Characterisation of selenium and tellurium nanoparticles produced by Aureobasidium pullulans using a multi-method approach
Introduction
Selenium (Se) and tellurium (Te) are metalloid elements that belong to the chalcogen group in Group 16 of the Periodic table. Neither selenium or tellurium are extracted as primary ores; but are recovered as by-products during the processing of base metal ores such as Cu, Pb, Bi, Fe and other metals [1]. They have an extremely low crustal abundance (Se 0.05 – 0.09 mg kg−1, Te 0.02 mg kg−1) and have been classified amongst the ‘critical’ elements due to a potential risk in their security of supply [2], [3], [4]. Both selenium and tellurium are of economic interest because of their applications in advanced technologies such as photovoltaic cells for solar energy, and thin film technologies [5], [6], [7]. To improve their supply, new methods of extraction have been investigated including chemical reduction [8,9], electrochemical processes [10,11], and microbial biorecovery [12], [13], [14]. Microbial biorecovery of Se and Te in their elemental forms may offer an environmentally sustainable, relatively low-cost method for their production but can be challenged by the low yield and tedious purification steps to obtain sufficient amounts. [15], [16], [17], [18]. Various species of bacteria and fungi have been investigated for the intracellular and extracellular biosynthesis of Se and Te NPs, as a means to biotransform oxyanions of both elements to less toxic forms; and their exploitation in environmental, industrial and medical applications [15], [16], [17], [18]. Se and Te NPs biosynthesised by fungi and bacteria have shown average diameters of 60–80 nm and 221 nm respectively, and may be associated with lipid, carbohydrate and/or protein on the surface of the produced Se and Te NPs [18,19]. Prior to any potential industrial application, there is a need to characterise biosynthesised NPs to determine their size, composition, distribution, and dispersibility. Various established kinds of metrological techniques are already in use for NP characterisation based on their quantification, separation and characterisation with each technique providing a specific kind of information [20]. In this study, separation of the NPs was achieved with asymmetric flow field-flow fractionation (AF4), an analytical technique which sequentially separates NPs in a thin channel in order of increasing particle size under the influence of a perpendicular crossflow [21], [22], [23], [24]. One advantage of the AF4 technique is its versatility to be simultaneously coupled with multiple detectors, with multi-angle light scattering (MALS) and inductively coupled plasma mass spectrometry (ICP-MS) detectors providing particle size data and element-specific detection respectively, and its ability to detect nano- and microparticles in the same run. For single particle inductively coupled plasma mass spectrometry (spICP-MS), sufficiently diluted suspensions are detected above a background signal and counted in fast time resolved analysis (TRA) mode using ultrafast integration times [25,26]. The introduction of a single NP into the plasma generates a packet of ions creating a pulse signal when it reaches the detector. The detector pulse is correlated to the total mass per particle and subsequently correlated to the particle size [27], [28], [29], [30].
The main advantages of spICP-MS are that it employs the power of an ICP plasma to completely destroy any biological matrix material; it uses very small sample volumes with sufficient dilution to ensure high sensitivity of signals above background and to avoid the detection of two particles in one measurement event. In this study, we demonstrated the need for complementary analytical techniques to characterise biogenic nanoparticles. In addition to the coupled AF4-UV-MALS-ICP-MS and spICP-MS analytical tools, the conventional and relatively straightforward techniques such as transmission electron microscopy (TEM) and dynamic light scattering (DLS) were employed to provide an enhanced analytical perspective for a better characterisation of biogenic Se/Te-NPs produced by the polymorphic fungus Aureobasidium pullulans when exposed to selenite and tellurite. Many fungi are capable of the reductive transformation of metalloid oxyanions, including Se and Te, to elemental forms which provides a biological system applicable to bioremediation and/or element biorecovery [13,31].
Section snippets
Biosynthesis of NPs by A. pullulans
Se and Te NPs biosynthesis by A. pullulans was performed following the protocol described by Liang et al. [18] All reagents and chemicals used were of analytical grade or better with all volumes measured gravimetrically. Liquid cultures were prepared in 250-mL Erlenmeyer conical flasks containing 100 mL nutrient medium on an orbital shaking incubator (Infors Multitron Standard, Rittergasse, Switzerland) at 125 rpm, 25 °C in the dark. AP1 agar medium was used as the growth medium with previously
Total biogenic SeNPs and TeNPs generated by A. pullulans
After 30-day incubation, nanoparticles were harvested from supernatants of A. pullulans after growth in AP1 medium amended with 1 mM Na2SeO3 (79 mg Se L−1)/Na2TeO3 (128 mg Te L−1). Total yields of SeNPs and TeNPs harvested from the supernatant were 81.6 ± 1.4 mg L−1, 21.5 ± 0.1 mg L−1 (n = 3) respectively. Previous research has already demonstrated that SeNPs and TeNPs can be located both intracellularly and extracellularly [18,33]. This work focused on the extracellular SeNPs and TeNPs
Conclusions
Selenium and tellurium nanoparticles were harvested from A. pullulans grown in liquid medium amended with sodium selenite and sodium tellurite for 30 days and characterised using different techniques. Based on the results presented in this work, the NPs have been separated with field-flow fractionation and characterised (particle size, shape, morphology, and distribution) with mass spectrometry and optical techniques. Various flow and sample parameters influence an optimal AF4 separation, and
Authorship contribution statement
Kenneth C Nwoko: Writing-original draft, review and editing, sample preparation, experimental design and instrumental analysis (AF4-UV-MALS-ICP-MS/MS), DLS, TEM.
Xinjin Liang: Growth and culture experiments, review, editing.
Magali AMJ Perez: Sample preparation and instrumental analysis (spICP-MS, TEM).
Eva Krupp: Supervision.
Geoffrey Michael Gadd: Conceptualization, supervision, funding acquisition, review, editing.
Jörg Feldmann: Conceptualization, supervision, funding acquisition, review,
Declaration of Competing Interest
None.
Acknowledgements
KCN acknowledges the receipt of research funding received from the University of Aberdeen's Elphinstone PhD studentship award (RG13451), and the Niger-Delta Development Commission (NDDC) Postgraduate Foreign Scholarship (NDDC/DEHSS/2015PGFS/IMS/09). JF and KCN gratefully acknowledge Postnova Analytics, UK for loaning of the AF4-UV-MALS system and technical support; and the Microscopy and Histology Facility at the Institute of Medical Sciences (IMS) Foresterhill, University of Aberdeen for TEM
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