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In the last years, the medicinal plant Perilla frutescens (L.) Britton has gained scientific interest because leaf extracts, due to the presence of rosmarinic acid and other polyphenols, have shown anti-allergic and skin protective potential in pre-clinical studies. Nevertheless, the lack of standardized extracts has limited clinical applications to date. In this work, for the first time, a standardized phytocomplex of P. frutescens, enriched in rosmarinic acid and total polyphenols, was produced through innovative in vitro cell culture biotechnology and tested. The activity of perilla was evaluated in an in vitro inflammatory model of human keratinocytes (HaCaT) by monitoring tight junctions, filaggrin, and loricrin protein levels, the release of pro-inflammatory cytokines and JNK MAPK signaling. In a practical health care application, the perilla biotechnological phytocomplex was tested in a multilayer model of vaginal mucosa, and then, in a preliminary clinical observation to explore its capacity to preserve vaginal mucosal integrity in women in peri-menopause. In keratinocytes cells, perilla phytocomplex demonstrated to exert a marked activity in epidermis barrier maintenance and anti-inflammatory effects, preserving tight junction expression and downregulating cytokines release through targeting JNK activation. Furthermore, perilla showed positive effects in retaining vaginal mucosal integrity in the reconstructed vaginal mucosa model and in vivo tests. Overall, our data suggest that the biotechnological P. frutescens phytocomplex could represent an innovative ingredient for dermatological applications.

The negative impact of using conventional fungicides in plant disease protection has increased the interest in safer alternatives such as plant secondary metabolites, generally having a better toxicological profile. However, cultivation conditions and plant material strongly affect the quality and quantity of secondary metabolites obtained from field grown plants, limiting the standardization needed for industrial production. Plant cell culture technology can provide highly homogeneous biomasses with specific chemical characteristics. A phytocomplex with high rosmarinic acid content (10.12% w/w) was obtained from a selected cell line of Salvia officinalis and was tested against the grapevine downy mildew pathogen, Plasmopara viticola. Grapevine leaf discs were sprayed with the phytocomplex at 5 g/L and then inoculated with P. viticola sporangia. Sporulation level on each disc was assessed after 7 days with an image processing software. The phytocomplex reduced by 95% the sporulation level compared to the control and was also more effective than rosmarinic acid alone, used at the same concentration found in the phytocomplex. Persistence of the phytocomplex was also assessed: when applied 5 days before inoculation, it reduced by 90% the sporulation level compared to the control. These results highlight the possibility to take advantage of cell culture techniques to produce safer pesticides with high quality standards.

Echinacea purpurea (L.) Moench is one of the most economically important medicinal plants, cultivated worldwide for its high medicinal value and with several industrial applications in both pharmaceutical and food industries. Thanks to its various phytochemical contents, including caffeic acid derivatives (CADs), E. purpurea extracts have antioxidant, anti-inflammatory, and immuno-stimulating properties. Among CADs, chicoric acid is one of the most important compounds which have shown important pharmacological properties. The present research was aimed at optimizing the production of chicoric acid in E. purpurea cell culture. Methyl jasmonate (MeJa) at different concentrations and for different duration of treatments was utilized as elicitor, and the content of total polyphenols and chicoric acid was measured. Several genes involved in the chicoric acid biosynthetic pathway were selected, and their expression evaluated at different time points of cell culture growth. This was performed with the aim of identifying the most suitable putative molecular markers to be used as a proxy for the early prediction of chicoric acid contents, without the need of expensive quantification methods. A correlation between the production of chicoric acid in response to MeJa and an increased response to oxidative stress was also proposed.

Plant biomasses growing in bioreactor could be developed as production systems for cosmetic ingredients, nutraceuticals and food additives. We previously reported that the red carrot cell line R4G accumulates high levels of anthocyanins, which are potent antioxidants with multiple health-promoting properties. To investigate the industrial potential of this cell line in detail, we tested extract for antioxidant and anti-inflammatory activity in the mouse monocyte/macrophage cell-line J774A.1 and in reconstructed skin tissue models. We also compared the R4G extract to commercial carrot extracts in terms of stability and metabolomic profiles. We found that the R4G extract have potent antioxidant and anti-inflammatory activities, protecting mammalian cells from the oxidative stress triggered by exposure to bacterial lipopolysaccharides and H2O2. The extract also inhibited the nuclear translocation of NF-κB in an epidermal skin model, and induced the expression of VEGF-A to promote the microcirculation in a dermal microtissue model. The anthocyanins extracted from R4G cells were significantly more stable than those found in natural red carrot extracts. Finally, we showed that R4G extract has similar metabolomic profile of natural extracts by using a combination of targeted and untargeted metabolomics analysis, demonstrating the safety of R4G carrot cells for applications in the nutraceutical and food/feed industries.

This study focused on the biological evaluation and chemical characterization of a new ingredient obtained by in vitro cell culture of Rhus coriaria L. An in vitro plant cell culture method permits to cultivate plant in a short period of time and to obtain extract with a high safety profile for the consumer, free from heavy metals, pesticides, aflatoxins, bacterial or fungal contamination. Through the selection of specific cell culture media, it was possible to obtain a Rhus coriaria cell line with a high content of gallic acid derivatives. The Rhus coriaria L. phytocomplex (RC-P), containing 7.6% w/w of acid gallic derivatives, was obtained by drying of plant cell biomass after 14 days of growth in the final selected culture medium. UPLC-ESI-MS and UPLC-DAD analysis allowed to identify numerous gallic acid derivatives, such as galloyl hexose, trigalloyl hexose and high molecular weight galloyl derivatives, and to quantify their overall content. The antioxidant activity of the RC-P was tested by DPPH assay and the wound healing activity was evaluated using a scratch wound healing test on human keratinocytes and fibroblasts. This work showed that RC-P could be a new effective cosmetic ingredient with antioxidant and skin repair activity.

Skin is being increasingly exposed to artificial blue light due to the extensive use of electronic devices, which can induce cell oxidative stress, causing signs of early photo aging. The Melissa officinalis phytocomplex is a new standardized cosmetic ingredient obtained by an in vitro plant cell culture with a high content of rosmarinic acid. In this study, we examine the activity of the Melissa officinalis phytocomplex to protect skin against blue light and infrared damages, evaluating the ROS (Radical Oxygen Species) level in keratinocyte cell line from human skin (HaCaT) and Nrf2 (Nuclear factor erythroid 2-related factor 2), elastin, and MMP1 (Matrix Metalloproteinase 1) immunostaining in living human skin explants ex vivo. This phytocomplex demonstrates antioxidant activity by reducing ROS production and thus the oxidant damage of the skin caused by UV and blue light exposure. In addition, it inhibits blue light-induced Nrf2 transcriptional activity, IR-induced elastin alteration, and IR-induced MMP-1 release. This Melissa officinalis phytocomplex is a new innovative active ingredient for cosmetic products that is able to protect skin against light and screen exposure damages and oxidative stress.