal revenue service agency). According to the results, 90.3% of the suspected medicines (n=31) were adulterated, and 65% exhibited higher concentrations of stanozolol than those indicated in the label. This work successfully established a new method for quantification of stanozolol using LC-QTOF-MS. This method aims at contributing to the identification and quantification of this anabolic androgenic steroid frequently seized by federal inspection agencies.

Counterfeit medicines: a pilot study for chemical profiling employing a different proposal of an usual technique

Gas chromatography (GC) is a gold standard technique used in forensic laboratories, including for the characterization of counterfeit medicines. When coupled simultaneously to flame ionization (FID) and mass detector (MS) allow the identification and quantification of medicines and drugs employing a single method, besides permitting the application of chemometric tools for forensic intelligence purposes. Here is presented a pilot project that developed and applied a qualitative method for the analysis of counterfeit medicines comprised by amphetamine-type stimulants and antidepressants, through a simple extraction procedure followed by GC-FID/MS analysis, with application of exploratory tools by Hierarchical Cluster Analysis (HCA) and Principal Component Analysis (PCA). The main purpose was to identify similarities between the all compounds detected in the irregular medicines allowing the traceability of illicit producers with the creation of a common data base. Through the analyses it was verified that different producers of counterfeit medicines labeled as Sibutramine, added a mixture of Caffeine and Benzocaine in their formulation, respecting the same ratio of 2.2:1. HCA was able to confirm these results, showing the presence of both falsifications in the same cluster, representing the best tool to identify similar characteristics among the samples – when compared to PCA. Other interesting finding was the use of Fluoxetine as a falsification of counterfeit medicines labeled as Sibutramine and Diethylpropion. Another seized sample labeled as “Nobesio Forte”, marketed as a mix of stimulants, showed only Caffeine and Lidocaine in its formulation. The pilot project applied primarily to 45 samples of counterfeit medicines containing amphetamine-type stimulants and antidepressants, showed the capability of perform the chemical profiling of counterfeit medicines in the solid form - powder, capsules and tablets. Further analysis can be performed for different types of medicines in solid form using the developed method, allowing the construction of a single database to perform the chemical profiling of counterfeit medicines, enabling the traceability of illicit producers.