Sporopollenin: Imaging and Spectroscopy - Literature review Example

Name: Professor: Date: Sporopollenin: Imaging and Spectroscopy Introduction Wakil et al. (2010), state that the word sporopollenin was coined by Zetzschel in description of the exteremely resilient exine of spores and pollen as well as other microspores. Archeologists have found pristine pollen grains which have survived inside sedimentary rocks for periods agreed upon as being over 500 millon years. There has also been found micro spores dated to be of at least 3.5 x 109 years. Most of the initial information regarding sporopollenin was from Brooks et al. The word sporopollenin is a chemical term and therefore has no reference to botanical taxonomy. Barrier et al. (2010) argue that the reason why there is little understanding of the precise structure of sporopollenin, both chemical and physical is due to the resilience and seeming indestructibility of the exine. There however been attempts to understand it by different researchers especially owing to its usefulness in aromatics and other areas. This paper reviews the existing literature on different aspects of sporopollenin. Which include its, structure, properties of sporopollenin, extraction of different components from sporopollenin including DNA among others, It uses, and their antioxidant properties . It also delves into the removal of metal ions from sporopollenin solution and also includes an examination of how metal bind to sporopollenin, what the effects are and what the perfect conditions are. There is, apparently a very wide range of sporopollenin from different organisms which in turn are varied according to the environment where they are found. This makes the literature varied depending on from which organisms and regions the sporopollenin examined was gathered. This combined, with the extreme difficulty encountered in breaching the exine makes concensus far between. The literature available is examined according to the parameters outlined above. Structure of sporopollenin Lorch et al. (2009), argue for the use of the infrared absorption spectra to establish the structure of sporopollenin. The spectra show a general likeness of all sporopollenin tested. Brooks also advocates the use of Pyrolysis for the investigation of sporopollenin. Spore-derived exines are partially permissive due to the presence of microspores. These microspores are instrumental in the extraction of other components of sporopollenin in order to remain with the exine. The microspores allow oxygen to pass through an occurrence that would be expected to lead to oxidation of the lipids and other components of sporopollenin. Oxidation does not occur under these circumstances which gives rise to the interest in study sporopollenin’s antioxidiant capabilities (Barrier, 2010). According to Neto et al. (2008) the commonly held position with regard to sporopollenin chemical and physical structure is that they consist of they contain a variety of aromatics and also unbranched aliphatics. Moliner et al. ( 2007) on the other hand observe that that there has already been established a strong similarity between green algae’s cell wall and spores from vascular plants this indicates that some sporopollenin, though not everything is known about them may have similar cell structure to some plant cells. They farther point out that studies of Osmunda regalis and Selaginella kraussiana which are both spores of Lycopodiacene pecies showed evidence of sporopollenin. The examination of the above spores among others shows that there is patterning on the exine of sporopollenin, particularly in pollen grains. The first observation of this feature was made by (Goodby et al., 2007: Len & Mackenzie, 2006). Ewing et al. (2006) further observe that Glycocalyx determines these specific characteristics on the surface of a microspore. Specific characteristics of the exine pattern which are also referred to as the template, are comprised of polyionic molecules forming agglutination bridges connecting near-spaced negatively charged areas on the compounds that form part of the exine. The surface also has filaments which are also refered to as lamellations which, after oxidative degradation, are exposed. This oxidative degradation was observed in mature Lycopodium clavatum spores (Lipka et al. 2004). Blinks et al. (2005) report that some sporopollenins have a molecular structure that is based on oxidative polymaer carotenoid esters and carotenoids. In this regard, Baunob et al. (2005) argues that sporopollenins are osmophilic contain aliphatic unsaturated C – C double bonds. Its reaction with basic dyes suggests that in it are weakly anionic groups which include acidic – enolic compounds. While this characteristic may lead to sporopollenin being confused with cutin and suberin during experiments, sporopollenin does not stain with Sudan IV while cutin and suberin do, thus the distinction between them. Another distinguishing factor is the fact that sporopollenin does not stain with phloroglucinol while cutin and suberin stain (Blinks et al. 2005). In geochemical terms, Ewing et al, 2003 observe that changes in the chemical and physical composition of sporopollenin depend on different rates of heating. Variation in heating has been acknowledged as the most important factor in these changes due to the chemical reactions they catalyse. According to the temperature at which pollen and spores carbonize is determined by the geothermal history of the subject up to that point of carbonization. Colour and chemical changes do not occur below 2000 of heat. Above 2200 all samples of sporopollenin, both pollen and spores, show considerable changes both in colour and chemical composition. They also produce soluble and unstable chemical products. Compared to dinoflagellates, changes in colour and chemical structures are different in sporopollenin. Compared to chitinozoa, Sporopollenin change faster. Research has, overtime, shown that in terms of chemical composition there exists great similarity between walls of spores and pollen from many types of plants across species. The xylan fraction in all of them is not well defined but constitutes approximately 10% of the entire pollen or spore wall while the outermost wall is resistant to chemicals. it however oxidizes easily in a combination of dicarboxylic acids (mono arid) which contain less than eighteen carbon atoms (Moliner et al. 2003). The characteristic of angiosperm pollen is unique especially with regard to its composition. This is because inside the pollen are complete sperm cells which are caused by asymmetric mitosis which is necessary for double fertilization as is necessary for the reproduction of plants with angiosperm cells. According to Barrier (2008), Sporopollenin exine capsules (SEC) are made from sporopollenin which has the same characteristics as polycarotenoid and consist of carbon, oxygen and hydrogen. This is what makes them almost completely resistant to corrosion from either acids or alkalis as illustrated by their ability to pass through digestive systems of animals intact. Due to the fact that they leave the digestive system completely unchanged, the sporopollenin that go into the animal’s alimentary canal have no adverse allergic effects on the animal. This property allows some aromatic sporopollenin to be used as spices, they give aroma to food without being digested even after ingestion. (Barrier, et al. 2013). The SEC are also elastic and can withstand a lot of compressive pressure after which they return to their original form. In their examination of the morphology of the pollen of species in the genus Oxytropis note that the grains are isopolar, subprolate and have a radially symmetric shape. The grains are ornamented differently in five categories depending with the climatic zones from which they come. There were ten pollen types in the region where this research was conducted. Len et al. (2006) described Oxytropis as fine, reticulate, subprolate and prolate. The characteristics are an indication of the similarities between the eleven species that the study investigated. Spores, particularly, bacterial spores are able to abide to stable surfaces of food processing plants owing to their physical characteristics and chemical composition. Certain spores, particularly from the Clostridium and Bacillus have appendages that are adapted for the formation of biofilm. The appendages are, however absent in B. mycoides. While appendages are not found in all of them, all spores have exosporia. Among the strains that have appendages, there is a variance in the number of appendages from strain to strain. B. cereus has a lower number of appendages (3 to 4) while B.thuringiensis has more appendages (12 to 18). The lengths and shapes of these appendages are different in the different strains in B. cereus the length ranges from 0.45 to 3.8 µm and it is tubular in shape. The diameter of the appendages is 13.6nm on averagely. While different strains are widely varied morphologically, the differences between them in terms of hydrophobicity is not as varied, all strains examined by Guilford at al. 2008 raged between 42.4 and 55.6 %. All of them, therefore, remain true to the to the characteristics of sporopollenin in as far as imperviousness is concerned. This is because the above indicated hydrophobicity range is moderately or highly hydrophobic. The presence of appendages and high hydrophobicity, they conclude, could be the reason why B.cerevs is so difficult to get rid of. Chambert et al. (2007) concluded that spores and pollen have a change of colour from pale yellow to medium brown in different ranks of coal the different shades are due to differences in thermal histories. The changes are also as a result of coalification as well as carbonization process. The coalification process also affects their reflective ability. When the pollen and spores are yet immature during carbonization the changes in colour are slow and relatively small. When they are mature, however, chemical components of the sporopollenin are broken down in order to produce hydrocarbons and the change is now more rapid from orange to brown. Carbonization also leads to the formation of aromatic rings. The table below give an illustration of the above information. Sample no Grid reference and location Formation/age Rv% a* b* L* Colour D8657.42 James Ross Island, Antarctic Peninsula Santa Marta Fm, Cretaceous 0.22 4.0 19.4 91.78 Pale yellow D8653.28 James Ross Island, Antarctic Peninsula Santa Marta Fm, Cretaceous 0.27 6.3 20.5 88.33 Pale yellow D8659.35 James Ross Island, Antarctic Peninsula Santa Marta Fm, Cretaceous 0.37 6.5 21.7 88.55 Pale yellow Yaverland 2 SZ 618 853, Isle of Wight, UK Vectis Fm. Cretaceous 0.37 10.5 27.3 84.76 Pale lemon yellow Watchet 14 ST 083 435, Watchet, Somerset, UK Westbury Fm, Late Triassic 0.40 6.4 22.8 90.37 Pale yellow Inn 23 NM 694 454, Loch Aline, Scotland Pabba Beds, Jurassic 0.56 8.5 23.6 87.98 Pale yellow Inn 37 NM 694 454, Loch Aline, Scotland Pabba Beds, Jurassic 0.67 9.6 23.8 82.98 Lemon yellow mv 87/66 NO 545 375, Midland Valley, Scotland Gedinnian, Early Devonian 0.73 20.6 32.2 61.56 Orange Papa 18.5 HU 1715 5905, Papa Stour, Shetland Eifelian, Mid Devonian 0.75 17.8 23.6 58.79 Orange mv 87/64 NO 538 394, Midland Valley, Scotland Gedinnian, Early Devonian 0.77 20.2 28.6 58.15 Orange Inn 8 NM 070 435, Inninmore, Scotland Westphalian B, Carboniferous 0.78 15.5 26.5 63.46 Orange Inn 9 NM 070 435, Inninmore, Scotland Westphalian B, Carboniferous 0.80 13.8 29.5 71.17 Yellow orange mv 88/1 NO 730 820, Midland Valley, Scotland Gedinnian, Early Devonian 0.96 19.2 26.9 58.8 Orange mv 87/88 NO 446 367, Midland Valley, Scotland Gedinnian, Early Devonian 1.04 14.8 12.6 50.87 Orange brown mv 87/91 NO 441 370, Midland Valley, Scotland Gedinnian, Early Devonian 1.04 18.8 18.6 48.19 Orange brown mv 87/89 NO 441 370, Midland Valley, Scotland Gedinnian, Early Devonian 1.09 17.4 13.2 40.67 Dark brown mv 85/44 NN 656 019, Midland Valley, Scotland Emsian, Early Devonian 1.28 14.9 6.3 41.01 Dark brown Foula 56 HT 960 415, Foula, Shetland Emsian, Early Devonian 1.50 10.4 2.0 37.37 Dark brown Foula 57 HT 958 414, Foula, Shetland Emsian, Early Devonian 1.80 11.9 6.0 32.78 Black Table 1 Figure one below shows the graphical representation of the above data. The measurement of the colour of and the reflective ability of sporopollenin was measured in order to establish the relationship between the two should it exist. The experiment to establish micro FT – IR alongside transmitted and reflected light microphotometry is intended to check this relationship. Extraction Paunov et al. (2007) observe that extraction of sporopollenin where separation with microspores is concerned involves its successive exposure to reagents including solvents and acid as well as caustic. This removes the intine cellulose wall after the sporopollenin has further been treated in order to produce a negative ninhydrin. Lorch et al (2009) further argue that sporopollenin may also be extracted by use of basic amino acids whether alphatic or aromatic. The extraction occurs when they are reacted with the likes of 1,3b-diamino propane and peptide chains. There exists a lot more literature on the extraction of spores than pollen due to the interest in spores that is due to the disease causing properties of some of their strains. The most researched of these include Bacillus anthracis, B. cereus among others. The heightened alert due to bio terrorism has increased these emphases. Lipka et al. (2004) provide a procedure for the extraction of exine shells from sporopollenin. The procedure involves suspension of loose powder particles of pollen in acetone and stirring the suspension for up to four hours in order to produce DFS. This is followed by followed by overnight drying. They suggest that the now defatted sample be then suspended in 750 cm3 of 6% potassium hydroxide acqueous solution and stirring done for up to six hours. In this procedure the next stage is filtration at grade four porosity. The resultant solid, they explain, should be washed three times in 300 cm3 of hot water. After washing with water, another wash in hot ethanol is required; two times in 300 cm3 and then the sporopolenin is refluxed in 750 cm3 of ethanol for up to two hours. Overnight drying of these overnight produces Base-hydrolysed Sporopollenin (BHS) which after suspension in750 cm3 of 85% orthophosphoric acid followed by stirring under reflux for seven days followed filtering at porosity grade 4. It is then washed five times in 250 cm3 of water, once in 250 cm3 acetone, once in 250cm3 , 2 molar acqueous hydrochloric acid, once in 250cm3 2 Molar acqueous sodium hydroxide and 250 cm3 of ethanol. To complete the extraction process sporopollenin is then dried in a vaccum over phosphorus oxide at 600 which gives 58-60g of sporopollenin exines. The sample from which these exines are acquired after the above mentioned process is 250 g of loose powder pollen.The filamentous structures in Lycopodium clavitum can be extracted by use of 2 aminolethanol as well as phenal extraction. Paunov et al. (2006); Dumolin et al. (2007) detail the process of extracting microcapsules from Lycopodium clavatum. They argue that this can be done using both organic and inorganic nanomaterials through preparation of nanoparticles on site. The illustration for this was done using pollen grains from both mosses and ferns. The extraction of nanomaterials is important since it allows for the access of the drugs and nutraceutical delivery systems there are they are also high quality food supplements which can either be ingested or injected the reaction for this extraction is conducted on the inside of the capsule through loading of the reagent into the exine of the capsule, the reagent used is normally Omega 3 fat. The method, they proposed, should be in three steps. The steps include the following Step1 involves putting the reagent or a mixture of reagents into the capsule. The next step involves the filtration and cleaning of capsules using pure solvent in order to rid the samples of the excess reagents. The capsules are then dispersed into a different reagent whose reaction with the first gives rise to a different reagent C which has low solubility. The third step is the is cleaning of the capsules of reagent B and redispersion of the capsules in water. In Sandbhou et al.(2004) experiment, sporopollenin from Lycopodium clavatum was used to establish sorption of sporopollenin through the employment of glutaraldehyde as the absorbent to a heavy metal ion. FTIR was used to examine the structure of the surface of the sporopollenin under observation. Thermal analysis was also used to check other parameters including the pH of the solution in question, molarity of the solution as well as different temperatures where reactions could be recorded. The experiment also involved the establishment of thermodynamic parameters. It was noted that the most efficient range of pH Carbon dioxide in this experiment was at pH5.5 while Nickel (II) and Copper (II) did best at pH 5. Application of Sporopollenin Spore derived exine shells can be used as antioxidant as well as conduits with antioxidant properties. When the exine is isolated from the spores through successive organic solvent treatment, the hollow capsule exine (Paunov et al. 2007). The exine shell acts as an antioxidiant which protects lipids from oxidation. Exine shells for the purpose of producing antioxidiants can be derived from any naturally occurring spores. The species that are suitable for these spores are presented in the following table together with their diameters. Spore Diameter Bacillus subtilis 1.2 μm Myosotis 2.4-5 μm Aspergillus niger 4 μm Penicillium 3-5 μm Cantharellus minor 4-6 μm Ganomerma 5-6.5 μm Agrocybe 10-14 μm Urtica dioica 10-12 μm Periconia 16-18 μm Epicoccum 20 μm Lycopodium clavatum 25 μm Lycopodium clavatum 40 μm Abies 125 μm Cucurbitapapo 200 μm Cuburbita 250 μm Another application of the exine shell is the increase the oxidative stability of an oxidative agent. Exine shells also are effective in protecting from effects from the elements. The fact that spore exines are inexpensive makes them suitable for use as delivery vehicles for active substances (Barrier 2010). Works cited Banoub, J. H., Newton, R. P., Esmans, E., Ewing, D. F., and Mackenzie, G., Recent developments in mass spectrometry for the characterization of nucleosides, nucleotides, oligonucleotides, and nucleic acids, Chem. Rev., 105, 1869-915. doi: 10.1021/cr030040w (2005) Barrier, S., Lobbert, A., Boasman, A. J., Boa, A. N. , Lorch, M., Atkin, S. L., and Mackenzie, G., Access to a primary aminosporopollenin solid support from plant spores, Green Chemistry, 12, 234-40. doi: 10.1039/b913215e (2010) Barrier,S., Rigby, A. S. Diego-Taboada, A. Thomasson, M. J,. Mackenzie, G and Atkin, S. L., Sporopollenin exines: A novel natural taste masking material, LWT-Food Sci. Technol., 43, 73-6. doi: 10.1016/j.lwt.2009.07.001 (2010). Binks, B. P., Clint, J. H., Mackenzie, G., Simcock, C. and Whitby, C. P. Naturally occurring spore particles at planar fluid interfaces and in emulsions, , Langmuir ,21, 8161-7. doi: 10.1021/la0513858 (2005) Chambert, S., Doutheau, A., Queneau, Y., Cowling, S. J., Goodby, J. W., and Mackenzie, G. Synthesis and thermotropic behavior of simple new glucolipid amides, , J. Carbohydr. Chem., ,26, 27-39. doi: 10.1080/07328300701252565 (2007) Dumoulin, F., Lafont, D., Huynh, T. L., Boullanger, P., Mackenzie, G., West, J. J. and Goodby, J. W., Synthesis and liquid crystalline properties of mono-, di- and tri-O-alkyl pentaerythritol derivatives bearing tri-, di- or monogalactosyl heads: The effects of curvature of molecular packing on mesophase formation,Chem.-Eur. J., 2007, 13, 5585-600. doi: 10.1002/chem.200601702 (2007) Ewing, D. F., Glacon, V., Len, C. and Mackenzie, G., Synthesis, conformation and antiviral activity of nucleoside analogues with the (2-hydroxy-1-phenylethoxy) methyl glycone - a family of nucleoside analogues related to d4T and aciclovir, New J. Chem ,29, 1461-8. doi: 10.1039/b510056a (2005) Ewing, D. F., Glacon, V., Mackenzie, G., Postel, D., and Len, C., Synthesis of acyclic bis-vinyl pyrimidines: a general route to d4T via metathesis, Tetrahedron, , 59, 941-5. doi: (2003) Laurent, N., Lafont, D., Dumoulin, F., Boullanger, P., Mackenzie, G., Kouwer, P. H. J. and Goodby, J. W., Synthesis of amphiphilic phenylazophenyl glycosides and a study of their liquid crystal properties, , J. Am. Chem. Soc. 125, 15499-506. doi: 10.1021/ja037347x (2003) Len, C. and Mackenzie, G. Synthesis of 2 ',3 '-didehydro-2 ',3 '-dideoxynucleosides having variations at either or both of the 2 '- and 3 '-positions, ,Tetrahedron,, 62, 9085-107. doi: 10.1016/j.tet.2006.07.050 (2006) Lorch,M., Thomasson, M. J., Diego-Taboada, A., Barrier, S., Atkin, S. L., Mackenzie, G., and Archibald, S. J, MRI contrast agent delivery using spore capsules: controlled release in blood plasma, Chem. Commun., 6442-4. doi: 10.1039/b909551a (2009). Molinier, V., Kouwer, P. J. J., Fitremann, J, Bouchu, A., Mackenzie, G., Queneau, Y., and Goodby, J. W., Shape dependence in the formation of condensed phases exhibited by disubstituted sucrose esters, ,Chem.-Eur. J., , 13, 1763-75. doi: 10.1002/chem.200600368 (2007) Molinier, V., Kouwer, P. H. J., Fitremann, J., Bouchu, A., Mackenzie, G., Queneau, Y., and Goodby, J. W., Self-organizing properties of monosubstituted sucrose fatty acid esters: The effects of chain length and unsaturation, Chem.-Eur. J., 12, 3547-57. doi: 10.1002/chem.200500773(2008) Neto, V., Granet, R., Mackenzie, G. and Krausz, P. Efficient synthesis of "Star-like" surfactants via "Click Chemistry" [3+2] copper (I)-catalyzed cycloaddition, J. Carbohydr. Chem., 27, 231-7. doi: 10.1080/07328300802105365 ( 2008) Paunov, V. N., Mackenzie, G. and Stoyanov, S. D. Sporopollenin micro-reactors for in-situ preparation, encapsulation and targeted delivery of active components, J. Mater. Chem., 17, 609-12. doi: 10.1039/b615865j ( 2007). Sandbhor, U., Kulkarni, P., Padhye, S., Kundu, G., Mackenzie, G., and Pritchard, R., Antimelanomal activity of the copper(II) complexes I-substituted 5-amino-imidazole ligands against B16F10 melanoma cells, Bioorg. Med. Chem. Lett., , 14, 2877-82. doi: Lipka, E., Daniel, C., Vaccher, M. P., Glacon, V., Ewing, D. Mackenzie, G., Len, C., Bonte, J. P. and Vaccher, C., Enantioseparation of new nucleoside analogs, related to d4T and acyclovir, by chiral capillary electrophoresis using highly sulfated beta-cyclodextrins, Electrophoresis, ,25, 444-53. doi: 10.1002/elps.200305706 (2004) Molinier, V., Kouwer, P. H. J., Queneau, Y., Fitremann, J., Mackenzie, G. and Goodby J. W., A bilayer to monolayer phase transition in liquid crystal glycolipids, , Chem. Commun., , 2860-1. doi: 10.1039/b308880d (2003) Wakil, A., Mackenzie, G., Diego-Taboada, A., Bell, J. G. and Atkin, S. L. Enhanced Bioavailability of Eicosapentaenoic Acid from Fish Oil After Encapsulation Within Plant Spore Exines as Microcapsules, Lipids,45, 645-9. doi: 10.1007/s11745-010-3427-y (2010). Read More

Blinks et al. (2005) report that some sporopollenins have a molecular structure that is based on oxidative polymaer carotenoid esters and carotenoids. In this regard, Baunob et al. (2005) argues that sporopollenins are osmophilic contain aliphatic unsaturated C – C double bonds. Its reaction with basic dyes suggests that in it are weakly anionic groups which include acidic – enolic compounds. While this characteristic may lead to sporopollenin being confused with cutin and suberin during experiments, sporopollenin does not stain with Sudan IV while cutin and suberin do, thus the distinction between them.

Another distinguishing factor is the fact that sporopollenin does not stain with phloroglucinol while cutin and suberin stain (Blinks et al. 2005). In geochemical terms, Ewing et al, 2003 observe that changes in the chemical and physical composition of sporopollenin depend on different rates of heating. Variation in heating has been acknowledged as the most important factor in these changes due to the chemical reactions they catalyse. According to the temperature at which pollen and spores carbonize is determined by the geothermal history of the subject up to that point of carbonization.

Colour and chemical changes do not occur below 2000 of heat. Above 2200 all samples of sporopollenin, both pollen and spores, show considerable changes both in colour and chemical composition. They also produce soluble and unstable chemical products. Compared to dinoflagellates, changes in colour and chemical structures are different in sporopollenin. Compared to chitinozoa, Sporopollenin change faster. Research has, overtime, shown that in terms of chemical composition there exists great similarity between walls of spores and pollen from many types of plants across species.

The xylan fraction in all of them is not well defined but constitutes approximately 10% of the entire pollen or spore wall while the outermost wall is resistant to chemicals. it however oxidizes easily in a combination of dicarboxylic acids (mono arid) which contain less than eighteen carbon atoms (Moliner et al. 2003). The characteristic of angiosperm pollen is unique especially with regard to its composition. This is because inside the pollen are complete sperm cells which are caused by asymmetric mitosis which is necessary for double fertilization as is necessary for the reproduction of plants with angiosperm cells.

According to Barrier (2008), Sporopollenin exine capsules (SEC) are made from sporopollenin which has the same characteristics as polycarotenoid and consist of carbon, oxygen and hydrogen. This is what makes them almost completely resistant to corrosion from either acids or alkalis as illustrated by their ability to pass through digestive systems of animals intact. Due to the fact that they leave the digestive system completely unchanged, the sporopollenin that go into the animal’s alimentary canal have no adverse allergic effects on the animal.

This property allows some aromatic sporopollenin to be used as spices, they give aroma to food without being digested even after ingestion. (Barrier, et al. 2013). The SEC are also elastic and can withstand a lot of compressive pressure after which they return to their original form. In their examination of the morphology of the pollen of species in the genus Oxytropis note that the grains are isopolar, subprolate and have a radially symmetric shape. The grains are ornamented differently in five categories depending with the climatic zones from which they come.

There were ten pollen types in the region where this research was conducted. Len et al. (2006) described Oxytropis as fine, reticulate, subprolate and prolate. The characteristics are an indication of the similarities between the eleven species that the study investigated. Spores, particularly, bacterial spores are able to abide to stable surfaces of food processing plants owing to their physical characteristics and chemical composition. Certain spores, particularly from the Clostridium and Bacillus have appendages that are adapted for the formation of biofilm.

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