Facile isolation and analysis of sporopollenin exine from bee pollen.
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ABSTRACT:
SUBMITTER: Hegedus K
PROVIDER: S-EPMC8113464 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Sporopollenin is a physically robust and chemically resilient biopolymer that comprises the outermost layer of pollen walls and is the first line of defense against harsh environmental conditions. The unique physicochemical properties of sporopollenin increasingly motivate the extraction of sporopollenin exine capsules (SECs) from pollen walls as a renewable source of organic microcapsules for encapsulation applications. Despite the wide range of different pollen species with varying sizes and wall thicknesses, faithful extraction of pollen-mimetic SECs has been limited to thick-walled pollen capsules with rigid mechanical properties. There is an unmet need to develop methods for producing SECs from thin-walled pollen capsules which constitute a large fraction of all pollen species and have attractive materials properties such as greater aerosol dispersion. Herein, we report the first successful extraction of inflated SEC microcapsules from a thin-walled pollen species (Zea mays), thereby overcoming traditional challenges with mechanical stability and loss of microstructure. Morphological and compositional characterization of the SECs obtained by the newly developed extraction protocol confirms successful protein removal along with preservation of nanoscale architectural features. Looking forward, there is excellent potential to apply similar strategies across a wide range of unexplored thin-walled pollen species.
| S-EPMC4908411 | biostudies-literature
Project description:Sporopollenin exine capsules (SECs) extracted from Lycopodium clavatum spores are an attractive biomaterial possessing a highly robust structure suitable for microencapsulation strategies. Despite several decades of research into SEC extraction methods, the protocols commonly used for L. clavatum still entail processing with both alkaline and acidolysis steps at temperatures up to 180?°C and lasting up to 7 days. Herein, we demonstrate a significantly streamlined processing regimen, which indicates that much lower temperatures and processing durations can be used without alkaline lysis. By employing CHN elemental analysis, scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), and dynamic image particle analysis (DIPA), the optimum conditions for L. clavatum SEC processing were determined to include 30?hours acidolysis at 70?°C without alkaline lysis. Extending these findings to proof-of-concept encapsulation studies, we further demonstrate that our SECs are able to achieve a loading of 0.170?±?0.01?g BSA per 1?g SECs by vacuum-assisted loading. Taken together, our streamlined processing method and corresponding characterization of SECs provides important insights for the development of applications including drug delivery, cosmetics, personal care products, and foods.
| S-EPMC4730194 | biostudies-literature
Project description:Sporopollenin, the polymer comprising the exine (outer solid shell) of pollen, is recognized as one of the most chemically and mechanically stable naturally occurring organic substances. The elastic modulus of sporopollenin is of great importance to understanding the adhesion, transport and protective functions of pollen grains. In addition, this fundamental mechanical property is of significant interest in using pollen exine as a material for drug delivery, reinforcing fillers, sensors and adhesives. Yet, the literature reports of the elastic modulus of sporopollenin are very limited. We provide the first report of the elastic modulus of sporopollenin from direct indentation of pollen particles of three plant species: ragweed (Ambrosia artemisiifolia), pecan (Carya illinoinensis) and Kentucky bluegrass (Poa pratensis). The modulus was determined with atomic force microscopy by using direct nanomechanical mapping of the pollen shell surface. The moduli were atypically high for non-crystalline organic biomaterials, with average values of 16 ± 2.5 GPa (ragweed), 9.5 ± 2.3 GPa (pecan) and 16 ± 4.0 GPa (Kentucky bluegrass). The amorphous pollen exine has a modulus exceeding known non-crystalline biomaterials, such as lignin (6.7 GPa) and actin (1.8 GPa). In addition to native pollen, we have investigated the effects of exposure to a common preparative base-acid chemical treatment and elevated humidity on the modulus. Base-acid treatment reduced the ragweed modulus by up to 58% and water vapour exposure at 90% relative humidity reduced the modulus by 54% (pecan) and 72% (Kentucky bluegrass). These results are in agreement with recently published estimates of the modulus of base-acid-treated ragweed pollen of 8 GPa from fitting to mechanical properties of ragweed pollen-epoxy composites.
| S-EPMC6170768 | biostudies-literature
Project description:Lycopodium clavatum sporopollenin exine capsules (SpECs) are known to both adsorb and absorb chemicals. The aim of the present work was to determine whether oestradiol (E2) is 'bioavailable' to bioindicator species, either pre-adsorbed to, or in the presence of, SpECs. SpEC uptake was confirmed for Daphnia magna and Dreissena bugensis. E2 levels varied among treatments for Caenorhabditis elegans though there was no relationship to SpEC load. E2 was not detected in D. bugensis tissues. Expression changes of general stress and E2-specific genes were measured. For C. elegans, NHR-14 expression suggested that SpECs modulate E2 impacts, but not general health responses. For D. magna, SpECs alone and with E2 changed Vtg1 and general stress responses. For D. bugensis, SpECS were taken up but no E2 or change in gene expression was detected after exposure to E2 and/or SpECs. The present study is the first to investigate SpECs and bound chemical dynamics.
| S-EPMC8556174 | biostudies-literature
Project description:Tripartite sporopollenin microcapsules prepared from pine pollen (Pinus sylvestris L. and Pinus nigra Arnold) were analysed with respect to the permeability of the different strata of the exine which surround the gametophyte and form the sacci. The sexine at the surface of the sacci is highly permeable for polymer molecules and latex particles with a diameter of up to 200 nm, whereas the nexine covering the gametophyte is impermeable for dextran molecules, with a Stokes' radius > or =4 nm (Dextran T 70), and for the tetravalent anionic dye Evans Blue (Stokes' radius = 1.3 nm). The central capsules obtained by dissolution of the sporoplasts showed strictly membrane-controlled exchange of non-electrolytes, with half-equilibration times in the range of minutes (monosaccharides, oligosaccharides) to hours (dextran molecules with Stokes' radii up to 2.5 nm). The dependence of the permeability coefficients of the nexine for non-electrolytes on Stokes' radius or molecular weight shows that the aqueous pores through the nexine are inhomogeneous with respect to their size, and that most pores are too narrow for free diffusion of sugar molecules. To explain the barrier function of the nexine for Evans Blue, it is assumed that at least the larger pores, which enable slow permeation of dextran molecules, contain negative charges.
| S-EPMC4243658 | biostudies-literature
Project description:Pollen grains are natural microcapsules comprised of the biopolymer sporopollenin. The uniformity and special tridimensional architecture of these sporopollenin structures confer them attractive properties such as high resistance and improved bioadhesion. However, natural pollen can be a source of allergens, hindering its biomedical applicability. Several methods have been developed to remove internal components and allergenic compounds, usually involving long and laborious processes, which often cannot be extended to other pollen types. In this work, we propose an abridged protocol to produce stable and pristine hollow pollen microcapsules, together with a complete physicochemical and morphological characterization of the intermediate and final products. The optimized procedure has been validated for different pollen samples, also producing sporopollenin microcapsules from Matricaria species for the first time. Pollen microcapsules obtained through this protocol presented low protein content (4.4%), preserved ornamented morphology with a nanoporous surface, and low product density (0.14 g/cm3). These features make them interesting candidates from a pharmaceutical perspective due to the versatility of this biomaterial as a drug delivery platform.
| S-EPMC8271440 | biostudies-literature
Project description:Pollen wall consists of several complex layers which form elaborate species-specific patterns. In Arabidopsis, the transcription factor ABORTED MICROSPORE (AMS) is a master regulator of exine formation, and another transcription factor, TRANSPOSABLE ELEMENT SILENCING VIA AT-HOOK (TEK), specifies formation of the nexine layer. However, knowledge regarding the temporal regulatory roles of TEK in pollen wall development is limited. Here, TEK-GFP driven by the AMS promoter was prematurely expressed in the tapetal nuclei, leading to complete male sterility in the pAMS:TEK-GFP (pat) transgenic lines with the wild-type background. Cytological observations in the pat anthers showed impaired callose synthesis and aberrant exine patterning. CALLOSE SYNTHASE5 (CalS5) is required for callose synthesis, and expression of CalS5 in pat plants was significantly reduced. We demonstrated that TEK negatively regulates CalS5 expression after the tetrad stage in wild-type anthers and further discovered that premature TEK-GFP in pat directly represses CalS5 expression through histone modification. Our findings show that TEK flexibly mediates its different functions via different temporal regulation, revealing that the temporal regulation of TEK is essential for exine patterning. Moreover, the result that the repression of CalS5 by TEK after the tetrad stage coincides with the timing of callose wall dissolution suggests that tapetum utilizes temporal regulation of genes to stop callose wall synthesis, which, together with the activation of callase activity, achieves microspore release and pollen wall patterning.
| S-EPMC7252695 | biostudies-literature
Project description:Background and aimsThe Arabidopsis thaliana pollen cell wall is a complex structure consisting of an outer sporopollenin framework and lipid-rich coat, as well as an inner cellulosic wall. Although mutant analysis has been a useful tool to study pollen cell walls, the ultrastructure of the arabidopsis anther has proved to be challenging to preserve for electron microscopy.MethodsIn this work, high-pressure freezing/freeze substitution and transmission electron microscopy were used to examine the sequence of developmental events in the anther that lead to sporopollenin deposition to form the exine and the dramatic differentiation and death of the tapetum, which produces the pollen coat.Key resultsCryo-fixation revealed a new view of the interplay between sporophytic anther tissues and gametophytic microspores over the course of pollen development, especially with respect to the intact microspore/pollen wall and the continuous tapetum epithelium. These data reveal the ultrastructure of tapetosomes and elaioplasts, highly specialized tapetum organelles that accumulate pollen coat components. The tapetum and middle layer of the anther also remain intact into the tricellular pollen and late uninucleate microspore stages, respectively.ConclusionsThis high-quality structural information, interpreted in the context of recent functional studies, provides the groundwork for future mutant studies where tapetum and microspore ultrastructure is assessed.
| S-EPMC4195548 | biostudies-literature
Project description:Exine, the outermost architecture of pollen walls, protects male gametes from the environment by virtue of its chemical and physical stability. Although much effort has been devoted to revealing the mechanism of exine construction, still little is known about it. To identify the genes involved in exine formation, we screened for Arabidopsis mutants with pollen grains exhibiting abnormal exine structure using scanning electron microscopy. We isolated 12 mutants, kaonashi1 (kns1) to kns12, and classified them into four types. The type 1 mutants showed a collapsed exine structure resembling a mutant of the callose synthase gene, suggesting that the type 1 genes are involved in callose wall synthesis. The type 2 mutant showed remarkably thin exine structure, presumably due to defective primexine thickening. The type 3 mutants showed defective tectum formation, and thus type 3 genes are required for primordial tectum formation or biosynthesis and deposition of sporopollenin. The type 4 mutants showed densely distributed baculae, suggesting type 4 genes determine the position of probacula formation. All identified kns mutants were recessive, suggesting that these KNS genes are expressed in sporophytic cells. Unlike previously known exine-defective mutants, most of the kns mutants showed normal fertility. Map-based cloning revealed that KNS2, one of the type 4 genes, encodes sucrose phosphate synthase. This enzyme might be required for synthesis of primexine or callose wall, which are both important for probacula positioning. Analysis of kns mutants will provide new knowledge to help understand the mechanism of biosynthesis of exine components and the construction of exine architecture.
| S-EPMC2566931 | biostudies-literature
Project description:Connecticut nursery pollen metabarcoding
| PRJNA548313 | ENA