Secretory Products in Petals of Centaurea cyanus L. Flowers: A Histochemistry, Ultrastructure, and Phytochemical Study of Volatile Compounds.

(1) Background: Centaurea cyanus L. is a medicinal plant whose flowers are widely used in herbal medicine. The aim of the study was to localise flower tissues that are responsible for the production of secretory products in petals and to analyse the volatile compounds. The volatile compounds of the flowers of this species have not been investigated to date. (2) Methods: Light, fluorescence, scanning and transmission electron microscopy techniques were used in the study. Lipophilic compounds were localised in the tissues using histochemical assays. Volatile compounds were determined with the use of solid phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS). (3) Results: The study showed production of secretion in the petal parenchyma, whose ultrastructure has features of a secretory tissue. The lipophilic secretion was localised in the cells and intercellular spaces of the parenchyma and in the walls and surface of epidermal cells, where it accumulated after release through cuticle microchannels. Sesquiterpenes were found to constitute the main group of volatile compounds, with the highest content of ß-caryophyllene (26.17%) and α-humulene (9.77%). (4) Conclusions: Given the presence of some volatile components that are often found in resins (caryophyllene, delta-cadinene) and the abundant secretion residues on the epidermal surface, we suppose that the C. cyanus secretion released by the flowers is a resinaceous mixture (oleoresin), which is frequently found in plants, as shown by literature data. This secretion may play an important role in the therapeutic effects of C. cyanus flowers.

Hemodialysis Effect on the Composition of the Eye Fluid of Cataract Patients.

Numerous reports have proven that dialysis patients experience disturbances in the levels of elements in biological fluids. Disturbances in the homeostasis of essential elements or the appearance of highly toxic elements are serious problems also in clinical ophthalmology. The purpose of this study was to investigate the influence of hemodialysis (HD) on the elemental composition of anterior chamber aqueous humor (AH) in patients undergoing cataract surgery. The study involved 22 patients. The control group enrolled 16 patients (age 75.68 ± 9.67, female 54.55%, male 45.45%) with cataract and normal kidney function (control), and the second group included six patients (age 70.33 ± 12.74, female 33.33%, male 66.67%) with cataract undergoing HD treatment. The elements quantification was established using an inductively coupled plasma optical emission spectrometer (ICP-MS). In the eye fluid of dialysis patients, there were increased levels of manganese (Mn) and mercury (Hg) and decreased levels of vanadium (V) and zinc (Zn). In addition, a statistically significant increase in the Hg/Zn and Hg/selenium (Se) ratios and a lowering of the iron (Fe)/Mn ratio were observed in the studied group in comparison to the control. The obtained results indicated the need for Zn and Se supplementation in order to eliminate the hazards caused by Hg toxicity. A lower level of V in the eye fluid of dialysis patients may have a positive effect on maintaining a calcium and phosphorus homeostasis. Our study gives a deep insight into changes of elements concentrations in AH induced by HD.

New Perspectives for Eye-Sparing Treatment Strategies in Primary Uveal Melanoma.

Uveal melanoma is the most common intraocular malignancy and arises from melanocytes in the choroid, ciliary body, or iris. The current eye-sparing treatment options include surgical treatment, plaque brachytherapy, proton beam radiotherapy, stereotactic photon radiotherapy, or photodynamic therapy. However, the efficacy of these methods is still unsatisfactory. This article reviews several possible new treatment options and their potential advantages in treating localized uveal melanoma. These methods may be based on the physical destruction of the cancerous cells by applying ultrasounds. Two examples of such an approach are High-Intensity Focused Ultrasound (HIFU)-a promising technology of thermal destruction of solid tumors located deep under the skin and sonodynamic therapy (SDT) that induces reactive oxygen species. Another approach may be based on improving the penetration of anti-cancer agents into UM cells. The most promising technologies from this group are based on enhancing drug delivery by applying electric current. One such approach is called transcorneal iontophoresis and has already been shown to increase the local concentration of several different therapeutics. Another technique, electrically enhanced chemotherapy, may promote drug delivery from the intercellular space to cells. Finally, new advanced nanoparticles are developed to combine diagnostic imaging and therapy (i.e., theranostics). However, these methods are mostly at an early stage of development. More advanced and targeted preclinical studies and clinical trials would be needed to introduce some of these techniques to routine clinical practice.

Phenolic Constituents of Lamium album L. subsp. album Flowers: Anatomical, Histochemical, and Phytochemical Study.

Flos Lamii albi has a high biological activity and is widely used in herbal medicine. The aim of the study was to characterize the secretory structures present in Lamium album subsp. album corolla and the location of phenolic compounds. Additionally, we carried out qualitative phytochemical analyses of flavonoids and phenolic acids. Light, fluorescence, and scanning electron microscopy were used to analyze the structure of the floral organs. The main classes of phenolic compounds and their localization were determined histochemically. Phytochemical analyses were performed with high-performance thin-layer chromatography (HPTLC) and high-performance liquid chromatography (HPLC). Six types of glandular trichomes were found which contained flavonoids, phenolic acids, and tannins. The phytochemical studies demonstrated the presence of caffeic, chlorogenic, ferulic, gallic, p-coumaric, protocatechuic, syringic, gentisic, and vanillic phenolic acids as well as rutoside, isoquercetin, and quercetin flavonoids. The corolla in L. album subsp. album has antioxidant properties due to the presence of various polyphenols, as shown by the histo- and phytochemical analyses. The distribution and morphology of trichomes and the content of phenolic compounds in the corolla have taxonomic, pharmacognostic, and practical importance, facilitating the identification of the raw material.

Aqueous humor selenium level and open-angle glaucoma.

PURPOSE: Selenium supplementation was seen to be linked to glaucoma disease in a previous study (Lillico A. JE, Reid M et al. (2002) Selenium Supplementation and Risk of Glaucoma in the NPC trial University of Arizona, Tucson, AZ., Arizona Cancer Center). As aqueous humor levels of selenium seemed to be associated with primary open-angle glaucoma (POAG), the aim of this study was to analyze concentrations of selenium in aqueous humor samples of patients with POAG and pseudoexfoliation glaucoma (PEXG) in comparison to normal samples. PATIENTS AND METHODS: Thirty-eight aqueous humor samples from patients undergoing cataract surgery were collected: Eleven patients with PEXG (age 65.8 ±â€¯10.69, female 6, male 5), 12 patients with POAG (age 65.3 ±â€¯10.50, female 7, male 5) and 15 patients without glaucoma (age 70.9 ±â€¯12.83, female 10, male 5, controls). Aqueous humor levels of selenium were measured by Flow-Injection-Inductively-Coupled-Plasma-DRC-Mass-Spectrometry (ICP-DRC-MS). RESULTS: Maximum likelihood estimation of the least squares means (LS-means) and the relative 95% confidence limits of selenium aqueous humor levels were 6.90 ±â€¯1.03 µg/L (control), 6.74 ±â€¯1.14 µg/L (POAG) and 8.25 ±â€¯1.18 µg/L (PEXG). The data were modeled using a generalized linear model (GLM) analysis, where selenium was set as dependent variable. The model was corrected for group differences in age and gender. The data show no differences among all the calculated differences between the least square means (LS means), taking in consideration the simultaneous 95% confidence limit and the multiple comparison tests with Tukey-Cramer adjustment. The evaluation of the model disclosed that POAG and PEXG patients had no significantly different aqueous humor selenium concentrations compared to controls and to each other. However, the quantile regression analysis of selenium aqueous humor levels showed differences in quantiles for open-angle glaucoma patients considering age and gender. CONCLUSION: As no significant difference in aqueous humor concentration of selenium was detected between open-angle glaucoma and controls, however, quantile analysis showed differences in quantiles levels for different age ranges in open-angle glaucoma patients, the trace element selenium seemed to be linked to glaucoma disease, yet not in a major role.

The New TLC Method for Separation and Determination of Multicomponent Mixtures of Plant Extracts.

The new mode of two-dimensional gradient thin layer chromatography (MGD-2D TLC) has been presented. Short distance development of sample in the first dimension leads to formation of the preconcentrated narrow zones. They are consecutively separated in the second dimension with the mobile phase gradient in several steps of development until the eluent reaches the further end of the chromatographic plate. The use of the above-mentioned technique allows isolating and then identifying the compounds of various polarity from the multicomponent mixture. The practical application of two-dimensional gradient thin layer chromatography has been performed for isolation of the two plant (Juniperus and Thymus) oils components as the examples of test mixtures. The experiments have been carried out with the use of silica gel plates as well as a normal phase condition. The results of solute separation with isocratic one-dimensional thin layer chromatography system have been compared with those of two-dimensional gradient system. It has been observed that application of the latter mode leads to almost triplicated number of zones in comparison with the former one. It is purposeful to apply the proposed mode to control the purity of the dominant component or components of the mixture.

Herbal preparation extract for skin after radiotherapy treatment. Part One--Preclinical tests.

Naran R is a herbal composition made of Plantago lanceolate folium, Malvae arboreae flos, Calendulae flos, Chamomillae inflorescentia, Lamii albi flos to prepare compresses or to wash skin with inflammations. The extract of this preparation is mixed to be applied as an ointment on patients' skin after radiotherapy. Experiments performed in vitro are part of pre-clinical tests with Naran R ointment. This study examined the impact of the plant composition for ethanol-water extract on human skin fibroblasts (HSF) culture. Samples of extract, prepared from patented amounts of herbs, were in the range of 25-225 µg/mL. Six methods were applied: standard spectrophotometric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, neutral red (NR) uptake assay, DPPH free radical scavenging test, labeling of cytoskeleton F-actin, staining of argyrophilic nucleolar organizer regions (AgNORs) and trypan blue coloration. The extract concentration 75 µg/mL was established as safe for application on human skin. In labeling of F-actin with rhodamine-phalloidin dye at this concentration the cytoskeleton was stable. The extract did not influence the membrane stability and had positive influence on the proliferation activity. It was confirmed in AgNOR test during incubation with extract, which led to formation of larger amount of smaller nucleolins. In DPPH scavenging activity test, the extract revealed over 8% higher free-radical scavenging activity in comparison to control. After trypan blue staining, the extract in concentration 125 µg/mL significantly lowered the cell viability. When the cytotoxic and anti-proliferative activity of the extracts were analyzed, MTT and Neutral Red (NR) methods were used. The cells' viability was maintained on a constant level (80-110%) after 24, 48 and 72 h of incubation. During all time of NR test (72 h) and even when 225 µg/mL of extract was applied, the viability of cells was in range 80-110% of control. Positive influence of the extract on investigated cells structure and proliferation, lack of toxicity and increasing anti-oxidant activity enable to consider this preparation as a natural remedy with potential application in skin therapy after radiation.

Assessment of eyebright (euphrasia officinalis L.) extract activity in relation to human corneal cells using in vitro tests.

BACKGROUND: Euphrasia officinalis L. is an herb traditionally used in folk medicine, mainly in the treatment of eye disorders. AIMS: The present study analyzed the activity of three extracts of E. officinalis L. (ethanol, ethyl acetate and heptane) on cultured human corneal epithelial cells (10.014 pRSV-T). STUDY DESIGN: In vitro study. METHODS: Toxicity, free radical scavenging activity and the immunomodulatory effects of the extracts were tested using the thiazolyl blue tetrazolium bromide (MTT) or Neutral Red, 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and ELISA tests, respectively. Moreover, nitric oxide levels and cytoskeleton architecture were analyzed after corneal cell incubation with the plant extracts. RESULTS: We show that the biological effect depended on both the concentration and the extraction solvent used. Heptane extracts, distinct from those in ethanol and ethyl acetate, were toxic to 10.014 pRSV-T cells at low concentrations (25 µg/mL) and did not demonstrate free radical scavenging effects. All tested extracts decreased pro-inflammatory cytokine expression (IL-1ß, IL-6 and TNF-α) and also anti-inflammatory IL-10 expression by human corneal cells when the extracts were added to the cell culture medium for 24 h. CONCLUSION: In conclusion, we show that the promising effects of the application of E. officinalis L. preparations as a supplementary therapy for eye disorders are associated with the ethanol and ethyl acetate extracts, not the heptane extract.