Marine versus Non-Marine Bacterial Exopolysaccharides and Their Skincare Applications.

Bacteria are well-known to synthesize high molecular weight polysaccharides excreted in extracellular domain, which constitute their protective microenvironment. Several bacterial exopolysaccharides (EPS) are commercially available for skincare applications in cosmetic products due to their unique structural features, conferring valuable biological and/or textural properties. This review aims to give an overview of bacterial EPS, an important group of macromolecules used in cosmetics as actives and functional ingredients. For this purpose, the main chemical characteristics of EPS are firstly described, followed by the basics of the development of cosmetic ingredients. Then, a focus on EPS production, including upstream and downstream processes, is provided. The diversity of EPS used in the cosmetic industry, and more specifically of marine-derived EPS is highlighted. Marine bacteria isolated from extreme environments are known to produce EPS. However, their production processes are highly challenging due to high or low temperatures; yield must be improved to reach economically viable ingredients. The biological properties of marine-derived EPS are then reviewed, resulting in the highlight of the challenges in this field.

Different Ways to On-Line Hyphenate Centrifugal Partition Chromatography and Mass Spectrometry: Application to Prenylated Xanthones from Garcinia mangostana.

Centrifugal partition chromatography is a liquid-liquid separation method well adapted for the fractionation or purification of natural compounds from plant extracts. However, following the preparative isolation, the fractions collected must be analysed by high-performance thin-layer chromatography or high-performance liquid chromatography to evaluate their composition and/or their purity. These additional steps are time-consuming and increase the risk of compound degradation. In order to get an instantaneous analysis of fraction content with structural information on the phytochemicals eluted, it is possible to hyphenate on-line centrifugal partition chromatography with mass spectrometry. Depending on the complexity of the extract, two different kinds of centrifugal partition chromatography-mass spectrometry coupling can be performed: centrifugal partition chromatography-mass spectrometry or centrifugal partition chromatography-high-performance liquid chromatography-mass spectrometry coupling. In the first case, one part of the centrifugal partition chromatography effluent is directly introduced in the mass spectrometry ionisation source to identify the eluted compounds, while in the second case, it is directed to a high-performance liquid chromatography-mass spectrometry system where compounds are first separated thanks to high-performance liquid chromatography and then identified using mass spectrometry.

Bioactive molecules in Kalanchoe pinnata leaves: extraction, purification, and identification.

Kalanchoe pinnata (Lam.) Pers. (syn. Bryophyllum pinnatum; family Crassulaceae) is a popular plant used in traditional medicine in many temperate regions of the world and particularly in South America. In Guyana, the leaves are traditionally used as an anti-inflammatory and antiseptic to treat coughs, ulcers, and sores. The purpose of this study was to implement a method for targeting and identifying molecules with antimicrobial activity, which could replace chemical preservatives in cosmetic applications. The leaves were extracted by a method based on pressurized liquid extraction (PLE), using different solvents. A study of antimicrobial activity and cytotoxicity tests were performed to select the most interesting extract. To isolate one or more active molecules, the selected crude extract was fractionated by centrifugal partition chromatography (CPC) and then antimicrobial activity and cytotoxicity of each fraction were tested under the same procedure. The last step consisted of identifying the main compounds in the most active fraction by LC-MS/MS.

Hyphenation of centrifugal partition chromatography with electrospray ionization mass spectrometry using an active flow-splitter device for characterization of flavonol glycosides.

Online coupling of centrifugal partition chromatography to electrospray ionization mass spectrometry (CPC/ESI-MS) was investigated for the separation and characterization of flavonol glycosides. Structural identification and purification monitoring of analytes on milligram scale were demonstrated to be possible by using an active flow-splitter device which transfers automatically and successively, at discrete frequencies, small aliquots of the chromatographic effluent to an independent auxiliary stream directed to an ESI quadrupole mass spectrometer. The CPC protocol used a biphasic solvent system composed of ethyl acetate/ethanol/water (4.5:1:4.5, v/v/v) in isocratic mode. During the separation process, continuous acquisition of mass spectral data of the isolated flavonols from the effluent was performed in the negative ion mode with an auxiliary stream composed of 50 mM ammonium acetate/ethanol (2:8, v/v) delivered by a secondary pump. To demonstrate the potential of this hyphenated technique, flavonol glycosides from an apple peel extract were identified, purified and quantitatively analyzed. Calibration curves and limits of detection are also detailed.

Pilot-scale ion-exchange centrifugal partition chromatography: purification of sinalbin from white mustard seeds.

The purification of p-hydroxybenzylglucosinolate (sinalbin) on a multigram scale from a crude aqueous extract of white mustard seeds (Sinapis alba var. concerta) was successfully achieved by scaling up a strong ion-exchange centrifugal partition chromatography (SIXCPC) laboratory procedure. Thus, the one-step sinalbin purification was performed with 2.35 g of crude extract in approximately 170 min (830 mg/h) up to 70.3 g in approximately 160 min (26.3 g/h) by switching from a 200 mL laboratory scale column to a 5.7 L pilot-scale column. The required biphasic solvent system contained ethyl acetate, n-butanol, and water in 3:2:5 v/v/v proportions, Aliquat 336 (trioctylmethyl ammonium chloride) was added to the organic stationary phase (80 mM) and acted as ion-exchanger. Potassium iodide in the aqueous mobile phase (80 mM) was used as sinalbin displacer. The 28.5 mass scale factor arose from the increase in mobile phase flow-rate (from 2 to 50 mL/min), from the higher mass of injected white mustard seed extract (from 12 to 350 g), and from the calculated productivity (from 830 mg to 26.3 g). These results demonstrate that industry scale production of glucosinolates is easily performed by SIXCPC, thus providing pure reference standards for pharmacology studies.

Blocking negative effects of senescence in human skin fibroblasts with a plant extract.

There is increasing evidence that senescent cells are a driving force behind many age-related pathologies and that their selective elimination increases the life- and healthspan of mice. Senescent cells negatively affect their surrounding tissue by losing their cell specific functionality and by secreting a pro-tumorigenic and pro-inflammatory mixture of growth hormones, chemokines, cytokines and proteases, termed the senescence-associated secretory phenotype (SASP). Here we identified an extract from the plant Solidago virgaurea subsp. alpestris, which exhibited weak senolytic activity, delayed the acquisition of a senescent phenotype and induced a papillary phenotype with improved functionality in human dermal fibroblasts. When administered to stress-induced premature senescent fibroblasts, this extract changed their global mRNA expression profile and particularly reduced the expression of various SASP components, thereby ameliorating the negative influence on nearby cells. Thus, the investigated plant extract represents a promising possibility to block age-related loss of tissue functionality.

Strong ion-exchange centrifugal partition chromatography as an efficient method for the large-scale purification of glucosinolates.

The glucosinolates sinalbin and glucoraphanin were purified by strong ion-exchange displacement centrifugal partition chromatography (SIXCPC). The optimized conditions involved the biphasic solvent system ethyl acetate/n-butanol/water (3:2:5, v/v), the lipophilic anion-exchanger Aliquat 336 (trioctylmethylammonium chloride, 160 and 408 mM) and a sodium iodide solution (80 and 272 mM) as displacer. Amounts as high as 2.4 g of sinalbin and 2.6g of glucoraphanin were obtained in one step in 2.5 and 3.5h respectively, starting from 12 and 25 g of mustard and broccoli seed aqueous extracts, using a laboratory scale CPC column (200 mL inner volume).

Preparative isolation of huperzines A and B from Huperzia serrata by displacement centrifugal partition chromatography.

The pH-zone refining centrifugal partition chromatography technique was used to separate the two acetylcholinesterase inhibitors huperzines A and B from a crude alkaloid extract of the club moss Huperzia serrata. Complete co-elution of huperzines A and B was initially observed with the well-known methyl tert-butyl ether-acetonitrile-water (4:1:5, v/v/v) solvent system with triethylamine (8mM) as the displacer and methane sulfonic acid (6mM) as the retainer. An efficient biphasic system was designed on the basis of solvent association that provided selectivity in the elution mode: n-heptane/ethyl acetate/n-propanol/water (5:15:35:45, v/v/v/v). Lowering the bridge solvent content (n-propanol) of this system increased the polarity difference between the two phases thus adapting it to the pH-zone refining mode. Thus, the purification of these compounds was achieved using the biphasic system n-heptane/ethyl acetate/n-propanol/water (10:30:15:45, v/v/v/v) with triethylamine (8mM) as the displacer and methane sulfonic acid (6mM) as the retainer.

Novel seco-dibenzopyrrocoline alkaloid from Cryptocarya oubatchensis.

[structure: see text] A novel seco-dibenzopyrrocoline alkaloid, named oubatchensine 6, and five phenanthroindolizidines (1-5) were isolated from Cryptocarya oubatchensis, and their structures were elucidated. Displacement centrifugal partition chromatography was used to purify compounds 1 and 6. Structure determination of the latter was carried out by mass spectrometry, NMR spectroscopy, quantum chemistry, and computer-assisted structure determination. Cytotoxic activity against KB cells was then investigated.

Multiple dual-mode centrifugal partition chromatography, a semi-continuous development mode for routine laboratory-scale purifications.

Nowadays, centrifugal partition chromatography (CPC) separations can be routinely achieved at the laboratory scale. The solvent system selection has been made easy, as generic sets of solvent systems are described in publications and books. This approach, however, generally reduces the scope of optimization strategies for two important parameters: selectivity and sample solubility. This can be very limiting for the preparative separation of structurally similar compounds. Multiple dual-mode (MDM) CPC has been developed to provide an easy-to-use alternative technique to circumvent this problem. A MDM separation consists of a succession of dual-mode runs (i.e. multiple inversion of stationary and mobile phase) that can only be achieved because both chromatographic phases are liquids. This original elution mode is thus a semi-continuous process with a classical sample injection and which only requires a single CPC column. Underlying mechanisms of MDM were studied using a model mixture of acenaphthylene and naphthalene. A mixture of two synthetic pairs of diastereomers was then successfully submitted to MDM CPC, in the framework of the synthesis of biologically active compounds.

Purification of antibiotics from the biocontrol agent Streptomyces anulatus S37 by centrifugal partition chromatography.

A novel actinomycete strain, Streptomyces anulatus S37, has been isolated from the rhizosphere of healthy Moroccan Vitis vinifera on the basis on its ability to promote grapevine growth and to induce natural defences against various phytopathogens. In the present work, the main bioactive metabolites produced by S. anulatus S37 were isolated. A crude n-BuOH extract of the S37 fermentation broth was firstly partitioned in a biphasic solvent system composed of n-heptane, methanol, and water (5:1.5:3.5, v/v). The most active organic fraction (1.1g) as revealed by TLC-bioautography was subsequently separated by a two-step centrifugal partition chromatography procedure. The first separation was performed in the ascending mode at 6mL/min with the biphasic solvent system n-heptane, ethyl acetate, methanol and water (2:1:2:1, v/v), to finally recover 40mg of a pure compound identified as streptochlorin by NMR spectroscopy. In a second separation, the solvent system n-heptane, acetonitrile, and water (5:5:4, v/v) was used in the ascending mode at 3mL/min to purify 135mg of nigericin and 53mg of piericidin A1. Assays performed with the three compounds have confirmed their inhibitory impact on the growth of Botryris cinerea in dual confrontation and also on V. vinifera L. plantlets.

Simultaneous presence of unsaturation and long alkyl chain at P'1 of Ilomastat confers selectivity for gelatinase A (MMP-2) over gelatinase B (MMP-9) inhibition as shown by molecular modelling studies.

Structural analogues of Ilomastat (Galardin), containing unsaturation(s) and chain extension carrying bulky phenyl group or alkyl moieties at P'1 were synthesized and purified by centrifugal partition chromatography. They were analyzed for their inhibitory capacity towards MMP-1, MMP-2, MMP-3, MMP-9 and MMP-14, main endopeptidases involved in tumour progression. Presence of unsaturation(s) decreased the inhibitory potency of compounds but, in turn increased their selectivity for gelatinases. 2b and 2d derivatives with a phenyl group inhibited preferentially MMP-9 with IC50 equal to 45 and 38 nM, respectively, but also display activity against MMP-2 (IC50 equal to 280 and 120 nM, respectively). Molecular docking computations confirmed affinity of these substances for both gelatinases. With aims to obtain a specific gelatinase A (MMP-2) inhibitor, P'1 of Ilomastat was modified to carry one unsaturation coupled to an alkyl chain with pentylidene group. Docking studies indicated that MMP-2, but not MMP-9, could accommodate such substitution; indeed 2a proved to inhibit MMP-2 (IC50=123 nM), while displaying no inhibitory capacity towards MMP-9.