Green Analytical Chemistry (GAC) metrics include a variety of criteria, such as the regent amounts and toxicity, energy consumption, generated waste, among others. The analytical greenness metric (AGREE) and its variant for sample preparation (AGREEprep) cover different aspects that contribute to the environmental sustainability of sample preparation. White Analytical Chemistry (WAC) considers not only environmental aspects but also analytical and practical aspects with a holistic vision based on a Red-Green-Blue color model. A case study is presented to assess the green and white profile of a method based on ultrasound-assisted extraction and determination of Mn and Fe in beef using microwave-induced plasma atomic emission spectroscopy (MP AES). The method was validated and resulted simple, fast without external heating using diluted acids. It was concluded that we should think in green sample preparation with the AGREEprep tool, as well as in white holistic assessments (WAC) as both constitute complementary tools.

Metabolomics studies are becoming increasingly common for understanding how plant metabolism responds to changes in environmental conditions, genetic manipulations and treatments. Despite the recent advances in metabolomics workflow, the sample preparation process still limits the high-throughput analysis in large-scale studies. Here, we present a highly flexible robotic system that integrates liquid handling, sonication, centrifugation, solvent evaporation and sample transfer processed in 96-well plates to automatize the metabolite extraction from leaf samples. We transferred an established manual extraction protocol performed to a robotic system, and with this, we show the optimization steps required to improve reproducibility and obtain comparable results in terms of extraction efficiency and accuracy. We then tested the robotic system to analyze the metabolomes of wild-type and four transgenic silver birch (Betula pendula Roth) lines under unstressed conditions. Birch trees were engineered to overexpress the poplar (Populus × canescens) isoprene synthase and to emit various amounts of isoprene. By fitting the different isoprene emission capacities of the transgenic trees with their leaf metabolomes, we observed an isoprene-dependent upregulation of some flavonoids and other secondary metabolites as well as carbohydrates, amino acid and lipid metabolites. By contrast, the disaccharide sucrose was found to be strongly negatively correlated to isoprene emission. The presented study illustrates the power of integrating robotics to increase the sample throughput, reduce human errors and labor time, and to ensure a fully controlled, monitored and standardized sample preparation procedure. Due to its modular and flexible structure, the robotic system can be easily adapted to other extraction protocols for the analysis of various tissues or plant species to achieve high-throughput metabolomics in plant research.

Reverse-phase solid-phase extraction (SPE) is regularly used for separating and purifying food-derived oligosaccharides and peptides prior to liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. However, the diversity in physicochemical properties of peptides may prevent the complete separation of the two types of analytes. Peptides present in the oligosaccharide fraction not only interfere with glycomics analysis but also escape peptidomics analysis. This work evaluated different SPE approaches for improving LC-MS/MS analysis of both oligosaccharides and peptides through testing on peptide standards and a food sample of commercial interest (proteolyzed almond extract). Compared with conventional reverse-phase SPE, mixed-mode SPE (reverse-phase/strong cation exchange) was more effective in retaining small/hydrophilic peptides and capturing them in the high-organic fraction and thus allowed the identification of more oligosaccharides and dipeptides in the proteolyzed almond extract, with satisfactory MS/MS confirmation. Overall, mixed-mode SPE emerged as the ideal method for simultaneously improving the identification of food-derived oligosaccharides and small peptides using LC-MS/MS analysis.

BACKGROUND: Nuclear magnetic resonance (NMR)-based metabolic profiling has been widely used in food and plant sciences. Despite its simplicity and inherent reproducibility, the determination of the appropriate pre-processing procedures greatly affects the obtained metabolic profile.
OBJECTIVE: The current study represents a detailed guide of use for untargeted NMR-based metabolic profiling of table olives (Olea europaea L.).
METHODS: Greek Kalamon table olives from different geographical origins were selected as reference materials. Differently treated samples were extracted using different solvents and/or solvent systems. Chemical profiles were evaluated with high-performance thin layer chromatography (HPTLC). Different deuterated solvents and sample concentrations were evaluated for the recording of optimal quality spectra.
RESULTS: The methanol extract of freeze-dried table olives was found to contain the most representative secondary metabolites, in higher concentrations, as well. The optimal deuterated solvent for the NMR analysis was methanol-d4 , while final sample concentration should be within the range of 10 to 15 mg/mL. Multivariate data analysis was also used to estimate and confirm the variation and clustering caused by different characteristics of the samples.
CONCLUSIONS: Results of the present study make evident the necessity for thorough planning and method development prior to any extensive metabolomic study based on NMR spectroscopy. Pre-processing and sample preparation stages seemed to greatly affect the metabolic profile and spectral quality in the case of table olives, which by extrapolation could apply to other food commodities. Nevertheless, the nature of the samples must be fully described in general, in order to proceed to solid conclusions.

OBJECTIVE: The goal of this study was to develop sample preparation method and validate the HPLC method for precise determination of paclitaxel (Ptx) in PLGA submicron particles conjugated with protein vector molecule.
METHODS: Ptx loaded PLGA submicron particles were formulated by a single emulsification method. PLGA submicron particles were conjugated with alpha fetoprotein third domain (rAFP3d) via standard carbodiimide technique. The obtained conjugate was analyzed using 1525 binary pump and 2487 UV-VIS detector system (Waters, USA) and Reprosil ODS C-18 analytical column with the dimensions of 150mm×4.6mm ID×5μm (Dr. Maisch GmbH, Germany). Sample preparation method was developed utilizing guard cartridge with С18 stationary phase (Phenomenex, USA). HPLC method was validated according to the international conference on harmonization guidelines.
RESULTS: Efficient sample preparation was achieved using 4% of DMSO pre-dissolution, following by 10min of centrifugation at 4500g. Ptx determination was performed using acetonitrile/0.1% phosphoric acid (50:50 v/v) mobile phase at a flow rate of 1.0mL/min, injection volume of 10μL, and at 227nm. The developed method showed linearity, accuracy and precision in the range from 0.03 to 360μg/mL, with LOD and LOQ values of 0.005 and 0.03μg/mL, respectively. The intra- and inter-day precisions presented RSD values of lower than 2%.
CONCLUSIONS: The validated method was successfully applied to calculate Ptx encapsulation efficacy and drug loading in the developed formulation.

Determining conditions optimal for host growth, maximal protein yield, and lysis buffer composition is of critical importance for the efficient purification of soluble and well-folded recombinant proteins suitable for functional and/or structural studies. Small-scale optimization of conditions for protein production and stability saves time, labor, and costs. Here we describe a protocol for quick protein production and solubility screen using TissueLyser II system from Qiagen enabling simultaneous processing of 96 protein samples, with application to recombinant proteins encompassing two intracellular domains of ethylene-recognizing sensor histidine kinase ETHYLENE RESPONSE1 (ETR1) from Arabidopsis thaliana. We demonstrate that conditions for expression and cell lysis found in our small-scale screen allow successful large-scale production of pure and functional domains of sensor histidine kinase, providing a strategy potentially transferable to other similar catalytic domains.

Solid-phase microextraction (SPME) is an alternative method to dialysis and ultrafiltration for the determination of plasma protein binding (PPB) of drugs. It is particularly advantageous for complicated analytes where standard methods are not applicable. Di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC) is a lead compound of novel thiosemicarbazone anti-cancer drugs, which entered clinical trials in 2016. However, this agent exhibited non-specific binding on filtration membranes and had intrinsic chelation activity, which precluded standard PPB methods. In this study, using a simple and fast procedure, we prepared novel SPME fibers for extraction of DpC based on a metal-free, silicon string support, covered with C18 sorbent. Reproducibility of the preparation process was demonstrated by the percent relative standard deviation (RSD) of ≤ 9.2% of the amount of DpC extracted from PBS by several independently prepared fibers. The SPME procedure was optimized by evaluating extraction and desorption time profiles. Suitability of the optimized protocol was verified by examining reproducibility, linearity, and recovery of DpC extracted from PBS or plasma. All samples extracted by SPME were analyzed using an optimized and validated UHPLC-MS/MS method. The developed procedure was applied to the in vitro determination of PPB of DpC at two clinically relevant concentrations (500 and 1000 ng/mL). These studies showed that DpC is highly bound to plasma proteins (PPB ≥ 88%) and this did not differ significantly between both concentrations tested. This investigation provides novel data in the applicability of SPME for the determination of PPB of chelators, as well as useful information for the clinical development of DpC. Graphical abstract.

This review article is focused on element composition of medicinal plants and herbs as well as their decoctions and infusions determined by atomic spectrometry methods. Considering quality and safety of these herbal beverages, widely practiced in herbalism for medicinal and supplementing purposes, element analysis is important, and quality of its results should not raise any doubts about reliability. Hence, original researches devoted to element analysis of decoctions and infusions of medicinal plants and herbs, published after 2000, have been surveyed in details, particularly focusing on sources of elements in medicinal plants, their availability for the intake during preparation of infusions and decoctions as well as different methodological aspects related to element analysis by atomic spectrometry, including sample pretreatment and preparation before measurements, calibration methods used, and verification of reliability of results.

The standard preparation technique for micro-sized samples is focused ion beam milling, most frequently using Ga+ ions. The main drawbacks are the required processing time and the possibility and risks of ion implantation. In contrast, ultrashort pulsed laser ablation can process any type of material with ideally negligible damage to the surrounding volume and provides 4 to 6 orders of magnitude higher ablation rates than the ion beam technique. In this work, a femtosecond laser was used to prepare wood samples from spruce for mechanical testing at the micrometre level. After optimization of the different laser parameters, tensile and compressive specimens were produced from microtomed radial-tangential and longitudinal-tangential sections. Additionally, laser-processed samples were exposed to an electron beam prior to testing to study possible beam damage. The specimens originating from these different preparation conditions were mechanically tested. Advantages and limitations of the femtosecond laser preparation technique and the deformation and fracture behaviour of the samples are discussed. The results prove that femtosecond laser processing is a fast and precise preparation technique, which enables the fabrication of pristine biological samples with dimensions at the microscale.

Drug product assay is one of several tests required for new drug products to ensure the quality of the product at release and throughout the life cycle of the product. Drug product assay testing is typically performed by preparing a composite sample of multiple dosage units to obtain an assay value representative of the batch. In some cases replicate composite samples may be prepared and the reportable assay value is the average value of all the replicates. In previously published work by Harrington et al. (2014) [5], a sample preparation composite and replicate strategy for assay was developed to provide a systematic approach which accounts for variability due to the analytical method and dosage form with a standard error of the potency assay criteria based on compendia and regulatory requirements. In this work, this sample preparation composite and replicate strategy for assay is applied to several case studies to demonstrate the utility of this approach and its application at various stages of pharmaceutical drug product development.