Eduardo Palacios

Anti-inflammatory and anti-bacterial activity, and CYTOTOXICITY of halloysite surfaces

Halloysite is a naturally-occurring nanomaterial occurring in the thousands of tons and that serves as biomaterial, with applications in the areas of biotechnology, pharmaceutical, and medical research. This study reports on the anti-inflammatory, cytotoxic, and anti-oxidant activity of halloysite Jarrahdale (collected at ∼ 45 km SE of Perth, Western Australia; JA), Dragon Mine (provided by Natural Nano Inc., Rochester, New York; NA), and Kalgoorie Archean (collected at Siberia, ∼ 85km NW of Kalgoorlie, West Australia; PA). Prior to biological testing, halloysites were characterized by 27Al and 29Si Nuclear Magnetic Resonance Spectroscopy, the anti-inflammatory activity was determined by (a) the mouse ear edema method, using 12-o-tetradecanoylphorbol-13-acetate (TPA) as anti-inflammatory agent; and (b) the myeloperoxidase enzymatic activity method (MPO). Cell viability was determined using the MTT method. Sample characterization by NMR method showed similar symmetry and atomic environments, with no evidence of distortion(s) due to shiftings in atomic ordering or electron density. The anti-inflammatory activity followed the order: PA>JA>NA, and remained invariant with time. Prolonged anti-inflammatory activity related inversely to surface area and lumen space. The low extent of infiltration at shorter reaction times confirmed a limiting number of active surface sites. EPR intensity signals followed the order: JA>NA>PA. The poor stabilization of RO species in PA suspensions was explained by tube alignment provoking occlusion, thus limiting transfer of H(+) or e(-) from-and-to the surface, and decreases in acidity associated to Al(oct). Cell viability (%) varied from one surface to the other, PA(92.3 ± 6.0), JA(84.9 ± 7.8), and NA(78.0 ± 5.6), but related directly to SBET values.

Role of bentonite clays on cell growth.

Bentonites, naturally occurring clays, are produced industrially because of their adsorbent capacity but little is known about their effects on human health. This manuscript reports on the effect of bentonites on cell growth behaviour. Bentonites collected from India (Bent-India), Hungary (Bent-Hungary), Argentina (Bent-Argentina), and Indonesia (Bent-Indonesia) were studied. All four bentonites were screened in-vitro against two human cancer cell lines [U251 (central nervous system, glioblastoma) and SKLU-1 (lung adenocarcinoma)] supplied by the National Cancer Institute (USA). Bentonites induced growth inhibition in the presence of U251 cells, and growth increment in the presence of SKLU-1 cells, showing that interactions between bentonite and cell surfaces were highly specific. The proliferation response for U251 cells was explained because clay surfaces controlled the levels of metabolic growth components, thereby inhibiting the development of high-grade gliomas, particularly primary glioblastomas. On the other hand, the proliferation response for SKLU-1 was explained by an exacerbated growth favoured by swelling, and concomitant accumulation of solutes, and their hydration and transformation via clay-surface mediated reactions.

Cinnabar-Preserved Bone Structures from Primary Osteogenesis and Fungal Signatures in Ancient Human Remains

The Red Queen remains (700 A.C.) found at Palenque, México, are examples of cinnabar (HgS) application to royal remains during pre-Hispanic times. The Red Queen remains are those of a ca. 30–35-yr-old female and present a striking similarity to the remains of another Mayan woman found at Copan, Honduras. Thus, covering the remains of royal women with HgS may have been a common practice in the Mayan civilization. High resolution microdiffraction and microscopic analysis of the Red Queen remains showed the presence of nanotubular organic minerals comparable in composition and molecular dimensions to collagen fibrils, and in spatial ordering to collagen fiber networks. Fungal structures are rare in the geological record because of poor preservation potential. Micrographs revealed the preservation of fungal signatures, with morphology comparable to parasitic fungal-coral matrix associations, consistent with the idea that fungal remains can be preserved in environments which contain high Hg concentrations. The well-preserved signatures of fungus-animal interactions and primary osteogenesis in the Red Queen remains are attributed to the long-term antibacterial activity of HgS and the association of sulfur components with nanotubular structures.

Lipid peroxidation and cytotoxicity induced by respirable volcanic ash.

This paper reports that the main component of respirable volcanic ash, allophane, induces lipid peroxidation (LP), the oxidative degradation of lipids in cell membranes, and cytotoxicity in murin monocyle/macrophage cells. Naturally-occurring allophane collected from New Zealand, Japan, and Ecuador was studied. The quantification of LP was conducted using the Thiobarbituric Acid Reactive Substances (TBARS) assay. The cytotoxic effect was determined by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide colorimetric assay. Electron-Paramagnetic Resonance (EPR) determinations of naturally-occurring allophane confirmed the incorporation in the structure and clustering of structural Fe(3+), and nucleation and growth of small-sized Fe (oxyhydr)oxide or gibbsite. LP induced by allophane varied with time, and solid concentration and composition, reaching 6.7 ± 0.2 nmol TBARS mg prot(-1). LP was surface controlled but not restricted by structural or surface-bound Fe(3+), because redox processes induced by soluble components other than perferryl iron. The reactivity of Fe(3+) soluble species stemming from surface-bound Fe(3+) or small-sized Fe(3+) refractory minerals in allophane surpassed that of structural Fe(3+) located in tetrahedral or octahedral sites of phyllosilicates or bulk iron oxides. Desferrioxamine B mesylate salt (DFOB) or ethylenediaminetetraacetic acid (EDTA) inhibited LP. EDTA acted as a more effective inhibitor, explained by multiple electron transfer pathways. Registered cell-viability values were as low as 68.5 ± 6.7%.

Antibacterial clay against gram-negative antibiotic resistant bacteria

Antibiotic resistant bacteria persist throughout the world because they have evolved the ability to express various defense mechanisms to cope with antibiotics and the immune system; thus, low-cost strategies for the treatment of these bacteria are needed, such as the usage of environmental minerals. This paper reports the antimicrobial properties of a clay collected from Brunnenberg, Germany, that is composed of ferroan saponite with admixtures of quartz, feldspar and calcite as well as exposed or hidden (layered at inner regions) nano Fe(0). Based on the growth curves (log phase) of six antibiotic resistant bacteria (4 gram-negative and 2 gram-positive), we concluded that the clay acted as a bacteriostat; however, the clay was only active against the gram-negative bacteria (except for resilient Klebsiella pneumonia). The bacteriostatic mode of action was evidenced by the initial lack of Colony Forming Units on agar plates with growth registered afterward, certainly after 24h, and can be explained because interactions between membrane lipopolysaccharides and the siloxane surfaces of the clay. Labile or bioavailable Fe in the clay (extracted by EDTA or DFO-B) induced the quantitative production of HO as well as oxidative stress, which, nevertheless, did not account for by its bacteriostatic activity.

Biological dissolution and activity of the Allende meteorite

This paper reports on the effect of the Allende meteorite on the integrity of biological material and addresses the question whether it can induce cell damage via oxidative stress and cell mortality. The reaction mechanisms addressed herein are studied using electron-paramagnetic resonance spectroscopy (EPR), high-resolution transmission electron microscopy, scanning electron microscopy and energy dispersive spectroscopy, high-resolution X-ray diffraction, and the assays for thiobarbituric acid reactive substances (TBARS) and cell viability using 3-(4,5)-dimethylthiazol-2-yel-2,5-diphenyltetrazolium bromide (MTT bromide). As determined by the TBARS assay, Allende specimens induced cell damage via oxidative stress. The contents of TBARS in suspensions containing 1000 ppm of Allende and Fe 1– x S were 6.8 ± 0.7 and 5.8 ± 0.6 nmol/mg protein, respectively. EPR experiments conducted on reaction mixtures containing Allende, 5,5-dimethyl-1-pyrroline- N -oxide (DMPO), and H 2 O 2 showed a quartet signal, a 1:2:2:1 intensity, and hyperfine coupling constants corresponding to a N = 1.49 mT and a H = 1.49 mT, a signature of the DMPO-OH adduct. The intensity of the signal depended on the concentration of the solids in suspension, while the formation of DMPO-OH was limited by H 2 O 2 . Experiments were conducted to test for the production of the DMPO-OH adduct from ferric ions, and the plausible generation of HO • . The role of ethanol (CH 3 CH 2 OH) as scavenger of HO • in Allende-DMPO suspensions was addressed. Results showed a six-line spectra, with hyperfine coupling constants a N = 15.8 G, a H = 22.6 G, and g = 2.0059, consistent with the formation of the DMPO-CH(OH)-CH 3 adduct, but not DMPO-OCH 2 CH 3 . We explain these findings as the result of formation of HO • onto (or in proximity to) the mineral surface, with CH 3 CH 2 OH competing with DMPO for HO ∞ , and ferric iron playing a lesser role in DMPO transformation. Our findings are congruent with reported radical-scavenging experiments for pyrite under anoxic conditions, concluding the formation of HO ∞ at surface defect sites. Experiments conducted in Allende–desferrioxamine B(DFO-B) suspensions showed the inhibition of the formation of HO • , by means of decreases in the DMPO-OH adduct signal, accounted for by the reaction between Fe(II) and HO • to form Fe(III) and competing reaction mechanisms at the structural Fe centers, confirming that the production of HO ∞ radicals is associated with iron centers and contributes to mineral dissolution. Small-sized magnetite domains present were recognized as catalytic sites for the production of HO ∞ radicals. The γ-Fe 3 O 4 domains present in the Allende matrix exhibited a submicron range, an elongated-hexagonal habit, and a high degree of crystallinity, supporting the presence of biogenic γ-Fe 3 O 4 . Cell viability was found to be susceptible to the distribution and atomic environment of structural Fe.

Effect of Functionalized Carbon Nanotubes and their Citric Acid Polymerization on Mesenchymal Stem Cells In Vitro

The effects of acid-functionalized and polycitric acid- (PCA-) polymerized carbon nanotubes (CNTs) in contact with the extracellular membrane of mesenchymal stem cells (MSC), a genetically unmodified cell line with differentiation capability, was evaluated with different cellular parameters. The modified CNTs show differences in the analyzed biological behaviors, that is, intracellular incorporation, cell proliferation, apoptosis, and cytotoxicity as compared with those unpolymerized nanotubes. Due to the reduced cellular uptake of polymerized CNTs, PCA-polymerized CNTs are less cytotoxic and are associated with less apoptotic cell death than the acid-functionalized ones. The effects of nitrogen-doped CNTs (CNx) is also reported, showing that functionalized undoped CNTs present strong stimulation of cell proliferation, whereas functionalized and polymerized CNxs stimulate an apoptotic behavior. The study of MSCs in contact with CNTs and PCA is challenging due to the complexity of its various signaling components. Our results provide basis for further studies aimed to understand the relevant role that the interaction of these nanotubes with extracellular membrane could have a crucial structure for tissue grafting.