Slug Mucosal Irritation Assay Research Articles

Chitosan-Based Thermogelling System for Nose-to-Brain Donepezil Delivery: Optimising Formulation Properties and Nasal Deposition Profile.

Donepezil nasal delivery strategies are being continuously investigated for advancing therapy in Alzheimer's disease. The aim of this study was to develop a chitosan-based, donepezil-loaded thermogelling formulation tailored to meet all the requirements for efficient nose-to-brain delivery. A statistical design of the experiments was implemented for the optimisation of the formulation and/or administration parameters, with regard to formulation viscosity, gelling and spray properties, as well as its targeted nasal deposition within the 3D-printed nasal cavity model. The optimised formulation was further characterised in terms of stability, in vitro release, in vitro biocompatibility and permeability (using Calu-3 cells), ex vivo mucoadhesion (using porcine nasal mucosa), and in vivo irritability (using slug mucosal irritation assay). The applied research design resulted in the development of a sprayable donepezil delivery platform characterised by instant gelation at 34 °C and olfactory deposition reaching a remarkably high 71.8% of the applied dose. The optimised formulation showed prolonged drug release (t1/2 about 90 min), mucoadhesive behaviour, and reversible permeation enhancement, with a 20-fold increase in adhesion and a 1.5-fold increase in the apparent permeability coefficient in relation to the corresponding donepezil solution. The slug mucosal irritation assay demonstrated an acceptable irritability profile, indicating its potential for safe nasal delivery. It can be concluded that the developed thermogelling formulation showed great promise as an efficient donepezil brain-targeted delivery system. Furthermore, the formulation is worth investigating in vivo for final feasibility confirmation.

Synthesis and Evaluation of Methacrylated Poly(2-ethyl-2-oxazoline) as a Mucoadhesive Polymer for Nasal Drug Delivery

Methacrylated poly(2-ethyl-2-oxazoline) (PEOZ) was synthesized by partial hydrolysis of 500 kDa PEOZ, and the resulting poly[(2-ethyl-2-oxazoline)-co-ethylenimine] P(EOZ-co-EI) was subsequently reacted with methacrylic anhydride. The successful synthesis of methacrylated PEOZ (MAPEOZ) was confirmed by proton nuclear magnetic resonance (1H NMR), infrared spectroscopy, and differential scanning calorimetry. The degrees of hydrolysis and methacrylation were determined by 1H NMR spectra. MAPEOZ exhibited temperature-responsive properties, which were dependent on the degree of methacrylation. On that basis, three soluble MAPEOZ derivatives with different degrees of methacrylation were selected and investigated in cell toxicity studies, showing no significant cytotoxicity against the HEK293 cell line. A slug mucosal irritation assay showed that PEOZ and MAPEOZ do not cause mucosal irritation. The presence of methacryloyl groups and residual amines had a remarkable synergistic effect on the mucoadhesive properties of these polymers. These poly(2-ethyl-2-oxazoline) derivatives have excellent potential as mucoadhesive materials for developing formulations for drug delivery via mucosal routes of administration.

Potential Mucosal Irritation Discrimination of Surface Disinfectants Employed against SARS-CoV-2 by Limacus flavus Slug Mucosal Irritation Assay

Preventive measures have proven to be the most effective strategy to counteract the spread of the SARS-CoV-2 virus. Among these, disinfection is strongly suggested by international health organizations’ official guidelines. As a consequence, the increase of disinfectants handling is going to expose people to the risk of eyes, mouth, nose, and mucous membranes accidental irritation. To assess mucosal irritation, previous studies employed the snail Arion lusitanicus as the mucosal model in Slug Mucosal Irritation (SMI) assay. The obtained results confirmed snails as a suitable experimental model for their anatomical characteristics superimposable to the human mucosae and the different easily observed readouts. Another terrestrial gastropod, Limacus flavus, also known as “ Yellow slug “, due to its larger size and greater longevity, has already been proposed as an SMI assay alternative model. In this study, for the first time, in addition to the standard parameters recorded in the SMI test, the production of yellow pigment in response to irritants, unique to the snail L. flavus, was evaluated. Our results showed that this species would be a promising model for mucosal irritation studies. The study conducted testing among all those chemical solutions most commonly recommended against the SARS-CoV-2 virus.

Nasal Powder Formulations: In-Vitro Characterisation of the Impact of Powders on Nasal Residence Time and Sensory Effects.

Nasal drug delivery is still primarily associated with locally-effective drugs, but next-generation products utilising the benefits of nasal administration—such as easy access to a relatively permeable mucosa, the presence of immunocompetent cells, and a direct route to the brain—are under investigation. Nasal powders offer the potential to improve the drugs’ effects by providing higher resistance against the mucociliary clearance, and thus prolonging the contact time of the drug with its target site. However, suitable and easy-to-use in-vitro setups tailored to the characterisation of this effect are missing. In this study, a selection of excipients for powder formulations were used to evaluate the applicability of different methods which investigate the influence on the contact time. The combination of the assessment of rheological properties, dynamic vapour sorption, and adhesiveness on agar–mucin plates was found to be a valuable predictive tool. For the additional assessment of the sensations associated with the close contact of powders and the mucosa, a slug mucosal irritation assay was conducted and adapted to powders. These methods are regarded as being especially useful for comparative screenings in early formulation development.

The safety profile of Bald\u2019s eyesalve for the treatment of bacterial infections

The rise in antimicrobial resistance has prompted the development of alternatives to combat bacterial infections. Bald’s eyesalve, a remedy used in the Early Medieval period, has previously been shown to have efficacy against Staphylococcus aureus in in vitro and in vivo models of chronic wounds. However, the safety profile of Bald’s eyesalve has not yet been demonstrated, and this is vital before testing in humans. Here, we determined the safety potential of Bald’s eyesalve using in vitro, ex vivo, and in vivo models representative of skin or eye infections. We also confirmed that Bald’s eyesalve is active against an important eye pathogen, Neisseria gonorrhoeae. Low levels of cytotoxicity were observed in eyesalve-treated cell lines representative of skin and immune cells. Results from a bovine corneal opacity and permeability test demonstrated slight irritation to the cornea that resolved within 10 min. The slug mucosal irritation assay revealed that a low level of mucus was secreted by slugs indicating moderate mucosal irritation. We obtained promising results from mouse wound closure experiments; no visible signs of irritation or inflammation were observed. Our results suggest that Bald’s eyesalve could be tested further on human volunteers to assess safety for topical application against bacterial infections.

Using the Slug Mucosal Irritation Assay to Investigate the Tolerability of Tablet Excipients on Human Skin in the Context of the Use of a Nipple Shield Delivery System.

Neonates are particularly challenging to treat. A novel patented drug delivery device containing a rapidly disintegrating tablet held within a modified nipple shield (NSDS) was designed to deliver medication to infants during breastfeeding. However concerns exist around dermatological nipple tolerability with no pharmaceutical safety assessment guidance to study local tissue tolerance of the nipple and the areola. This is the first Slug Mucosal Irritation (SMI) study to evaluate irritancy potential of GRAS excipients commonly used to manufacture rapidly disintegrating immediate release solid oral dosage form METHODS: Zinc sulphate selected as the antidiarrheal model drug that reduces infant mortality, was blended with functional excipients at traditional levels [microcrystalline cellulose, sodium starch glycolate, croscarmellose sodium, magnesium stearate]. Slugs were exposed to blends slurried in human breast milk to assess their stinging, itching or burning potential, using objective values such as mucus production to categorize irritation potency RESULTS: Presently an in vivo assay, previously validated for prediction of ocular and nasal irritation, was used as an alternative to vertebrate models to anticipate the potential maternal dermatological tolerability issues to NSDS tablet components. The excipients did not elicit irritancy. However, mild irritancy was observed when zinc sulphate was present in blends. These promising good tolerability results support the continued investigation of these excipients within NSDS rapidly disintegrating tablet formulations. Topical local tolerance effects being almost entirely limited to irritation, the slug assay potentially adds to the existing preformulation toolbox, and may sit in between the in vitro and existing in vivo assays.

Predicting intramuscular discomfort with the Slug Mucosal Irritation assay

Predicting intramuscular discomfort with the Slug Mucosal Irritation assay

The Slug Mucosal Irritation (SMI) assay: A tool for the evaluation of nasal discomfort

In this research project, the Slug Mucosal Irritation (SMI) assay was applied to predict nasal discomfort, investigating the correlation between responses in slugs and humans.Several SMI experiments and a Human Nose Irritation Test (HNIT) were performed with five NaCl solutions (0.4%, 1.3%, 2.6%, 5.4% and 10.4%) and two benzalkonium chloride solutions (BAC 0.02% and BAC 0.05%). In the HNIT, subjective evaluation of clinical discomfort was performed by 24 participants at several time points.Analyzes reveal that (1) a significant positive association existed between immediate stinging reaction reported by the participants and the mean total mucus production of the slugs (Spearman’s Rank correlation=0.963, p<0.001); (2) NaCl 0.4% was best tolerated in both tests; (3) a concentration-response effect was observed for NaCl and BAC solutions; (4) NaCl 10.4% induced the highest mucus production in the slugs and received higher sting scores for immediate discomfort in the HNIT; (5) stinging sensations decreased rapidly in time and (6) based on these results a new classification prediction model for nasal applications was established.In conclusion, the SMI assay is a promising evaluation method for clinical nasal discomfort. Screening (prototype) formulations with this assay allows formula optimization prior to a clinical trial.

The slug mucosal irritation (SMI) assay: Development of a screening tool for the evaluation of ocular discomfort caused by shampoos

In this research, the slug mucosal irritation (SMI) assay was applied to predict ocular discomfort caused by shampoos to investigate the correlation between responses in slugs and humans.Several SMI experiments and a human eye irritation test (HEIT) were performed with 1 artificial tear solution (ArtTear) and 5 shampoos (A–E; 5%-dilution). In the HEIT, evaluation was performed by participants and an ophthalmologist at several time points.Analyses reveal that (1) a significant positive association existed between immediate stinging reaction reported by the participants and the mean total mucus produced by the slugs (MTMP) (Spearman’s Rank correlation=0.986, p<0.001); (2) ArtTear was best tolerated in both tests; (3) moreover, all shampoos induced higher reactions than ArtTear and water; (4) Shampoo A induced the highest MTMP and received higher scores for immediate discomfort; (5) B was the best tolerated shampoo in both tests, while C, D and E resulted in more pronounced reactions; (6) lacrimation was found not to be statistically correlated with discomfort sensations reported by the participants.The SMI assay is a promising evaluation method for discomfort in the human eye. Screening prototype (eye) formulations with this assay allows formula optimization prior to a HEIT.

New aspects of the Slug Mucosal Irritation assay: predicting nasal stinging, itching and burning sensations

Stinging, itching and/or burning (SIB) sensations cannot be detected by animal tests or in vitro models. In the past, the Slug Mucosal Irritation (SMI) assay demonstrated a relation between an increased mucus production in slugs and an elevated incidence of SIB sensations in humans. A new 1-day SMI test procedure was developed focusing on the prediction of these short-term sensations. The objective of this study was to verify whether this new procedure is capable predicting mucosal tolerance of several marketed nasal formulations using the slug Arion lusitanicus. Irritation and tissue damage were quantified with a 5-day repeated exposure study by means of the mucus produced and proteins and enzymes released. The new protocol predicted SIB sensations by means of mucus production. The effects of six liquid nasal formulations were tested with both protocols, while five physiologic saline solutions were only tested with the new protocol to optimize it. None of the tested liquid nasal formulations resulted in tissue damage; however, exposure to the different formulations had a clear effect on the mucus production of the slugs and moderate discomfort was observed in some cases. These effects were due to the active ingredient, the presence of benzalkonium chloride as a preservative or the hyperosmolality of the formulation. For the most part results agreed with clinical data found in literature. It was concluded that the SMI assay, and the new 1-day protocol in particular, is a good tool to predict nasal clinical discomfort.

Influence of environmental conditions on production of volatiles by Trichoderma atroviride in relation with the sick building syndrome

A Trichoderma atroviride strain was isolated from a water-damaged office and the production of microbial volatile organic compounds (MVOCs) was investigated by means of headspace solid phase microextraction GC–MS. Different growth conditions (substrate, temperature, relative humidity) were selected, resembling indoor parameters, to elucidate a possible relationship between MVOCs, produced by Trichoderma atroviride, and the Sick Building Syndrome. In general, the range of MVOCs and the emitted quantities were larger on malt extract agar (MEA) than on wallpaper and plasterboard. Particular attention was dedicated to the volatile marker 6-pentyl-2-pyrone, a compound produced in high quantities on MEA, and its mucosal irritation potency was shown in a slug mucosal irritation assay. Some compounds characteristic for growth on specific building materials were detected, e.g. 2-ethylcyclopentanone, menthone, iso-menthone and trans- p-menth-2-en-7-ol on plasterboard and 4-heptanone and 1-octen-3-ol on wallpaper. Relative humidity and substrate had a more important effect on MVOC production than temperature.

Influence of reaction medium during synthesis of Gantrez® AN 119 nanoparticles for oral vaccination

Two synthesis methods of poly(methyl vinyl ether-co-maleic anhydride) (Gantrez® AN 119) nanoparticles (NP) (used for oral vaccination) were compared. Wheat germ agglutinin (WGA) was used as ligand to enhance the bioadhesive properties of NP and β-galactosidase as antigen. The first method encapsulated β-galactosidase in NP by co-precipitation in an acetone/water mixture containing 44% acetone. In the second method, antigen addition occurred in 100% acetone. To improve stability, NP were crosslinked with 1,3-diaminopropane. The stability of WGA-conjugated NP with encapsulated antigen diminished at lower pH and when decreasing the amount of crosslinker. The binding type between WGA and polymer depended on the synthesis method: predominantly ionic bonds were formed using the 44% acetone method, whereas synthesis via the 100% acetone method resulted in covalent bonds. The biological activity of the WGA coating, evaluated via a pig gastric mucin binding test, was lower in NP prepared via the 100% acetone method. No release of native antigen was detected after hydrolysis of NP, due to the covalent antigen binding during antigen encapsulation and the high reactivity of the polymer. Moreover, the mucosal irritation capacity was evaluated upon nanoparticle hydrolysis using a slug mucosal irritation assay. Herein, hydrolysed NP of the 44% acetone method were classified as mild irritative.

Mucosal Irritation Potential of Personal Lubricants Relates to Product Osmolality as Detected by the Slug Mucosal Irritation Assay

The slug mucosal irritation assay has recently been used as a sensitive measure of mucus membrane tolerance for vaginal microbicide products and carriers. In the current study, it was determined whether musosal irritation potency of personal lubricants is related to varying product osmolalities. Five commercial lubricants with an osmolality range were evaluated using the previously validated slug mucosal irritation assay. Specifically, Arion lusitanicus were treated with lubricants over 5 days to quantify mucus production and tissue damage, allowing assignment of each product into an irritation potency category (none, mild, moderate, or severe). The irritation potency (assessed by the mucus production) of the lubricants showed a significant, quadratic relationship with the product osmolality (P = 0.001; R (2) = 0.99). Femglide, a hypo-osmotic lubricant (32 mOsm/kg), caused a negative mucus production. Pré, an iso-osmotic lubricant (316 mOsm/kg), caused no changes. Two moderately hyperosmotic lubricants, Replens and K-Y jelly (2143 and 2463 mOsm/kg), induced mild and moderate irritation, respectively. The highly hyperosmotic lubricant Astroglide (5848 mOsm/kg) resulted in severe irritation and tissue damage. Commonly used personal lubricants show a full range of mucosal irritation potential, which is related to product osmolality.

164 Evaluation of the effects of chronic subcytotoxic surfactant exposure to a human corneal cell line

In this research, the slug mucosal irritation (SMI) assay was applied to predict ocular discomfort caused by shampoos to investigate the correlation between responses in slugs and humans.Several SMI experiments and a human eye irritation test (HEIT) were performed with 1 artificial tear solution (ArtTear) and 5 shampoos (A–E; 5%-dilution). In the HEIT, evaluation was performed by participants and an ophthalmologist at several time points.Analyses reveal that (1) a significant positive association existed between immediate stinging reaction reported by the participants and the mean total mucus produced by the slugs (MTMP) (Spearman’s Rank correlation = 0.986, p < 0.001); (2) ArtTear was best tolerated in both tests; (3) moreover, all shampoos induced higher reactions than ArtTear and water; (4) Shampoo A induced the highest MTMP and received higher scores for immediate discomfort; (5) B was the best tolerated shampoo in both tests, while C, D and E resulted in more pronounced reactions; (6) lacrimation was found not to be statistically correlated with discomfort sensations reported by the participants.The SMI assay is a promising evaluation method for discomfort in the human eye. Screening prototype (eye) formulations with this assay allows formula optimization prior to a HEIT.