Novel nanostructured lipid carriers loading Apigenin for anterior segment ocular pathologies.

Dry eye disease (DED) is a chronic multifactorial disorder of the ocular surface caused by tear film dysfunction and constitutes one of the most common ocular conditions worldwide. However, its treatment remains unsatisfactory. While artificial tears are commonly used to moisturize the ocular surface, they do not address the underlying causes of DED. Apigenin (APG) is a natural product with anti-inflammatory properties, but its low solubility and bioavailability limit its efficacy. Therefore, a novel formulation of APG loaded into biodegradable and biocompatible nanoparticles (APG-NLC) was developed to overcome the restricted APG stability, improve its therapeutic efficacy, and prolong its retention time on the ocular surface by extending its release. APG-NLC optimization, characterization, biopharmaceutical properties and therapeutic efficacy were evaluated. The optimized APG-NLC exhibited an average particle size below 200 nm, a positive surface charge, and an encapsulation efficiency over 99 %. APG-NLC exhibited sustained release of APG, and stability studies demonstrated that the formulation retained its integrity for over 25 months. In vitro and in vivo ocular tolerance studies indicated that APG-NLC did not cause any irritation, rendering them suitable for ocular topical administration. Furthermore, APG-NLC showed non-toxicity in an epithelial corneal cell line and exhibited fast cell internalization. Therapeutic benefits were demonstrated using an in vivo model of DED, where APG-NLC effectively reversed DED by reducing ocular surface cellular damage and increasing tear volume. Anti-inflammatory assays in vivo also showcased its potential to treat and prevent ocular inflammation, particularly relevant in DED patients. Hence, APG-NLC represent a promising system for the treatment and prevention of DED and its associated inflammation.

Gene expression regulating lipopolysaccharide and lipid metabolic processes in porcine oral mucosal cells in long-term primary in vitro culture.

Lipids are an alternative energy source for cells and provide structural integrity in cell membrane and their metabolism is regulated with the use of different pathways, such as integrin signalling, oxidative stress, mechanical stress, and pH changes. All of those processes take place in the oral mucosa which is subject to different environmental impacts. In this study, porcine buccal pouch mucosal cells (pBPMCs) were used during long-term primary in vitro culture. The cultured cells were collected at 7, 15 and 30 days of IVC and subsequently transferred to RNA isolation. In the results of the following microarray analysis, we analyzed the genes detected, belonging to ontology groups, such as "cellular lipid metabolic process", "response to lipid" and "response to lipopolysaccharides. All of the genes involved in these ontological groups were expressed at higher levels at 7 days of IVC and substantially decreased in expression at days 15 and 30 of primary culture. We observed new genes, which may be recognized as markers in regulation of lipid metabolism in mucosal cells in vitro. The results suggested that the biochemical mechanism-involved lipids were accompanied by increased enzymatic activation and synthesis of crucial growth factors reaching high activity at day 7 of culture, which is also well documented as a stage of tissue regeneration period within oral mucosa. Therefore, this "biochemical fingerprint" may be an additional checkpoint of the integrity, resistance and easy adaptability of oral tissues, which are important conditions of success in tissue engineering and grafting for tissue reconstruction.

Chromatographic identification of a new flavin derivative in plain yogurt.

The presence of flavin derivatives in plain yogurt was assessed by high-performance liquid chromatography (HPLC) and thin-layer chromatography (TLC). The total amount of flavins in yogurts produced by different companies was variable and oscillated between 150.0 and 218.8 microg/100 g. Riboflavin (RF) was the predominant flavin. Besides RF, flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN), 7alpha-hydroxyriboflavin (7alpha-HRF), 4'- or 5'-D-riboflavin-beta-D-galactoside (RFgal), and traces of 10-formylmethylflavin (10-FMF) and 10-hydroxyethylflavin (10-HEF) have been found. It is known that RFgal may be obtained using enzymes or cultures of different microorganisms, but its presence in foodstuffs has not been demonstrated, yet.

Chromatographic determination of flavin derivatives in baker's yeast.

The presence of flavin derivatives in baker's yeast was tested by high-performance liquid chromatography and thin-layer chromatography. In yeast samples, besides flavin adenine dinucleotide and flavin mononucleotide, small amounts of riboflavin and traces of 10-formylmethylflavin have been found. Total amount of flavins was calculated to be 17.9 +/- 2.9 micrograms/g of fresh yeast. The distribution of flavin adenine dinucleotide, flavin mononucleotide, riboflavin and 10-formylmethylflavin in total flavin content were estimated to be 71.5%, 25.8%, 1.7% and below 0.05%, respectively. In some samples we have additionally detected small amounts (0.8% of total flavins) of new flavin derivative which has been identified as 4',5'-riboflavin cyclic phosphate by means of its chromatographic and chemical behaviour. This compound seems to be a product of flavin adenine dinucleotide degradation and probably has been earlier mistaken for flavin mononucleotide. Its formation is dependent on pH conditions.