The interweaving roles of mineral and microbiome in shaping the antibacterial activity of archaeological medicinal clays.

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal Earths (MEs), natural aluminosilicate-based substances (largely kaolinite and montmorillonite), have been part of the European pharmacopoeia for well over two millennia; they were used generically as antidotes to 'poison'. AIM OF THE STUDY: To test the antibacterial activity of three Lemnian and three Silesian Earths, medicinal earths in the collection of the Pharmacy Museum of the University of Basel, dating to 16th-18th century and following the methodology outlined in the graphical abstract. To compare them with natural clays of the same composition (reference clays) and synthetic clays (natural clays spiked with elements such as B, Al, Ti and Fe); to assess the parameters which drive antibacterial activity, when present, in each group of samples. MATERIALS AND METHODS: a total of 31 samples are investigated chemically (ICP-MS), mineralogically (both bulk (XRD) and at the nano-sized level (TEM-EDAX)); their organic load (bacterial and fungal) is DNA-sequenced; their bioactivity (MIC60) is tested against Gram-positive, S. aureus and Gram-negative, P. aeruginosa. RESULTS: Reference smectites and kaolinites show no antibacterial activity against the above pathogens. However, the same clays when spiked with B or Al (but not with Ti or Fe) do show antibacterial activity. Of the six MEs, only two are antibacterial against both pathogens. Following DNA sequencing of the bioactive MEs, we show the presence within of a fungal component, Talaromyces sp, a fungus of the family of Trichocomaceae (order Eurotiales), historically associated with Penicillium. Talaromyces is a known producer of the exometabolite bioxanthracene B, and in an earlier publication we have already identified a closely related member of the bioxanthracene group, in association with one of the LE samples examined here. By linking fungus to its exometabolite we suggest that this fungal load may be the key parameter driving antibacterial activity of the MEs. CONCLUSIONS: Antibacterial activity in kaolinite and smectite clays can arise either from spiking natural clays with elements like B and Al, or from an organic (fungal) load found only within some archaeological earths. It cannot be assumed, a priori, that this organic load was acquired randomly and as a result of long-term storage in museum collections. This is because, at least in the case of medicinal Lemnian Earth, there is historical evidence to suggest that the addition of a fungal component may have been deliberate.

Greco-Roman mineral (litho)therapeutics and their relationship to their microbiome: The case of the red pigment miltos.

This paper introduces a holistic approach to the study of Greco-Roman (G-R) lithotherapeutics. These are the minerals or mineral combinations that appear in the medical and scientific literature of the G-R world. It argues that they can best be described not simply in terms of their bulk chemistry/mineralogy but also their ecological microbiology and nanofraction component. It suggests that each individual attribute may have underpinned the bioactivity of the lithotherapeutic as an antibacterial, antifungal or other. We focus on miltos, the highly prized, naturally fine, red iron oxide-based mineral used as a pigment, in boat maintenance, agriculture and medicine. Five samples (four geological (from Kea, N. Cyclades) and one archaeological (from Lemnos, NE Aegean)) of miltos were analyzed with physical and biological science techniques. We show that: a. Kean miltos and Lemnian earth/miltos must have been chemically and mineralogically different; b. Lemnian miltos must have been more effective as an antibacterial against specific pathogens (Gram + and Gram - bacteria) than its Kean counterpart; c. two samples of Kean miltos, although similar, chemically, mineralogically and eco-microbiologically (phylum/class level), nevertheless, displayed different antibacterial action. We suggest that this may constitute proof of microbial ecology playing an important role in effecting bioactivity and, interestingly, at the more specific genus/species level. From the perspective of the historian of G-R science, we suggest that it may have been on account of its bioactivity, rather than simply its 'red-staining' effect, that miltos gained prominent entry into the scientific and medical literature of the G-R world.

Electrophoretic deposition of ZnO/alginate and ZnO-bioactive glass/alginate composite coatings for antimicrobial applications.

Two organic/inorganic composite coatings based on alginate, as organic matrix, and zinc oxide nanoparticles (n-ZnO) with and without bioactive glass (BG), as inorganic components, intended for biomedical applications, were developed by electrophoretic deposition (EPD). Different n-ZnO (1-10 g/L) and BG (1-1.5 g/L) contents were studied for a fixed alginate concentration (2 g/L). The presence of n-ZnO was confirmed to impart antibacterial properties to the coatings against gram-negative bacteria Escherichia coli, while the BG induced the formation of hydroxyapatite on coating surfaces thereby imparting bioactivity, making the coating suitable for bone replacement applications. Coating composition was analyzed by thermogravimetric analysis (TG), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and energy dispersive X-ray spectroscopy (EDS) analyses. Scanning electron microscopy (SEM) was employed to study both the surface and the cross section morphology of the coatings. Polarization curves of the coated substrates made in cell culture media at 37 °C confirmed the corrosion protection function of the novel organic/inorganic composite coatings.

A stable droplet reactor for high temperature nanocrystal synthesis.

We report a versatile capillary-based droplet reactor for the controlled synthesis of nanoparticles over a wide range of flow conditions and temperatures. The reactor tolerates large flow-rate differentials between individual reagent streams, and allows droplet composition to be varied independently of residence time and volume. The reactor was successfully applied to the synthesis of metal (Ag), metal-oxide (TiO(2)) and compound semiconductor (CdSe) nanoparticles, and in each case exhibited stable droplet flow over many hours of operation without fouling, even for reactions involving solid intermediates. For CdSe formed by the reaction of Cd oleate and Se, highly controlled growth could be achieved at temperatures of up to 250 °C, with emission spectra varying smoothly and reproducibly with temperature and flow-rate. The droplet reactor showed exceptional stability when operated under constant flow-rate and temperature conditions, yielding particles with well-defined band-edge emission spectra that did not vary over the course of a full day's continuous operation.

Struvite crystallisation and recovery using a stainless steel structure as a seed material.

A metallic system acting as a seed substrate has been designed and developed in order to assess its efficiency in recovering phosphorus as struvite. The device, consisting of two concentric stainless steel meshes, was immerged in the upper section of a pilot crystallisation reactor fed with synthetic liquors (MgCl(2) x 6H(2)O, NH(4)H(2)PO(4),) for 2h. Apart from soluble PO(4)-P removals which remained in the range 79-80% with or without application of the metallic system, it was found that under the specific operating conditions tested the meshes were capable of accumulating struvite at a rate of 7.6 gm(-2)h(-1), hence reducing significantly the amount of fine particles remaining in solution from 302.2 to 12 mg L(-1) when compared to trials without mesh.

Kinetics of struvite precipitation: effect of the magnesium dose on induction times and precipitation rates.

The presence of white deposits in specific areas of wastewater treatment plants is generally the consequence of the spontaneous formation of a mineral called struvite. Struvite forms when the levels of phosphate, ammonium and magnesium naturally available in wastewater effluents reach a minimum molar ratio 1:1:1 under specific conditions of pH, temperature and mixing energy. Originally regarded as a phenomenon to be controlled or eliminated, struvite has been lately identified as an alternative way of removing and recovering P from wastewater effluents and generating a product identified as an excellent base for the production of slow release fertilisers. Chemical and physical principles of struvite precipitation and the development of crystallisation technologies have been widely investigated. However, little interest has been given to kinetics of struvite precipitation. In the present work the kinetics of struvite formation have been investigated at both laboratory and pilot scale in synthetic solutions containing Mg(2+), NH4(+), and PO4(3-) ions in a molar ratio 1:2:2 at room temperature. These different tests have used pH measurements to assess the impact of water chemistry on induction times, and more precisely the influence of magnesium levels on kinetic rates. Experimental results and kinetic calculations revealed that the control of the magnesium dose initially present in solution is decisive of the speed at which struvite nucleates.

Impact of reactor operation on success of struvite precipitation from synthetic liquors.

A pilot scale reactor was designed and developed to study struvite crystallisation principles. The present work focuses on the possible impact of the reactor's operating parameters on struvite characteristics, and evaluates the performances of the process in removing phosphorus. Struvite precipitation from synthetic liquors was investigated under various situations including: pH, magnesium dosing, addition of foreign ions such as calcium and increasing retention time. Small variations of all these parameters were found to have significant effects on struvite crystal characteristics and/or production. For instance, an increase of pH from 10.0 to 10.5 favoured the formation of Mg3(PO4)2.22H2O rather than struvite. For molar ratios Ca:Mg above 1:1, calcium ions competed with magnesium to form an amorphous calcium phosphate, hence inhibiting struvite formation. With regards to crystal growth, the process showed some limitations. Indeed, large amounts of fines were produced, and crystal rarely grew over 100 microm under optimum conditions. Based on those observations, zeta-potential measurements of struvite crystals were investigated. Results revealed highly negative zeta-potential values for all experiments, indicating that this may be a limitation to struvite tendency for agglomeration.

Use of bone meal amendments to immobilise Pb, Zn and Cd in soil: A leaching column study.

The aim of this study is to test the stabilisation of metals in contaminated soils via the formation of low-solubility metal phosphates. Bone apatite, in the form of commercially available bone meal, was tested as a phosphate source on a mine waste contaminated made-ground with high levels of Pb, Zn and Cd. Triplicate leaching columns were set up at bone meal to soil ratios of 1:25 and 1:10, in addition to unamended controls, and were run for 18 months. The columns were irrigated daily with a synthetic rain solution at pH of 2, 3, and 4.4. After 100 days, the leachate Pb, Zn and Cd concentrations of all amended columns were significantly reduced. For 1:10 treatments, release of these metals was suppressed throughout the trial. For 1:25 treatments, Zn and Cd concentrations in the leachates began to increase after 300 days. DTPA and water extractions showed that Pb and Cd were more strongly held in the amended soils. This study concludes that the complexity of soil processes and the small quantities of metals sequestered precluded determination of a metal immobilisation mechanism.

Effect of bone meal (calcium phosphate) amendments on metal release from contaminated soils--a leaching column study.

Metal-contaminated soil may be remediated in situ by the formation of highly insoluble metal phosphates if an appropriate phosphorus (P) source can be found. Leaching column experiments have been carried out to assess the suitability of bone meal as such a source. Bone meal additions reduced metal release from a contaminated soil, increased soil and leachate pH and decreased soil leachate toxicity. Minimal P leaching occurred from the soil. The data are consistent with a proton consuming bone meal (calcium phosphate) dissolution reaction followed by the formation of metal phosphates. Although, no metal phosphates were observed to form using X-ray diffraction of scanning electron microscopy this could be due to their low concentration. Relatively low (1:50 bone meal:soil) concentrations of fine (90-500 microns) bone meal would appear to be an effective treatment for metal-contaminated soils.