Overcoming Challenges in Pediatric Formulation with a Patient-Centric Design Approach: A Proof-of-Concept Study on the Design of an Oral Solution of a Bitter Drug.

Designing oral formulations for children is very challenging, especially considering their peculiarities and preferences. The choice of excipients, dosing volume and palatability are key issues of pediatric oral liquid medicines. The purpose of the present study is to develop an oral pediatric solution of a model bitter drug (ranitidine) following a patient centric design process which includes the definition of a target product profile (TPP). To conclude on the matching of the developed solution to TPP, its chemical and microbiological stability was analyzed over 30 days (stored at 4 °C and room temperature). Simulation of use was accomplished by removing a sample with a syringe every day. Taste masking was assessed by an electronic tongue. The developed formulation relied on a simple taste masking strategy consisting in a mixture of sweeteners (sodium saccharine and aspartame) and 0.1% sodium chloride, which allowed a higher bitterness masking effectiveness in comparison with simple syrup. The ranitidine solution was stable for 30 days stored at 4 °C. However, differences were noted between the stability protocols (unopened recipient and in-use stability) showing the contribution of the simulation of use to the formation of degradation products. Stock solution was subjected to acid and alkali hydrolysis, chemical oxidation, heat degradation and a photo degradation stability assessment. The developed pediatric solution matched the TPP in all dimensions, namely composition suitable for children, preparation and handling adapted to hospital pharmaceutical compounding and adequate stability and quality. According to the results, in-use stability protocols should be preferred in the stability evaluation of pediatric formulations.

Manual or automated measuring of antipsychotics' chemical oxygen demand.

Antipsychotic (AP) drugs are becoming accumulated in terrestrial and aqueous resources due to their actual consumption. Thus, the search of methods for assessing the contamination load of these drugs is mandatory. The COD is a key parameter used for monitoring water quality upon the assessment of the effect of polluting agents on the oxygen level. Thus, the present work aims to assess the chemical oxygen demand (COD) levels of several typical and atypical antipsychotic drugs in order to obtain structure-activity relationships. It was implemented the titrimetric method with potassium dichromate as oxidant and a digestion step of 2h, followed by the measurement of remained unreduced dichromate by titration. After that, an automated sequential injection analysis (SIA) method was, also, used aiming to overcome some drawbacks of the titrimetric method. The results obtained showed a relationship between the chemical structures of antipsychotic drugs and their COD values, where the presence of aromatic rings and oxidable groups give higher COD values. It was obtained a good compliance between the results of the reference batch procedure and the SIA system, and the APs were clustered in two groups, with the values ratio between the methodologies, of 2 or 4, in the case of lower or higher COD values, respectively. The SIA methodology is capable of operating as a screening method, in any stage of a synthetic process, being also more environmentally friendly, and cost-effective. Besides, the studies presented open promising perspectives for the improvement of the effectiveness of pharmaceutical removal from the waste effluents, by assessing COD values.

Evaluation of ionic liquids as alternative solvents for aldolase activity: Use of a new automated SIA methodology.

An automated methodology is proposed for the evaluation of a set of ionic liquids (ILs) as alternative reaction media for aldolase based synthetic processes. For that, the effect of traditionally used organic solvents and ILs on the activity of aldolase was studied by means of a novel automated methodology. The implemented methodology is based on the concept of sequential injection analysis (SIA) and relies on the aldolase based cleavage of d-fructose-1,6 diphosphate (DFDP), to produce dihydroxyacetone phosphate (DHAP) and d-glyceraldehyde-3-phosphate (G3P). In the presence of FeCl3, 3-methyl-2-benzothiazoline hydrazine (MBTH) forms a blue cation that can be measured at 670nm, by combination with G3P. The influence of several parameters such as substrate and enzyme concentration, temperature, delay time and MBTH and FeCl3 concentration were studied and the optimum reaction conditions were subsequently selected. The developed methodology showed good precision and a relative standard deviation (rsd) that does not exceed 7% also leading to low reagents consumption as well as effluent production. Resorting to this strategy, the activity of the enzyme was studied in strictly aqueous media and in the presence of dimethylformamide, methanol, bmpyr [Cl], hmim [Cl], bmim [BF4], emim [BF4], emim [Ac], bmim [Cl], emim [TfMs], emim [Ms] and Chol [Ac] up to 50%. The results show that the utilization of ILs as reaction media for aldolase based organic synthesis might present potential advantages over the tested conventional organic solvents. The least toxic IL found in this study was cho [Ac] that causes a reduction of enzyme activity of only 2.7% when used in a concentration of 50%. Generally, it can be concluded that ILs based on choline or short alkyl imidazolium moieties associated with biocompatible anions are the most promising ILs regarding the future inclusion of these solvents in synthetic protocols catalyzed by aldolase.

Improved activity of α-chymotrypsin in mixed micelles of cetyltrimethylammonium bromide (CTAB) and ionic liquids: a kinetic study resorting to sequential injection analysis.

In the present work an automatic methodology, based on sequential injection analysis, (SIA) for the kinetic evaluation of α-chymotrypsin (α-CT) activity in cetyltrimethylammonium bromide (CTAB)/ionic liquid (IL) mixed micelles, was developed. The α-CT-catalysed hydrolysis of N-succinyl-l-phenylalanine-p-nitroanilide (SPpNA) was monitored by following the change in absorbance at 410nm brought about by the formation of p-nitroaniline (pNA). The influence of parameters such as reagents concentration, flow rate, as well as reaction coil length, on the sensitivity and performance of the SIA system were studied and the optimum reaction conditions were subsequently selected. The effect of CTAB/IL micelles on the catalytic constant Kcat and apparent Michaelis-Menten constant (KM) for SPpNA hydrolysis was then studied. The kinetic assays evidence that CTAB/ILs mixed micellar systems can induce α-CT superactivity. In order to perform a critical evaluation of the obtained results, CMC and average micellar size of CTAB/hmim[Cl], CTAB/bmim[Cl], CTAB/bmpyr[Cl] and CTAB/bmpy[Cl] mixed micelles were evaluated by fluorescence and dynamic light scattering, respectively. The SIA methodology showed to be an interesting tool for evaluation of α-CT activity in mixed micelles as it proved to be robust and exhibited good repeatability in all the assay conditions leading also to a reduction of the consumption of solvents as well as of effluent production.

Automated carboxylesterase assay for the evaluation of ionic liquids' human toxicity.

Aiming the prediction of ionic liquids' (ILs) human toxicity, an automated carboxylesterase activity assay was developed. The method was implemented on a sequential injection analysis (SIA) system and relied on the hydrolysis of 4-methylumbelliferyl acetate by the enzyme, to produce the fluorescent compound 4-methylumbelliferone (λ(exc)=365 nm; λ(em)=460 nm). The adopted approach was based on activity/inhibition assays in which the reduction of enzyme activity was denoted by a decrease of fluorescence. The activity assays were performed in aqueous media and in the presence of increasing concentrations of seven commercially available ILs and sodium bis(p-nitrophenyl) phosphate, a strong inhibitor of carboxylesterase. The inhibitory effects were quantified by means of the calculation of half maximal inhibitory effective concentration (EC(50)) values that were between 38 and 134 µmol L(-1). This allowed us to perform some considerations about the toxicity of the compounds. The less and more toxic ILs found in this study were emim [Ms] and tbph [Ms], respectively. The developed SIA methodology is robust and exhibits good repeatability in all the tested conditions (rsd<5.0%, n=10). At the same time a reduction of costs was attained by reduced consumption of reagents and minimal effluent production (2 mL per cycle).