This study presents the fabrication of beta-type Ti-28Nb-35.4Zr (TNZ) and graphene nanoplatelet (GNP)-reinforced TNZ composites (0.1 and 0.2 wt.%) via direct metal deposition (DMD) for orthopedic implant applications. The effects of GNP addition on microstructure, mechanical strength, tribological behavior, corrosion resistance, and cytocompatibility were systematically evaluated. All DMD-processed samples primarily exhibited beta-phase, while GNP incorporation promoted alpha '' martensite formation due to oxygen uptake during processing. The 0.1 wt.% GNP addition refined the grain structure, enhancing compressive yield strength by 16% (863 MPa) and maintaining high ductility with a maximum strain >55%. Hardness increased by 17% for the TNZ-0.1GNP composite, which had the lowest wear volume (0.03 & micro;m(3)) and minimal debris formation. Electrochemical testing in Hanks' balanced salt solution revealed enhanced corrosion resistance for TNZ-0.1GNP with a corrosion rate of 3.88 & micro;m y(-1) due to synergistic passivation and barrier effects. The reduced water contact angle (74 degrees +/- 5 degrees) indicated improved hydrophilicity and favorable surface energy for biointeractions. Furthermore, human osteoblast-like SaOS2 cells showed thriving adhesion, proliferation, and complete surface coverage on TNZ-0.1GNP after 7 days of culture. Overall, DMD-processed TNZ-0.1GNP composite exhibited promising mechanical, wear resistance, corrosion and biocompatible performance as load-bearing orthopedic implant materials.

Additive manufacturing (AM) of polymeric materials is rapidly transforming the biomedical field by enabling the fabrication of patient-specific, anatomically complex structures with precise control over internal architecture. Polymers are especially attractive for AM of biomedical devices due to their cost-effectiveness, abundance, low density, and tunable mechanical and degradation properties, supporting diverse applications in soft and hard tissue engineering, microfluidics, and drug delivery. However, many medical-grade polymers interact poorly with mammalian cells and tissues due to the lack of bioactive surface functional groups, which can hinder their performance in biomedical applications that rely on cell-material interactions such as tissue regeneration. This review systematically surveys physical, chemical, and biomimetic surface modification techniques for AM-compatible medical polymers to improve biomedical applications and targeted functionalities. While much attention has been paid in the literature to surface modification in bone tissue engineering, functional coatings incorporating bioactive molecules and nanoparticles further provide antibacterial, anti-inflammatory, and pro-regenerative functions. A major emphasis of this review is the synergy between AM and surface engineering, enabling simultaneous optimization of internal architecture and surface bioactivitycapabilities fundamentally unattainable by conventional manufacturing techniques. Finally, challenges such as sterilization compatibility and long-term stability of surface modifications are discussed as key to clinical translation.

Indigenous foods are increasingly being incorporated into contemporary diets, yet many lack dietary safety data and documented histories of use. This provides uncertainty for suppliers and poses challenges for food regulators assessing traditionally used foods entering modern markets. We present a case study of an Australian native grain (ganalay; Astrebla lappacea), traditionally used by the Gamilaraay Peoples, and compare it to commonly consumed wheat using in vitro bioassays and chemical analyses. The native grain exhibited no greater cytotoxicity, oxidative stress, or inflammatory signalling than wheat in human monocyte and hepatocyte bioassays. Chemical analysis showed contaminant levels were below tolerable limits, and no chemical classes of concern were detected. These findings suggest that the tested grain sample poses no greater risk than wheat within the in vitro and chemical contexts examined. This study demonstrates a practicable, low-cost safety assessment framework that supports the inclusion of traditionally used Indigenous foods within contemporary diets, while enabling Indigenous-led development of culturally significant species. The framework provides an initial evidence base that can be interpreted alongside cultural knowledge and histories of use, offering an accessible pathway for communities seeking to understand the safety of their foods, regardless of whether commercialisation is an intended outcome.

The addition of zinc oxide (ZnO) nanofillers to 3D printable poly(lactic acid) (PLA) filaments for material extrusion (MEX) additive manufacturing (fused filament fabrication, FFF, a.k.a. fused deposition modelling, FDM) has the potential to enable the fabrication of biomedical devices with embedded antibacterial functionality. This work investigates the biological properties, mainly the biodegradability, antibacterial activity, and cytotoxicity of 3D printed PLA-ZnO nanocomposites containing between 1 wt% to 5 wt% of either untreated or silane-treated filler. This study demonstrated that the concentration and surface properties of the filler control the matrix degradation rate, which directly influences the release rate of ZnO and Zn2+, which in turn governs the antibacterial properties of the nanocomposites. All nanocomposites showed excellent antibacterial properties (> 99% reduction in bacteria) against both gram-positive (Staphylococcus aureus) and gram-negative (Escherichia coli) strains. Potential cytotoxic effects against human immune THP-1 cells were only evident at the highest filler loading (5 wt%), whereas nanocomposites with < 5 wt% filler loading were non-cytotoxic after 7 days of exposure. The 3D printed PLA-ZnO nanocomposites produced in this study show potential for use in clinical settings, with nanocomposites having filler loadings of < 2 wt% being the most appropriate candidates due to their excellent antibacterial properties while showing comparable biocompatibility to pristine PLA.

Pure zinc (Zn) and Zn alloys exhibit poor strength and weak ductility in their as-cast state, limiting their use in bone implants. Consequently, various alloying elements, such as rare earth elements (REE), and thermomechanical processes such as hot rolling (HR) have been explored to enrich their properties. This study reports the effect of gadolinium (Gd) (0.2-0.6 wt%) on the microstructure, mechanical properties, corrosion resistance, and biocompatibility of a Zn-3Cu-0.2Li (ZCL) alloy prepared via casting and HR. The results showed that Gd addition generated a new secondary phase of GdZn12 and HR refined the grain size, which together significantly enhanced the strength and the ductility of the ZCL-xGd alloys. The HR ZCL-0.6Gd alloy displayed a tensile yield strength (CTYS) of 245 MPa, an ultimate tensile strength (CUTS) of 361 MPa, and an elongation (epsilon) of 33 %. Overall, the HR ZCL-xGd alloys demonstrated superior tensile properties with a CTYS in the range of 228-248 MPa, CUTS in the range of 341-371 MPa, and epsilon in the range of 23-33 %, meeting orthopedic implant requirements. The resistance to corrosion of HR ZCL-xGd alloys decreased as a function of Gd level, with the HR ZCL-0.6Gd showing the highest degradation rate (60.5 mu m/y in Hanks' solution for 30 days). Cytocompatibility tests indicated an insignificant level of toxicity in relation to osteoblast-like SaOS2 cells. The HR ZCL-0.6Gd alloy exhibited optimal strength, biodegradability, and biocompatibility for bone implants.

Poly(lactic acid)-zinc oxide (PLA-ZnO) nanocomposites for fused filament fabrication have potential applications in the biomedical field as they combine the bio-compatibility of PLA with the antibacterial properties of ZnO. This work investigates the effects of masterbatch mixing strategy, ZnO concentration and ZnO surface treatment (silanisation) on the printability and the mechanical performance of the nanocomposites as a pre-requirement to the wider uptake of these materials. The results showed that the printability decreased as the filler loading increased. However, the surface treatment of the ZnO powder enhanced the matrix-filler interfacial interactions and reduced the thermal degradation of PLA. This ameliorated the printability and the tensile properties of the nanocomposites filled with up to 5 wt.% of ZnO. Moreover, despite the additional thermal treatment, melt-mixing prevented the degradative effect induced by the solvent used for solvent mixing. Future work will focus on assessing the antibacterial properties of the nanocomposite FFF parts.

Summary Climate change is threatening our current food systems and countries around the world are seeking alternative food sources to safeguard against potential food insecurity and nutrient deficiency concerns. Various traditional foods used by Aboriginal groups of Australia have previously been shown to be highly nutritious, contain beneficial functional properties and can thrive in the harsh climatic conditions found across Australia. As interest in these traditional foods grows, Aboriginal groups are looking at developing a range of foods for modern markets. However, for many of these foods, there is still little known about their nutritional and functional properties. In this study, we report the nutrition composition and antioxidant properties of a native Australian grain species that is currently being developed for commercial markets by a collective of Aboriginal Traditional Custodians and compare its properties to a commonly consumed wheat variety. Compared with wheat, the native grain species contained 2‐fold greater protein and total fats, and higher levels of essential minerals and trace elements, including 8‐fold iron levels and >2.5‐fold calcium, magnesium, zinc, copper and manganese levels. Functionally, the native grain contained 2.4‐fold greater polyphenol content and displayed greater antioxidant potential in exposed human monocyte cultures. Importantly, the native grain was also found to contain a very low gluten content. Altogether, this shows that the native grain is nutritionally and functionally superior to commonly consumed wheat for the measured parameters and could potentially serve as a grain alternative or be used to fortify current wheat‐based and gluten‐free products.

Hemp is low-tetrahydrocannabinol (THC) Cannabis sativa that is cultivated globally for food and fibre. Hemp seeds and seed-oil are popularly consumed individually or utilised within cooking, due to their flavour, nutrition, and functional food benefits. The functional food properties of hemp seed include its beneficial fatty acid (FA) profile and antioxidant activity, which are associated with reduced inflammation that benefits chronic conditions including cardiovascular disease and cancer. Maximising these functional food properties can offer extensive health benefits. Biostimulants - such as molasses - are fertiliser additives that are associated with improvements to plant growth, yield, and biochemical composition; however, they have been scarcely applied to hemp. Furthermore, due to the complex composition of biostimulants, understanding their modes of action poses a significant challenge, impeding the optimization of their usage and the realization of associated benefits. Accordingly, this study aimed to characterise the impact of molasses on the functional food properties of hemp seeds and explore biostimulant-induced biochemical changes as indicators of modes of effect. Although molasses treatment did not significantly alter the FA profile (p = 0.960 for omega-6:omega-3 FA ratio), the antioxidant capacity (as measured by ABTS) was significantly increased (3.8-fold increase, p = 0.008). Metabolite profiling and statistical modelling indicated that this change was likely associated with increases in several lipophilic antioxidant metabolites, including hydroxycinnamic acid amides and cannabisins. Comparison of root, leaf, seed, and sugar leaf tissue changes in phytohormones and metabolites indicated that the growth of hemp with molasses was predominantly associated with the induction of the plant's endogenous stress response within the roots. These results indicate that the biostimulant molasses is a beneficial fertiliser additive for enhancing the functional food (antioxidant) potential of hemp seeds, thereby improving the health-benefits imparted to consumers.

As interest in Australian native products continues to grow worldwide, Aboriginal and Torres Strait Islander peoples (First Peoples) are striving to be industry leaders in the production of their traditional foods that are being developed for commercial markets. To successfully gain market approval both within Australia and globally, food regulatory authorities require at least a documented history of safe use to indicate dietary safety. Moreover, many countries also require compositional analysis and safety data to further support their safe human consumption. However, safety data are lacking for many of these traditional food items and the history that surrounds their safe use has rarely been recorded in written form, but rather passed on through cultural practices and language. This review evaluates the suitability of current frameworks for assessing the dietary safety of traditional foods and highlights the food-safety regulatory hurdles currently felt by First Peoples and their businesses attempting to enter the Australian native foods industry. These issues also extend to the requirements of food regulatory authorities around the world, when assessing the market eligibility of traditional food items. Potential solutions to these problems are discussed, including new proposed processes that can be incorporated into the current food regulatory frameworks. Importantly, these proposed processes would allow the dietary risk assessment of traditional foods to be completed in a manner that better accommodates the stories, traditional knowledge and interests of First Peoples, while also meeting the safety data requirements set out by regulatory bodies both within Australia and around the world.

In this study, new beta Ti-28Nb-35.4Zr (hereafter denoted TNZ) and multi-walled carbon nanotubes (MWCNTs; 0.1 wt.%) reinforced TNZ composite (hereafter denoted TNZCNT) were manufactured for bone implant applications via direct metal deposition (DMD). The effect of MWCNTs addition was systematically investigated on the microstructure and resultant mechanical, nano-tribological, and biocompatibility properties of TNZ. Results indicated that the microstructures of TNZ and TNZCNT composite were primarily composed of beta along with localized alpha '' martensite phases. TNZ and TNZCNT composite exhibited average compressive yield strengths of 706 MPa and 833 MPa, respectively, with outstanding plastic deformation ability (>55%) without forming cracks and fractured surfaces under high compressive loads. Average nanohardness of TNZ and TNZCNT com-posite was measured as 2.9 GPa and 3.4 GPa, respectively. Compared to average wear volume of TNZ counterpart (0.19 mu m3), nano-tribological tests also revealed higher resistance of TNZCNT composite to wear (0.07 mu m3) owing to its higher hardness. MTS assay revealed that both TNZ and TNZCNT composite exhibit high viabilities of SaOS2 cells measured as 97.6% and 107.6%, respectively, after 7 d of cell culturing. Moreover, TNZCNT composite exhibited thriving adhesion and spreading of SaOS2 cells showing their growth and proliferation on its surface after cell culture for 1 and 7 d, demonstrating its extraordinary biocompatibility. Overall, owing to their appropriate mechanical, nano-tribological, and biocompatibility properties, the DMD-manufactured TNZ and TNZCNT composite displayed promising potential to be utilized as a candidate material for load-bearing implant applications.

Zinc (Zn)‐based materials reveal inadequate mechanical properties as orthopedic biodegradable implantable materials, which limits their biomedical applications. Herein, Zn– x Mg (magnesium) composites ( x = 0.5, 1.0, 1.5, and 2.0 wt%) reinforced with 0.2 wt% graphene nanoplatelets (GNP) are obtained via powder metallurgy and hot‐pressing sintering. The addition of 0.5–2.0 wt% Mg into Zn–0.2 wt% GNP composite resulted in the formation of Mg 2 Zn 11 and MgZn 2 phases without any additional intermetallic carbide phases. The hot‐pressing sintered (HPS) Zn–0.5Mg–0.2GNP composite exhibits a compressive yield strength of 169 ± 18 MPa, an ultimate compressive strength of 270 ± 39 MPa, a compressive strain of 17 ± 6%, and a microhardness of 86 ± 2 HV. Electrochemical corrosion testing reveals that corrosion resistance of HPS Zn– x Mg–0.2GNP composites decreases with increasing Mg content, while immersion tests in Hanks’ balanced salt solution for 30 d indicate that the degradation rate increases from 0.032 to 0.141 mm y −1 by increasing Mg content from 0 to 2.0 wt%. In vitro cytotoxicity assessments using SaOS2 human osteoblast cells show >90% viability following exposure over 5 d to 12.5% extract concentrations of all HPS composites. The HPS Zn–0.5Mg–0.2GNP composite exhibits the appropriate material properties for biodegradable bone‐implant applications.

Zinc (Zn)-based alloys have emerged as promising new biodegradable materials owing to their moderate corrosion rates and potential biological functionalities. Nevertheless, as-cast pure Zn and its alloys possess low mechanical strength, poor ductility and low hardness, which hinders their biomedical application. In this study, biodegradable Zn-3Cu-0.4Li (ZCL) and ZCL-xSc (x = 0.20, 0.35 and 0.55 wt.%) alloys were fabricated by casting and further hot-rolled (HR), with the aim of acquiring satisfactory mechanical, corrosion and biocompatibility properties for orthopedic implant applications. Results indicated that the mechanical properties of HR ZCL and ZCL-xSc alloys were significantly improved compared to the as-cast alloys. The HR ZCL-0.20Sc alloy showed the best combination of mechanical properties: yield strength of 277 MPa; ultimate tensile strength of 337 MPa; elongation of 40%; and a microhardness of 113 HV. The nanohardness of HR ZCL-xSc alloys increased from 1.64 to 2.43 GPa with increasing Sc content from 0 to 0.55 wt.%. Degradation rates of HR ZCL-xSc alloys gradually increased with increasing Sc content, with the HR ZCL-0.55Sc alloy showing the lowest corrosion resistance and the highest degradation rate of 50.1 mu m/y after immersion in Hanks' balanced salt solution for 30 d. In vitro cytocompatibility assessment using human osteoblast-like SaOS2 cells showed high cell viabilities after 1 d of exposure to undiluted extracts of HR ZCL and ZCL-xSc alloys, and also after 5 d with 50% diluted extracts. Overall, the HR ZCL-0.20Sc alloy has great potential as a biodegradable bone-implant material, due to excellent mechanical, satisfactory corrosion and good biocompatibility properties.

This study systematically investigated the effect of equal channel angular pressing (ECAP) on the microstructure, mechanical, corrosion, nano-tribological properties and biocompatibility of a newly developed β Ti-28Nb-35.4Zr (hereafter denoted TNZ) alloy. Results indicated that ECAP of the β TNZ alloy refined its microstructure by forming ultrafine grains without causing stress-induced phase transformation, leading to formation of a single β phase. The ECAP-processed TNZ alloy exhibited a compressive yield strength of 960 MPa, and high plastic deformation capacity without fracturing under compression loads. Potentiodynamic polarization tests revealed the higher tendency of ECAP-processed TNZ alloys to form passive oxide films on its surface, which exhibited a lower corrosion rate (0.44±0.07 µm/y) in Hanks' balanced salt solution compared to its as-cast counterpart (0.71±0.10 µm/y). Nanotribological testing also revealed higher resistance of the ECAP-processed TNZ alloy to abrasion, wear and scratching, when compared to its as-cast counterpart. Cytocompatibility and cell adhesion assessments of the ECAP-processed TNZ alloys showed a high viability (111%) of human osteoblast-like SaOS2 cells after 7 d of culturing. Moreover, the ECAP-processed TNZ alloy promoted adhesion and spreading of SaOS2 cells, which exhibited growth and proliferation on alloy surfaces. In summary, significantly enhanced mechanical, corrosion, and biological properties of ECAP-processed TNZ alloy advocate its suitability for load-bearing implant applications. STATEMENT OF SIGNIFICANCE: Equal channel angular pressing (ECAP) provides a unique combination of enhanced mechanical and functional properties of materials by optimizing their microstructures and phase transformations. This study investigated the mechanical, nano-tribological, corrosion, and biocompatibility properties of a newly developed β Ti-28Nb-35.4Zr (TNZ) alloy processed via ECAP. Our findings indicated that ECAP of the β TNZ alloy refined its microstructure by forming ultrafine grains without causing stress-induced phase transformation. Compared to its as-cast counterpart, ECAP-processed TNZ exhibited significantly enhanced compressive yield strength, plastic deformation capacity, hardness, wear, and corrosion properties. Moreover, in vitro cytocompatibility and cell adhesion studies revealed high cellular viabilities, growth and proliferation of osteoblast-like SaOS2 cells on the ECAP-processed TNZ alloy.

Maternal diet is critical for offspring development and long-term health. Here we investigated the effects of a poor maternal diet pre-conception and during pregnancy on metabolic outcomes and the developing hypothalamus in male and female offspring at birth. We hypothesised that offspring born to dams fed a diet high in fat and sugar (HFSD) peri-pregnancy will have disrupted metabolic outcomes. We also determined if these HFSD-related effects could be reversed by a shift to a healthier diet post-conception, in particular to a diet high in omega-3 polyunsaturated fatty acids (ω3 PUFAs), since ω3 PUFAs are considered essential for normal neurodevelopment. Unexpectedly, our data show that there are minimal negative effects of maternal HFSD on newborn pups. On the other hand, consumption of an ω3-replete diet during pregnancy altered several developmental parameters. As such, pups born to high-ω3-fed dams weighed less for their length, had reduced circulating leptin, and also displayed sex-specific disruption in the expression of hypothalamic neuropeptides. Collectively, our study shows that maternal intake of a diet rich in ω3 PUFAs during pregnancy may be detrimental for some metabolic developmental outcomes in the offspring. These data indicate the importance of a balanced dietary intake in pregnancy and highlight the need for further research into the impact of maternal ω3 intake on offspring development and long-term health.

Agonism of the G protein-coupled bile acid receptor "Takeda G-protein receptor 5" (TGR5) aids in attenuating cholesterol accumulation due to atherosclerotic progression. Although mammalian bile compounds can activate TGR5, they are generally weak agonists, and more effective compounds need to be identified. In this study, two marine bile compounds (5β-scymnol and its sulfate) were compared with mammalian bile compounds deoxycholic acid (DCA) and ursodeoxycholic acid (UDCA) using an in vitro model of TGR5 agonism. The response profiles of human embryonic kidney 293 cells (HEK293) transfected to overexpress TGR5 (HEK293-TGR5) and incubated with subcytotoxic concentrations of test compounds were compared to nontransfected HEK293 control cells using the specific calcium-binding fluorophore Fura-2AM to measure intracellular calcium [Ca2+]i release. Scymnol and scymnol sulfate caused a sustained increase in [Ca2+]i within TGR5 cells only, which was abolished by a specific inhibitor for Gαq protein (UBO-QIC). Sustained increases in [Ca2+]i were seen in both cell types with DCA exposure; this was unaffected by UBO-QIC, indicating that TGR5 activation was not involved. Exposure to UDCA did not alter [Ca2+]i, suggesting a lack of TGR5 bioactivity. These findings demonstrated that both scymnol and scymnol sulfate are novel agonists of TGR5 receptors, showing therapeutic potential for treating atherosclerosis.

BACKGROUND:Cholesterol crystallization within an atherosclerotic plaque significantly contributes to the acceleration of plaque rupture - a problematic event due to the current lack of specific treatments to prevent such formations. Modelling this pathogenic process is also difficult due to the lack of suitable experimental models that enable quantitative analysis of crystal formation and bioactivity screening of potential therapeutic compounds.AIM:To develop an in vitro human cell model of cholesterol crystallization combined with an imaging system that incorporates both quantitative analysis and real-time continuous imaging of cholesterol crystal formation.METHODS AND RESULTS:An enhanced in vitro model of cholesterol crystallization was developed through the use of acetylated low-density lipoprotein (AcLDL) and 7-ketocholesterol as agents of foam cell induction within a human THP-1 monocytic cell line. Advanced confocal and polarizing microscopies were incorporated into the model so as to allow for quantitation of cholesterol crystallization, with the lipid-loaded group producing significantly greater numbers of cholesterol crystals than the untreated group. The utility of this system was also demonstrated by investigating the effects of the cholesterol-lowering drug lovastatin and therapeutic bile compound ursodeoxycholic acid (UDCA), showing that these drugs influence different aspects of cholesterol crystal formation.CONCLUSIONS:The in vitro human THP-1 monocyte model of cholesterol crystallization provides an effective and efficient means of quantitating cholesterol crystallization in the pre-clinical stage of research. The model also allows for the screening of potentially therapeutic compounds that may be used in attenuating or preventing cholesterol crystallization.

The authors would like to correct a typographical error in their recently published paper [...]

Magnesium (Mg) and its alloys are considered promising biodegradable implant materials because of their strength and natural degradation in the human body. However, the high corrosion rate of pure Mg in the physiological environment leads to rapid degradation before adequate bone healing. This mismatch between bone healing and the degradation of Mg implants supports the development of new Mg alloys with the addition of other suitable alloying elements in order to achieve simultaneously high corrosion resistance and desirable mechanical properties. This study systematically investigates the microstructure, mechanical properties, corrosion behavior, and biocompatibility of Mg-based alloys with the addition of different concentrations of scandium (Sc), i.e., Mg-0.6Zr-0.5Sr-xSc (x = 0.5, 1, 2, 3 wt.%). Results indicated that high concentration of Sc in strontium (Sr)-containing Mg alloys can alter their microstructures by suppressing the intermetallic phases along the grain boundaries and improve the corrosion resistance by forming chemically stable Sc oxide layers on the surfaces of the Mg alloys. Cytotoxicity assessment revealed that the Sc containing Mg alloys did not significantly alter the viability of human osteoblast-like SaOS2 cells. This study highlights the advantages of using Sc as an alloying element to simultaneously tune Mg alloys with higher strength and slower degradation. Statement of significance Rare earth elements such as scandium (Sc) with both a high solid-solubility and strong affinity towards oxygen can improve the mechanical and corrosion properties of magnesium (Mg) alloys. However, the feasibility of Sc-containing Mg alloys as biodegradable implant materials is scarcely reported. This study investigates the effects of different Sc concentrations on the mechanical, corrosion, and biocompatibility properties of Mg-Zr-Sr-Sc alloys. Our findings indicated that the addition of Sc significantly improves the mechanical and corrosion properties of Mg-Zr-Sr alloys. Moreover, in vitro cytotoxicity assessment of the Mg-Zr-Sr-Sc alloys did not show any adverse effects on the viability of osteoblast-like cells. (C) 2019 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The effectiveness of decontamination procedures used for the removal of external drug contamination in forensic hair analysis is an ongoing debate. This investigation evaluated wash methods complying with Society of Hair Testing (SoHT) guidelines and their capacity to remove cocaine (COC) and methamphetamine (MA) from artificially contaminated hair. The most effective decontamination method was determined using a systematic approach, involving (1) an initial washing solvent screen, (2) optimization of wash duration, (3) comparison of sequential wash methods, and (4) reanalysis of clinical hair samples. For analysis, hair was subjected to micro-pulverized methanolic extraction prior to quantitation by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Methanol (MeOH) and 0.1 M phosphate buffer (pH 6) were the most effective organic and aqueous solvents, respectively, removing 28%-38% of COC and 16%-31% of MA. Wash durations longer than 30-60 minutes did not remove additional amounts, and a more efficient sequential wash method was subsequently developed. Despite this, the interpretation of reportable results relative to the SoHT cut-off levels was unchanged for most clinical hair samples reanalyzed after washing by agitation for 30 minutes with MeOH. These findings highlight the inability of decontamination solvents to completely remove external COC and MA contamination from hair, including wash methods adhering to SoHT guidelines.