A fresh perspective on infrared spectroscopy as a prescreening method for molecular and stable isotopes analyses on ancient human bones.

Following the development of modern genome sequencing technologies, the investigation of museum osteological finds is increasingly informative and popular. Viable protocols to help preserve these collections from exceedingly invasive analyses, would allow greater access to the specimens for scientific research. The main aim of this work is to survey skeletal tissues, specifically petrous bones and roots of teeth, using infrared spectroscopy as a prescreening method to assess the bone quality for molecular analyses. This approach could overcome the major problem of identifying useful genetic material in archaeological bone collections without resorting to demanding, time consuming and expensive laboratory studies. A minimally invasive sampling of archaeological bones was developed and bone structural and compositional changes were examined, linking isotopic and genetic data to infrared spectra. The predictive model based on Infrared parameters is effective in determining the occurrence of ancient DNA (aDNA); however, the quality/quantity of aDNA cannot be determined because of the influence of environmental and local factors experienced by the examined bones during the burial period.

An Atomistic Model Describing the Structure and Morphology of Cu-Doped C-S-H Hardening Accelerator Nanoparticles.

Calcium silicate hydrate (C-S-H) is the main binding phase in Portland cement. The addition of C-S-H nanoparticles as nucleation seeds has successfully been used to accelerate the hydration process and the precipitation of binding phases either in conventional Portland cement or in alternative binders. Indeed, the modulation of the hydration kinetics during the early-stage dissolution-precipitation reactions, by acting on the nucleation and growth of binding phases, improves the early strength development. The fine-tuning of concrete properties in terms of compressive strength and durability by designed structural modifications can be achieved through the detailed description of the reaction products at the atomic scale. The nano-sized, chemically complex and structurally disordered nature of these phases hamper their thorough structural characterization. To this aim, we implement a novel multi-scale approach by combining forefront small-angle X-ray scattering (SAXS) and synchrotron wide-angle X-ray total scattering (WAXTS) analyses for the characterization of Cu-doped C-S-H nanoparticles dispersed in a colloidal suspension, used as hardening accelerator. SAXS and WAXTS data were analyzed under a unified modeling approach by developing suitable atomistic models for C-S-H nanoparticles to be used to simulate the experimental X-ray scattering pattern through the Debye scattering equation. The optimization of atomistic models against the experimental pattern, together with complementary information on the structural local order from 29Si solid-state nuclear magnetic resonance and X-ray absorption spectroscopy, provided a comprehensive description of the structure, size and morphology of C-S-H nanoparticles from the atomic to the nanometer scale. C-S-H nanoparticles were modeled as an assembly of layers composed of 7-fold coordinated Ca atoms and decorated by silicate dimers and chains. The structural layers are a few tens of nanometers in length and width, with a crystal structure resembling that of a defective tobermorite, but lacking any ordering between stacking layers.

Urea-functionalized amorphous calcium phosphate nanofertilizers: optimizing the synthetic strategy towards environmental sustainability and manufacturing costs.

Nanosized fertilizers are the new frontier of nanotechnology towards a sustainable agriculture. Here, an efficient N-nanofertilizer is obtained by post-synthetic modification (PSM) of nitrate-doped amorphous calcium phosphate (ACP) nanoparticles (NPs) with urea. The unwasteful PSM protocol leads to N-payloads as large as 8.1 w/w%, is well replicated by using inexpensive technical-grade reagents for cost-effective up-scaling and moderately favours urea release slowdown. Using the PSM approach, the N amount is ca. 3 times larger than that obtained in an equivalent one-pot synthesis where urea and nitrate are jointly added during the NPs preparation. In vivo tests on cucumber plants in hydroponic conditions show that N-doped ACP NPs, with half absolute N-content than in conventional urea treatment, promote the formation of an equivalent amount of root and shoot biomass, without nitrogen depletion. The high nitrogen use efficiency (up to 69%) and a cost-effective preparation method support the sustainable real usage of N-doped ACP as a nanofertilizer.

On the amorphous layer in bone mineral and biomimetic apatite: A combined small- and wide-angle X-ray scattering analysis.

The occurrence of an amorphous calcium phosphate layer covering the crystalline apatite core has been suggested to be an intrinsic feature of both bone mineral and synthetic biomimetic analogs. However, an exahustive quantitative picture of the amorphous-crystalline relationship in these materials is still missing. Here, we present a multiple scale modelling that combines small-angle X-ray scattering (SAXS) and synchrotron wide-angle X-ray total scattering (WAXTS) analyses to investigate the amorphous-crystalline spatial interplay in bone sample and biomimetic carbonated nano-apatites. SAXS analysis indicates the presence of a single morphology consisting of tiny nanoplates (NPLs) and provides a measure of their thickness (falling in the 3-5 nm range). WAXTS analysis was performed by developing atomistic models of apatite NPLs incorporating lattice strain, mostly attributed to the carbonate content, and calculating the X-ray patterns using the Debye Scattering Equation. Upon model optimization, the size and strain parameters of the crystalline platelets were derived and the amorphous component, co-existing with the crystalline one, separated and quantified (in the 23-33 wt% range). Notably, the thickness of the apatite core was found to exhibit nearly null (bone) or minor (< 0.5 nm, biomimetic samples) deviations from that of the entire NPLs, suggesting that the amorphous material remains predominantly distributed along the lateral sides of the NPLs, in a core-crown-like arrangement. The lattice strain analysis indicates a significant stiffness along the c axis, which is comparable in bone and synthetic samples, and larger deformations in the other directions. STATEMENT OF SIGNIFICANCE: Current models of bone mineral and biomimetic nanoapatites suggest the occurrence of an amorphous layer covering the apatitic crystalline nanoplates in a core-shell arrangement. By combining X-ray scattering techniques in the small and wide angle regions, we propose a joint atomic-to-nanometre scale modelling to investigate the amorphous-crystalline interplay within the nanoplates. Estimates are extracted for the thickness of the entire nanoplates and the crystalline core, together with the quantification of the amorphous fraction and apatite lattice strain. Based on the thickness matching, the location of the amorphous material mostly along the edges of the nanoplates is inferred, with a vanishing or very thin layer in the thickness direction, suggesting a core-crown-like arrangement, with possible implications on the mineral surface reactivity.

The role of nanoparticle structure and morphology in the dissolution kinetics and nutrient release of nitrate-doped calcium phosphate nanofertilizers.

Bio-inspired synthetic calcium phosphate (CaP) nanoparticles (NPs), mimicking the mineral component of bone and teeth, are emergent materials for sustainable applications in agriculture. These sparingly soluble salts show self-inhibiting dissolution processes in undersaturated aqueous media, the control at the molecular and nanoscale levels of which is not fully elucidated. Understanding the mechanisms of particle dissolution is highly relevant to the efficient delivery of macronutrients to the plants and crucial for developing a valuable synthesis-by-design approach. It has also implications in bone (de)mineralization processes. Herein, we shed light on the role of size, morphology and crystallinity in the dissolution behaviour of CaP NPs and on their nitrate doping for potential use as (P,N)-nanofertilizers. Spherical fully amorphous NPs and apatite-amorphous nanoplatelets (NPLs) in a core-crown arrangement are studied by combining forefront Small-Angle and Wide-Angle X-ray Total Scattering (SAXS and WAXTS) analyses. Ca2+ ion release rates differ for spherical NPs and NPLs demonstrating that morphology plays an active role in directing the dissolution kinetics. Amorphous NPs manifest a rapid loss of nitrates governed by surface-chemistry. NPLs show much slower release, paralleling that of Ca2+ ions, that supports both detectable nitrate incorporation in the apatite structure and dissolution from the core basal faces.

Engineering Biomimetic Calcium Phosphate Nanoparticles: A Green Synthesis of Slow-Release Multinutrient (NPK) Nanofertilizers.

Biomimetic calcium phosphate nanoparticles (CaP) have been actively used in biomedicine, due to their high biodegradability and biocompatibility. However, much less progress has been made regarding their application in precision agriculture, i.e., for the controlled delivery of active species to plants. Herein, we report a straightforward and green synthetic method to dope CaP with potassium (K) and nitrogen (N, as nitrate and urea). By modulating the synthetic conditions in terms of maturation time (at 37 °C) and doping, we prepared K- and N-doped nanoparticles in the form of either amorphous calcium phosphate (ACP) or nanocrystalline apatite (Ap) and studied the impact of the dopants on the ACP-to-Ap transformation pathways. Importantly, we found out that ACP, isolated at low maturation times, incorporates nitrogen (in the form of nitrate and urea) to a larger extent than Ap (2.6 vs 1.1 wt %, respectively). Multinutrient nanofertilizers (so-called nanoU-NPK) with the following composition (wt %) were obtained: Ca (23.3), P (10.1), K (1.5), NO3 (2.9), and urea (4.8). The nanoU-NPK provides a slow and gradual release of the most important plant macronutrients (NPK), with N in two chemical forms, and different kinetics. The concentration of nutrients supplied by 10 g L-1 of nanoU-NPK to the media after 1 week (in mg L-1) was Ca (27.0), P (6.2), K (41.0), NO3 (134.0), and urea (315.0). Preliminary tests on durum wheat have shown that the application of nanoU-NPK allows reducing the amount of nitrogen supplied to the plants by 40% with respect to a conventional treatment, without affecting the final kernel weight per plant. The application of these slow-release NPK nanofertilizers is a promising strategy toward enhancing the efficiency of the fertilization, complying with the concept of precision agriculture.

A universal curve of apatite crystallinity for the assessment of bone integrity and preservation.

The reliable determination of bioapatite crystallinity is of great practical interest, as a proxy to the physico-chemical and microstructural properties, and ultimately, to the integrity of bone materials. Bioapatite crystallinity is used to diagnose pathologies in modern calcified tissues as well as to assess the preservation state of fossil bones. To date, infrared spectroscopy is one of the most applied techniques for bone characterisation and the derived infrared splitting factor (IRSF) has been widely used to practically assess bioapatite crystallinity. Here we thoroughly discuss and revise the use of the IRSF parameter and its meaning as a crystallinity indicator, based on extensive measurements of fresh and fossil bones, virtually covering the known range of crystallinity degree of bioapatite. A novel way to calculate and use the infrared peak width as a suitable measurement of true apatite crystallinity is proposed, and validated by combined measurement of the same samples through X-ray diffraction. The non-linear correlation between the infrared peak width and the derived ISRF is explained. As shown, the infrared peak width at 604 cm-1 can be effectively used to assess both the average crystallite size and structural carbonate content of bioapatite, thus establishing a universal calibration curve of practical use.

Raman hyperspectral imaging as an effective and highly informative tool to study the diagenetic alteration of fossil bones.

Retrieving the pristine chemical or isotopic composition of archaeological bones is of great interest for many studies aiming to reconstruct the past life of ancient populations (i.e. diet, mobility, palaeoenvironment, age). However, from the death of the individual onwards, bones undergo several taphonomic and diagenetic processes that cause the alteration of their microstructure and composition. A detailed study on bone diagenesis has the double purpose to assess the preservation state of archaeological bones and to understand the alteration pathways, thus providing evidence that may contribute to evaluate the reliability of the retrieved information. On these bases, this research aims to explore the effectiveness of Raman hyperspectral imaging to detect types, extent and spatial distribution of diagenetic alteration at the micro-scale level. An early-Holocene bone sample from the Al Khiday cemetery (Khartoum, Sudan) was here analysed. Parameters related to the collagen content, bioapatite crystallinity and structural carbonate content, and to the occurrence of secondary mineral phases were calculated from Raman spectra. The acquired data provided spatially-resolved information on both the preservation state of bone constituents and the diagenetic processes occurring during burial. Given the minimal sample preparation, the easy and fast data acquisition and the improvement of system configurations, micro-Raman spectroscopy can be extensively applied as a screening method on a large set of samples in order to characterise the preservation state of archaeological bones. This technique can be effectively applied to identify suitable and well preserved portions of the analysed sample on which perform further analyses.

Correction: Long-distance connections in the Copper Age: New evidence from the Alpine Iceman's copper axe.

[This corrects the article DOI: 10.1371/journal.pone.0179263.].

Differentiating between long and short range disorder in infra-red spectra: on the meaning of "crystallinity" in silica.

Local atomic disorder and crystallinity are structural properties that influence greatly the resulting chemical and mechanical properties of inorganic solids, and are used as indicators for different pathways of material formation. Here, these structural properties are assessed in the crystals of quartz based on particle-size-related scattering processes in transmission infra-red spectroscopy. Independent determinations of particle size distributions in the range 2-100 µm of a single crystal of quartz and defective quartz with highly anisotropic micro-crystallites show that particle sizes below the employed wavelength (approx 10 µm) exhibit asymmetric narrowing of absorption peak widths, due to scattering processes that depend on the intra-particle structural defects and long range crystallinity. In particular, we observe that the 1079 cm-1 peak could be used to assess crystallinity, because it shows an asymmetric peak shape shift toward a higher wavelength, depending on the crystallite size. We observe that the 694 cm-1 peak could be used to assess local atomic disorder as it does not show scattering and peak shape changes when absorption effects dominate, below 2 µm. We propose coupling particle size assessments with infra-red peak shape analysis as a method to characterize crystallinity and short range order for studying recrystallization in natural silica, as well as defectivity in many different types of silicas used for industrial and technological applications.

Long-distance connections in the Copper Age: New evidence from the Alpine Iceman's copper axe.

25 years after the discovery in the Ötztal Italian Alps, the 5,300-year-old mummy keeps providing key information on human biological and medical conditions, aspects of everyday life and societal organization in the Copper Age. The hand axe found with the body of the Alpine Iceman is one of the rare copper objects that is firmly dated to the early Copper Age because of the radiocarbon dating of the axe wooden shaft. Here we report the measurement of the lead isotope ratios of the copper blade. The results unambiguously indicate that the source of the metal is the ore-rich area of Southern Tuscany, despite ample evidence that Alpine copper ore sources were known and exploited at the time. The experimental results are discussed within the framework of all the available coeval archaeometallurgical data in Central-Southern Europe: they show that the Alps were a neat cultural barrier separating distinct metal circuits. The direct evidence of raw metal or object movement between Central Italy and the Alps is surprising and provides a new perspective on long-distance relocation of goods and relationships between the early Copper Age cultures in the area. The result is in line with the recent investigations re-evaluating the timing and extent of copper production in Central Italy in the 4th millennium BC.

Late Pleistocene/Early Holocene Evidence of Prostatic Stones at Al Khiday Cemetery, Central Sudan.

The recovery of three stone-like ovoid objects within the burial of a pre-Mesolithic (Late Pleistocene/Early Holocene) individual at Al Khiday cemetery (Central Sudan) raises the question of the nature and origin of these objects. The position in which the objects were found in relation to the human skeleton suggested a pathological condition affecting the individual, possibly urinary bladder, kidney stones or gallstones. To solve this issue, a multi-analytical approach, consisting of tomographic, microstructural and compositional analyses, was therefore performed. Based on their microstructure and mineralogical composition, consisting of hydroxylapatite and whitlockite, the investigated stones were identified as primary (endogenous) prostatic calculi. In addition, the occurrence of bacterial imprints also indicates on-going infectious processes in the individual. This discovery of the earliest known case of lithiasis extends the appearance of prostatic stones into the Late Pleistocene/Early Holocene, a disease which therefore can no longer be considered exclusive to the modern era, but which also affected prehistoric individuals, whose lifestyle and diet were significantly different to our own.