Name: extraction of plant-based capsules for microencapsulation applications
Uploaded: 2016-11-09T14:00:00.000+00:00
Duration: 10 min 54 s
Description: Watch this Scientific Journal Video about Extraction of Plant-based Capsules for Microencapsulation Applications at JoVE.com

This work was supported by the National Research Foundation (NRF-NRFF2011-01) and the National Medical Research Council (NMRC/CBRG/0005/2012).

L. Sporopollenin ekzin Capsule&#39;lerden 1. Ekstraksiyon (sn) clavatum Sporlar Not: SEC çıkarma işlemi bir yanıcı toz (L. clavatum), sıcak aşındırıcı asitler ve yanıcı solventler, dolayısıyla uygun kişisel koruyucu donanımlar (gözlük, yüz maskesi, eldiven, laboratuvar önlüğü), kullanım onaylanan risk değerlendirmesini ve bertaraf gerektirir yetkili laboratuvar personeli tarafından kimyasallar esastır. <ol><li> Spore yağları azaltır <ol><li> Bir cam kondenser, su dolaşım sistemi ve su banyosu (Şekil 1) kullanarak bir davlumbaz bir reflü set-up hazırlayın. </li><li> 100 g L. tartılır uzakta herhangi bir ateşleme kaynağından toz oluşturarak olmadan clavatum sporlar. Not: Ticari olarak elde edildi Burada kullanılan sporların (Malzeme Listesi). </li><li> Manyetik bir karıştırma çubuğu ile 1 L&#39;lik bir politetrafloroetilen (PTFE) yuvarlak dipli bir şişeye yavaş yavaş Aktarım sporlar. </li><li> sporlar için 500 ml aseton eklePTFE şişesi ve homojen bir süspansiyon oluşturmak için hafifçe çalkalanır. </li><li> 50 ° C&#39;de su banyosu grubu PTFE içine yerleştirilir ve geri akış yoğunlaştırıcısı bağlanabilir. </li><li> 200 rpm&#39;de karıştırılarak 6 saat aseton içinde geri gerçekleştirmek ve daha sonra oda sıcaklığında soğumaya bırakın. </li><li> vakum altında filtre kağıdı kullanılarak süspansiyon filtre edin ve büyük (15 cm çapında) bir cam Petri kapları içinde yağdan arındırılmış sporlar toplar. </li><li> Çözücü buharlaştırma için delikli alüminyum folyo ile kaplayarak oda sıcaklığında (davlumbaz) de sporlar kurutun. </li></ol></li><li> asidoliz <ol><li> % 85 deiyonize (Di) su (h / h) fosforik asit hazırlamak ve 1 L&#39;lik bir PTFE şişeye yağsız kuru sporların aktarın. </li><li> yağı alınmış sporların fosforik asit (% 85 h / h) 500 ml ilave edilir. </li><li> 70 ° C&#39;de bir su banyosu grubu PTFE içine yerleştirilir ve geri akış yoğunlaştırıcısı bağlanabilir. </li><li> 30 saat süre ile fosforik asit içinde hafif bir geri akış (70 ° C) gerçekleştirmek ve daha sonra izinOda sıcaklığında soğutulur. </li><li> 500 ml ılık distile su kullanılarak süspansiyon sulandırmak ve filtre kağıdı kullanılarak vakumla süzme ile saniye toplamak. </li><li> bir dizi yıkamaya tabi gerçekleştirmek için 3 L cam behere saniye aktarın. </li></ol></li><li> SEC Yıkama <ol><li> Davlumbaz saniye içeren 3 L cam behere koyun. </li><li> 10 dakika için hafifçe karıştırılarak sn 800 ml sıcak (50 ° C), DI su ilave edilir. </li><li> filtre kağıdı ve bir vakum filtrasyonu kurulumu kullanılarak süspansiyon filtre. </li><li> Temiz 3 L cam beher saniye toplayın. </li><li> Yineleyin 1.3.1 adımları. 1.3.4 için. Beş kere. </li><li> 10 dakika için hafifçe karıştırılarak sn 600 ml sıcak (45 ° C) aseton ekleyin. </li><li> Vakum altında süzme ile saniye toplamak ve temiz bir 3 L&#39;lik bir cam behere saniye aktarın. </li><li> Yineleyin 1.3.6 adımları. ve 1.3.7. Sıcak 600 ml (50 ° C), 2 M hidroklorik asit kullanılarak. </li><li> Yineleyin 1.3.6 adımları. ve 1.3.7. ho 600 ml kullanılarakt (50 ° C), 2 M sodyum hidroksit. </li><li> Tekrarlayın 1.3.1 ve 1.3.4 sekiz kez yineleyin. </li><li> Yineleyin 1.3.6 adımları. ve 1.3.7. Sıcak 600 ml (45 ° C) aseton kullanılmıştır. </li><li> Yineleyin 1.3.6 adımları. ve 1.3.7. Sıcak 600 ml (45 ° C) etanol kullanılmıştır. </li><li> Yineleyin 1.3.6 adımları. ve 1.3.7. Sıcak 800 ml (50 ° C) DI suyun kullanımı. </li><li> su içeriği kaldırmak için 24 saat boyunca oda sıcaklığında bir çeker ocak içinde altı büyük cam Petri kapları ve kuru temiz sn aktarın. </li><li> 60 ° C ve 10 saat süre ile ya da sabit bir ağırlığa kadar, 1 mbar vakumda bir vakumlu fırın içinde saniye kurutulur. </li><li> kurutulmuş saniye toplayın ve bir polipropilen şişe aktarın. </li><li> kuru kabine saniye saklayın. </li></ol></li></ol> SECS 2. Karakterizasyonu <ol><li> Farklı aşamalarında üretilen sporları ve Secs kullanarak taramalı elektron mikroskobu analizi 41 gerçekleştirin. </li><li> Farklı aşamalarında üretilen sporları ve Secs kullanarak element analizi 41 gerçekleştirin. DRy element analizi önce en az 1 saat süre ile 60 ° C &#39;de, tüm örnekler ve 6.25 bir çarpma faktörü ile, yüzde azot kullanılarak protein içeriğini hesaplamak. 11, her numune için üç kez ölçümleri kullanılarak sonuçlar elde edilir. </li><li> Bir dinamik görüntü parçacık analizörü kullanılarak Mikromeritik özellikleri analiz yapmak. 41 </li><li> Konfokal lazer tarama mikroskobu kullanarak, işlenmemiş Tarama işlenir ve FITC-BSA-yüklü SEC sn. 41 </li></ol> Vakum Yükleme Tekniği ile 3. biyomakromolekülün Enkapsulasyon <ol><li> 50 ml&#39;lik bir polipropilen tüpü içine distile su ve transferi ile 125 mg, 1.2 ml / ml bovin serum albümini (BSA) çözeltisi hazırlayın. </li><li> Saniye 150 mg tartılır ve BSA solüsyonuna aktarın. </li><li> homojen bir çözelti meydana getirmek üzere, 10 dakika boyunca tüp vorteksleyin. </li><li> filtre kağıdı kullanılarak tüp örtün. </li><li> 1 mbar&#39;da vakumla 4 saat dondurularak kurutma şişesine ve kuru tüp aktarın. </li><li> 3.1 adımları gerçekleştirin3.5. üç bağımsız gruplar için. </li><li> BSA yüklü saniye toplayın ve bir havan kullanarak topakların kaldırın. </li><li> 50 ml&#39;lik bir tüp BSA yüklü saniye aktarın ve artık BSA kaldırmak için 2 ml DI su ekleyin. </li><li> 5 dakika boyunca 4704 xg santrifüj yoluyla saniye toplayın ve supernatant atın. </li><li> bir zamanlar DI suyla yıkamayı tekrarlayın. </li><li> Filtre kağıdı kullanarak BSA yüklü saniye içeren tüp örtün. </li><li> bir dondurucu (-20 ° C) saniye aktarın ve 1 saat dondurmak. </li><li> Daha fazla karakterizasyon kadar dondurucuda 24 saat ve mağaza için saniye kurumaya dondurun. </li><li> adımlarda 3.1 aynı prosedürü kullanarak BSA olmadan plasebo Secs hazırlayın. 3.13 için. </li><li> 3.1 adımlarda aynı prosedürü kullanarak FITC-BSA yüklü saniye hazırlayın. 3.13 için. </li></ol> Kapsülleme Etkinliğinin Belirlenmesi 4. <ol><li> 2 ml polipropilen tüplerde BSA yüklü saniye 5 mg tartılır. </li><li> 1.4 mi fosfataz takımı için Phospha eklemeTe, 5 dakika boyunca salin pH 7.4 (PBS) ve vorteks tamponlu. </li><li> Sonda 10 sn 3 döngüleri için% 40 genlik süspansiyon sonikasyon. </li><li> 0.45 mikron Polyethersulfone (PES) filtre kullanarak çözüm Filtre. </li><li> adımlarda 4.1 prosedürün aynısını. 4.4&#39;e. plasebo Secs kullanarak. </li><li> boş olarak plasebo ile 280 nm&#39;de absorbansı ölçülür. </li><li> BSA miktarı, PBS içinde BSA standart bir eğrisi (200 ila 1000 ug / ml) ile kapsüllenmiş ölçmek. 42 </li></ol>

<ol>
	<li>Paunov, V. N., Mackenzie, G., Stoyanov, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sporopollenin+micro-reactors+for+in-situ+preparation,+encapsulation+and+targeted+delivery+of+active+components.">Sporopollenin micro-reactors for in-situ preparation, encapsulation and targeted delivery of active components.</a> J. Mater. Chem. 17 (7), 609-612 (2007).</li><li>Barrier, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Physical and chemical properties of sporopollenin exine particles. Doctoral dissertation thesis. , (2008).</li><li>Beckett, S. T., Atkin, S. L., Mackenzie, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dosage+Form.">Dosage Form.</a> US Patent. , (2009).</li><li>Lorch, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MRI+contrast+agent+delivery+using+spore+capsules:+controlled+release+in+blood+plasma.">MRI contrast agent delivery using spore capsules: controlled release in blood plasma.</a> Chem. Comm. (42), 6442-6444 (2009).</li><li>Wakil, A., Mackenzie, G., Diego-Taboada, A., Bell, J. G., Atkin, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+bioavailability+of+eicosapentaenoic+acid+from+fish+oil+after+encapsulation+within+plant+spore+exines+as+microcapsules.">Enhanced bioavailability of eicosapentaenoic acid from fish oil after encapsulation within plant spore exines as microcapsules.</a> Lipids. 45 (7), 645-649 (2010).</li><li>Barrier, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sporopollenin+exines:+A+novel+natural+taste+masking+material.">Sporopollenin exines: A novel natural taste masking material.</a> LWT-Food Sci. Technol. 43 (1), 73-76 (2010).</li><li>Barrier, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Viability+of+plant+spore+exine+capsules+for+microencapsulation.">Viability of plant spore exine capsules for microencapsulation.</a> J. Mater. Chem. 21 (4), 975-981 (2011).</li><li>Hamad, S. A., Dyab, A. F., Stoyanov, S. D., Paunov, V. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Encapsulation+of+living+cells+into+sporopollenin+microcapsules.">Encapsulation of living cells into sporopollenin microcapsules.</a> J. Mater. Chem. 21 (44), 18018-18023 (2011).</li><li>Diego-Taboada, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sequestration+of+edible+oil+from+emulsions+using+new+single+and+double+layered+microcapsules+from+plant+spores.">Sequestration of edible oil from emulsions using new single and double layered microcapsules from plant spores.</a> J. Mater. Chem. 22 (19), 9767-9773 (2012).</li><li>Diego-Taboada, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protein+free+microcapsules+obtained+from+plant+spores+as+a+model+for+drug+delivery:+Ibuprofen+encapsulation,+release+and+taste.">Protein free microcapsules obtained from plant spores as a model for drug delivery: Ibuprofen encapsulation, release and taste.</a> Mater. Chem. B. 1 (5), 707-713 (2013).</li><li>Atwe, S. U., Ma, Y., Gill, H. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pollen+grains+for+oral+vaccination.">Pollen grains for oral vaccination.</a> J. Control. Release. 194, 45-52 (2014).</li><li>Mackenzie, G., Beckett, S., Atkin, S., Diego-Taboada, A., Gaonkar, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ch+24.">Ch 24.</a> Microencapsulation in the Food Industry: A Practical Implementation Guide. , 283-297 (2014).</li><li>Diego-Taboada, A., Beckett, S. T., Atkin, S. L., Mackenzie, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hollow+Pollen+Shells+to+Enhance+Drug+Delivery.">Hollow Pollen Shells to Enhance Drug Delivery.</a> Pharmaceutics. 6 (1), 80-96 (2014).</li><li>Ma, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+of+a+novel+rape+pollen+shell+microencapsulation+and+its+use+for+protein+adsorption+and+pH-controlled+release.">Preparation of a novel rape pollen shell microencapsulation and its use for protein adsorption and pH-controlled release.</a> J. Microencapsul. 31 (7), 667-673 (2014).</li><li>Archibald, S. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+does+iron+interact+with+sporopollenin+exine+capsules?+An+X-ray+absorption+study+including+microfocus+XANES+and+XRF+imaging.">How does iron interact with sporopollenin exine capsules? An X-ray absorption study including microfocus XANES and XRF imaging.</a> J. Mater. Chem. B. 2 (8), 945-959 (2014).</li><li>Southworth, D., et al., Blackmore, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ch+10.">Ch 10.</a> Microspores Evolution and Ontogeny: Evolution and Ontogeny. , 193-212 (2013).</li><li>Stanley, R. G., Linskens, H. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Pollen: Biology, Biochemistry, Management. , 307 (1974).</li><li>Ariizumi, T., Toriyama, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+regulation+of+sporopollenin+synthesis+and+pollen+exine+development.">Genetic regulation of sporopollenin synthesis and pollen exine development.</a> Annu. Rev. Plant Biol. 62, 437-460 (2011).</li><li>Mundargi, R. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Natural+Sunflower+Pollen+as+a+Drug+Delivery+Vehicle.">Natural Sunflower Pollen as a Drug Delivery Vehicle.</a> Small. 12 (9), 1167-1173 (2015).</li><li>Mundargi, R. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lycopodium+Spores:+A+Naturally+Manufactured,+Superrobust+Biomaterial+for+Drug+Delivery.">Lycopodium Spores: A Naturally Manufactured, Superrobust Biomaterial for Drug Delivery.</a> Adv. Funct. Mater. 26 (4), 487-497 (2015).</li><li>Beckett, S. T., Atkin, S. L., Mackenzie, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Topical+Formulations+Containing+Sporopollenin.">Topical Formulations Containing Sporopollenin.</a> US Patent. , (2007).</li><li>Zetzsche, F., Huggler, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Untersuchungen+&#252;ber+die+Membran+der+Sporen+und+Pollen+I.+1.+Lycopodium+clavatum+L.">Untersuchungen &#252;ber die Membran der Sporen und Pollen I. 1. Lycopodium clavatum L.</a> Liebigs Ann. Chem. 461 (1), 89-109 (1928).</li><li>Zetschke, F., Kaelin, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Untersuchungen+&#252;ber+die+membran+der+sporen+und+pollen+v.+4.+Zur+autoxydation+der+sporopollenine.">Untersuchungen &#252;ber die membran der sporen und pollen v. 4. Zur autoxydation der sporopollenine.</a> Helv. Chim. Acta. 14 (1), 517-519 (1931).</li><li>Zetzsche, F., Vicari, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Untersuchungen+&#252;ber+die+Membran+der+Sporen+und+Pollen+III.+2.+Picea+orientalis+Pinus+silvestris+L.,+Corylus+Avellana+L.">Untersuchungen &#252;ber die Membran der Sporen und Pollen III. 2. Picea orientalis Pinus silvestris L., Corylus Avellana L.</a> Helv. Chim. Acta. 14 (1), 62-67 (1931).</li><li>Zetzsche, F., Kalt, P., Liechti, J., Ziegler, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zur+Konstitution+des+Lycopodium-Sporonins,+des+Tasmanins+und+des+Lange-Sporonins.+XI.+Mitteilung+&#252;ber+die+Membran+der+Sporen+und+Pollen.">Zur Konstitution des Lycopodium-Sporonins, des Tasmanins und des Lange-Sporonins. XI. Mitteilung &#252;ber die Membran der Sporen und Pollen.</a> Journal f&#252;r Praktische Chemie. 148 (9-10), 267-286 (1937).</li><li>Shaw, G., Yeadon, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+studies+on+the+constitution+of+some+pollen+and+spore+membranes.">Chemical studies on the constitution of some pollen and spore membranes.</a> Grana. 5 (2), 247-252 (1964).</li><li>Shaw, G., Yeadon, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+studies+on+the+constitution+of+some+pollen+and+spore+membranes.">Chemical studies on the constitution of some pollen and spore membranes.</a> J. Chem. Soc. C Org. , 16-22 (1966).</li><li>Dom&#236;nguez, E., Mercado, J. A., Quesada, M. A., Heredia, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pollen+sporopollenin:+degradation+and+structural+elucidation.">Pollen sporopollenin: degradation and structural elucidation.</a> Sex. Plant Reprod. 12 (3), 171-178 (1999).</li><li>Mundargi, R. C., Tan, E. L., Seo, J., Cho, N. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Encapsulation+and+controlled+release+formulations+of+5-fluorouracil+from+natural+Lycopodium+clavatum+spores.">Encapsulation and controlled release formulations of 5-fluorouracil from natural Lycopodium clavatum spores.</a> J. Ind. Eng. Chem. 36, 102-108 (2016).</li><li>Orhan, I., K&#252;peli, E., &#350;ener, B., Yesilada, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Appraisal+of+anti-inflammatory+potential+of+the+clubmoss,+Lycopodium+clavatum+L.">Appraisal of anti-inflammatory potential of the clubmoss, Lycopodium clavatum L.</a> J. Ethnopharmacol. 109 (1), 146-150 (2007).</li><li>Baytop, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Therapy with medicinal plants in Turkey. , 334-335 (1999).</li><li>Pathak, S., Banerjee, A., Paul, S., Khuda-Bukhsh, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protective+potentials+of+a+plant+extract+(Lycopodium+clavatum)+on+mice+chronically+fed+hepato-carcinogens.">Protective potentials of a plant extract (Lycopodium clavatum) on mice chronically fed hepato-carcinogens.</a> Indian J. Exp. Biol. 47 (7), 602-607 (2009).</li><li>Ma, X., Gang, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+lycopodium+alkaloids.">The lycopodium alkaloids.</a> Nat. Prod. Rep. 21 (6), 752-772 (2004).</li><li>Bishayee, K., Chakraborty, D., Ghosh, S., Boujedaini, N., Khuda-Bukhsh, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lycopodine+triggers+apoptosis+by+modulating+5-lipoxygenase,+and+depolarizing+mitochondrial+membrane+potential+in+androgen+sensitive+and+refractory+prostate+cancer+cells+without+modulating+p53+activity:+signaling+cascade+and+drug-DNA+interaction.">Lycopodine triggers apoptosis by modulating 5-lipoxygenase, and depolarizing mitochondrial membrane potential in androgen sensitive and refractory prostate cancer cells without modulating p53 activity: signaling cascade and drug-DNA interaction.</a> Eur. J. Pharmacol. 698 (1), 110-121 (2013).</li><li>Durdun, C., Papuc, C., Crivineanu, M., Nicorescu, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antioxidant+potential+of+Lycopodium+clavatum+and+Cnicus+benedictus+hydroethanolic+extracts+on+stressed+mice.">Antioxidant potential of Lycopodium clavatum and Cnicus benedictus hydroethanolic extracts on stressed mice.</a> Scientific Works-University of Agronomical Sciences and Veterinary Medicine, Bucharest Series C, Veterinary Medicine. 57 (3), 61-68 (2011).</li><li>Banerjee, J., Biswas, S., Madhu, N. R., Karmakar, S. R., Biswas, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+better+understanding+of+pharmacological+activities+and+uses+of+phytochemicals+of+Lycopodium+clavatum:+A+review.">A better understanding of pharmacological activities and uses of phytochemicals of Lycopodium clavatum: A review.</a> J. Pharmacogn. Phytochem. 3 (1), 207-210 (2014).</li><li>Mundargi, R. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extraction+of+sporopollenin+exine+capsules+from+sunflower+pollen+grains.">Extraction of sporopollenin exine capsules from sunflower pollen grains.</a> RSC Adv. 6 (20), 16533-16539 (2016).</li><li>Mathias, P. M., Chen, C. C., Walters, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modeling+Polyethylene+Fractionation+Using+the+Statistical+Associating+Fluid+Theory.">Modeling Polyethylene Fractionation Using the Statistical Associating Fluid Theory.</a> , (2000).</li><li>El-Bayaa, A., Badawy, N., Gamal, A., Zidan, I., Mowafy, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Purification+of+wet+process+phosphoric+acid+by+decreasing+iron+and+uranium+using+white+silica+sand.">Purification of wet process phosphoric acid by decreasing iron and uranium using white silica sand.</a> J. Hazar. Mater. 190 (1), 324-329 (2011).</li><li>Carr, J., Zhang, L., Davis, M., Ravishankar, S., Flieg, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scale+Controlling+Chemical+Additives+for+Phosphoric+Acid+Production+Plants.">Scale Controlling Chemical Additives for Phosphoric Acid Production Plants.</a> Procedia Engineering. 83, 233-242 (2014).</li><li>Mundargi, R. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Eco-friendly+streamlined+process+for+sporopollenin+exine+capsule+extraction.">Eco-friendly streamlined process for sporopollenin exine capsule extraction.</a> Sci. Rep. 6, 1-14 (2016).</li><li>Smith, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Fundamentals of Fourier transform infrared spectroscopy. , 182 (2011).</li></ol>

Yazarlar ifşa hiçbir şey yok.

Bu çalışmada, L. SEC çıkarıldığı sistematik analizinde clavatum sporlar sunulmaktadır ve bu rapor ayrıca, yüksek işlem sıcaklığına (180 ° C) gerektiren, mevcut protokolü aksine, önceden var olan, yaygın olarak kullanılan protokol. 11, bir önemli ölçüde kolaylaştırılmasını gerçekleştirirken daha kaliteli kapsüller üretmek mümkün olduğunu göstermektedir ve uzun bir süreç süresi (7 gün), 11 mevcut SEC çıkarma işleme optimizasyonu öncelik...

Orada mikrokapsülleme uygulamalarında kullanılmak üzere bitki sporlarının ve polenlerin elde edilen doğal bitki bazlı kapsüller önemli ilgi. Olan Doğada 1-15, sporlar ve polen sert çevre koşullarına karşı duyarlı genetik malzemeler için koruma sağlar. Bitki sporlar ve polen temel yapısı tipik olarak bir dış kabuk tabaka (ekzin), bir iç muhafaza tabakası (intine), ve iç sitoplazmik malzeme içerir. Ekzin sporopolleninin olarak ifade edilen bir kimyasal sağlam bir biyopolimerin 1,9,10,13,16 oluşur ve intine, öncelikle, selülozik malzeme oluşur. 16-18 boş kapsüller çeşitli işlemler 7,9 ile izole edilebilir sitoplazmik malzemenin çıkarılması için proteinler ve intine katmanı. 2,12,16 Bu Sporopollenin ekzin kapsüller (sn) nedeniyle dar bir büyüklük dağılımı ve homojen morfolojisi sentetik enkapsülantların için çekici bir alternatif sağlar. 7,9,13,19,20Bu Lycopodium clavatum gibi çeşitli bitki türlerinden saniye elde etmek üzere standart işlemlerinin geliştirilmesi, ilaç verme, gıda ve kozmetik alanlarında mikrokapsülleme geniş bir uygulama yelpazesi için potansiyel açmaktadır. 6,10-13,21 Sn elde etmek amacıyla, araştırmacılar, ilk organik çözücüler ile sporlar ve polen muamele edilmiş ve sitoplazmik içeriğini çıkarmak için alkalin çözeltiler içinde geri akışa alınmıştır. 22-25 Ancak, geride kalan kapsülün yapısı hala selülozik intine tabaka ihtiva ettiği belirlendi. Bu çıkarmak için, araştırmacılar, SEC intine çıkarılması tercih edilen bir yöntem haline fosforik asit, birkaç gün boyunca 22-25 hidroklorik asit, sıcak sülfürik asit, ya da sıcak fosforik asit ile uzun asidik hidroliz işlemi araştırdık. 2 Bununla birlikte, devam eden yılda araştırma, çeşitli sporlar ve polenler beni işleme sert karşı direnç de¤iflik derecelerde olduğunu göstermiştirthods yaygın olarak kullanılan. 26,27 Bazı sporlar ve polenler tamamen bozulmuştur ve kuvvetli alkali çözeltiler içinde tüm yapısal bütünlüğünü kaybeden veya ağır güçlü asidik çözeltiler içinde hasar. yapan 16 işleme koşulları SEC yanıt olarak değişkenlik kimyasal önemsiz varyasyonlar nedeniyle yapı ve türler arasındaki Sporopollenin ekzin malzemenin ekzin morfolojisi. 28 nedeniyle Sporopollenin ekzin kapsüller (sn) sağlamlığına değişkenlik nedeniyle, spore ve polen her tür için işleme koşullarını optimize etmek için gereklidir. L. türünden bitki sporlar clavatum saniyelik en çok çalışılan tek kaynağı haline gelmiş. . Bunun nedeni yaygın kullanılabilirlik, düşük maliyet, monodispersiteye ve kimyasal dayanıklılık öncelikle olduğu önerilmiştir 9,29 sporların rahatlıkla 1 gruplaşmaların şeklinde sporoplasmic içeriği hasat ve ihtiva edilebilir - 2 mikron hücre organellerinin ve biomolecules. 11 L. clavatum sporlar terapötik geniş bir uygulama yelpazesi için doğal bir toz yağlama maddesinin, kozmetikler için 30,31 bir baz olarak 30 ve bitkisel ilaç 32-36 kullanılmıştır. L. elde SEC sn clavatum sporlar ve işlendikten sonra polen. 2 diğer türlerden saniyeden daha fazla işleme karşı daha dayanıklı olduğu gösterilmiştir, elde edilen SEC sn Komplike microridge yapıları ve yüksek morfolojik düzenlilik kapsüllenmesi için büyük bir iç boşluğu sağlarken korumak için gösterilmiştir. 7 çalışmalar göstermektedir L. clavatum SEC sn ilaçlar, 10,13 aşılar, 11 protein, 7,14 hücreleri, 8 yağlar, 5-7,9 ve gıda takviyeleri kapsüllenmesi için kullanılabilir. 5,15 Gözlemlenen SEC yükleme verimleri, geleneksel ile karşılaştırıldığında nispeten yüksektir kapsülleme malzemeleri. 7 de rapor faydaları vardırSEC kapsülleme böyle, 6,10 zevke maskelemek için ve oksidasyona karşı doğal bir koruma bir dereceye sağlamak. Mevcut çalışmalarda 12 yetenek, L. için en sık kullanılan SEC ekstraksiyon yöntemi olarak clavatum dört ana adımlar dayanmaktadır. Birinci adım sporlar yağdan arındırmak için, 50 ° C &#39;de en fazla 12 saat boyunca, aseton solvent geri akan. 11 adım sitoplazma ve proteinli maddelerin çıkarılması için 120 ° C&#39;ye kadar 12 saat boyunca% 6, potasyum hidroksit, alkalin geri akan. 11 Adım üç 180 ° C&#39;de en fazla 7 gün boyunca,% 85 fosforik asit içinde asit geri akış selülozik intine malzeme kaldırmaktır. 11 Dördüncü adım kalan olmayan ekzin malzemeyi çıkarmak için, su, çözücü maddeler, asitler ve bazlar kullanılarak kapsamlı yıkama işlemi olup ve kimyasal artıklar. kapsülleme uygulamalara ilişkin SEC çıkarma ana hedefleri, ücretsiz fro sitoplazmik malzemenin boş kapsül üretmek için vardırPotansiyel alerjik proteinler m ve morfolojik olarak bozulmamış. 2,37 Ancak, sanayi üretim açısından, tür enerji verimliliği, üretim süresi, güvenlik, ve ortaya çıkan atık gibi ek ekonomik ve çevresel faktörleri dikkate almak da önemlidir. enerji verimliliği ile ilgili olarak, yüksek sıcaklıklar ve uzun işlem süreleri hem üretim maliyetlerini yanı sıra çevresel ayak izi etkiler. Üretim süresi ve gerçekleştirme süresi işleme karlılığı etkileyen en önemli faktörlerdir. Özellikle kaygı yüksek sıcaklık fosforik asit işleme güvenlik sorunları artırır ve toplu dönüş zamanlarında önemli altyapı bakım artışları ve gecikmelere yol açan aşındırıcı ölçekleme neden olduğu bilinmektedir olmasıdır. 38-40 Mümkünse, yol açabilir gerekli adımların sayısını en aza indirmek önemli ölçüde üretilen atık azaltılması. Ancak, yaygın L. dört aşamalı süreç kullanılan clavatum SEC çıkarma sadece EV vardırAraştırmanın on yıllardır olved ve küçük gerçek proses optimizasyonu olmuştur. Son zamanlarda, Mundargi ve ark., 41 sistematik değerlendirilmesi ve en sık bildirilen SEC çıkarma tekniklerinden biri optimize ederek bu alanda devam eden çalışmalara önemli bir katkı yaptı. Bu çalışmada ilk bölümünde: spor yağsızlaşma 6 saat 50 ° C&#39;de aseton işleme kullanılarak gösterilmiştir; sporoplasm ve intine kaldırma işlemleri 30 saat boyunca 70 ° C&#39;de% 85 fosforik asit işleme kullanılarak gösterilmiştir; Su, çözücüler, asit ve baz ile geniş bir yıkama bakiye sporoplasmic içeriğinin kaldırılması göstermek için kullanılır; ve SEC kurutma konveksiyon kurutma ve vakum kurutma fırını kullanılarak gösterilmiştir. Üçüncü bölümde, SEC vakum yükleme, bir model protein vakum yükleme kullanılarak araştınlır, ardından bovin serum albümini (BSA), yıkama ve liyofilizasyon BSA-sek yüklendi. Dördüncü bölümde, determinBSA kapsülleme verimlilik ation sonication soruşturma, santrifüj kullanılarak ve UV / Vis spektrometresi gösterilmiştir.

<table><tbody><tr><td>&lt;em&gt;Lycopodium clavatum&lt;/em&gt; spores (s-type)</td><td>Sigma</td><td>19108-500G-F</td><td></td></tr><tr><td>Bovine serum albumin</td><td>Sigma</td><td>A2153-50G</td><td></td></tr><tr><td>FITC-conjugated BSA</td><td>Sigma</td><td>A9771-250MG</td><td></td></tr><tr><td>Phosphoric acid (85% w/v)</td><td>Sigma</td><td>438081-2.5L</td><td></td></tr><tr><td>Hydrochloric acid</td><td>Sigma</td><td>V800202&amp;nbsp;</td><td></td></tr><tr><td>Sodium hydroxide</td><td>Sigma</td><td>S5881-1KG&amp;nbsp;</td><td></td></tr><tr><td>Acetone</td><td>Sigma</td><td>V800022</td><td></td></tr><tr><td>Ethanol</td><td>AcME&amp;nbsp;</td><td>C000356</td><td></td></tr><tr><td>Deionized water</td><td>Millipore purified water&amp;nbsp;</td><td></td><td></td></tr><tr><td>Qualitative filter paper (grade No. 1, cotton cellulose)</td><td></td><td></td><td></td></tr><tr><td>Polystyrene microspheres (50 &amp;plusmn; 1 &amp;micro;m)&amp;nbsp;</td><td>Thermoscientific (CA, USA)</td><td>4250A</td><td></td></tr><tr><td>Vectashield&amp;nbsp;</td><td>Vector labs (CA, USA)</td><td>H-1000</td><td></td></tr><tr><td>Sticky-slides, D 263 M Schott glass, No.1.5H (170 &amp;mu;m, 25 mm x 75 mm) unsterile glass slide</td><td>Ibidi GmbH (Munich, Germany)</td><td>10812</td><td></td></tr><tr><td>Commercial Lycopodium SECs (L-type)</td><td>Polysciences, Inc. (PA, USA)</td><td>16867-1</td><td></td></tr><tr><td>Heating plates</td><td>IKA, Germany</td><td></td><td></td></tr><tr><td>Scanning electron microscope</td><td>Jeol, Japan</td><td>&amp;nbsp;JFC-1600</td><td></td></tr><tr><td>Elemental analyzer&amp;nbsp;</td><td>Elementar, Germany</td><td>VarioEL III</td><td></td></tr><tr><td>FlowCam: The benchtop system</td><td>Fluid Imaging Technologies, USA</td><td>FlowCamVS</td><td></td></tr><tr><td>Confocal laser scanning microscope</td><td>Carl Zeiss, Germany</td><td>LSM710</td><td></td></tr><tr><td>Freeze dryer</td><td>Labconco, USA&amp;nbsp;</td><td></td><td></td></tr><tr><td>UV Spectrometer</td><td>Boeco, Germany</td><td>S220</td><td></td></tr></tbody></table>

extraction of plant-based capsules for microencapsulation applications

Bitki bazlı sporlar veya polen elde edilen mikrokapsüller mikroenkapsülasyon uygulamaları çeşitli bir yelpazede için sağlam bir platform sağlar. İç sporoplasmic içeriğini kaldırmak üzere sporlar ve polen işlenirken Sporopollenin ekzin kapsüller (sn) elde edilir. Elde edilen içi boş mikro kapsüller Mikromeritik muntazamlık yüksek derecede sergilerler ve özellikle bitki türlerine ilişkin karmaşık mikro özelliklerini korumaktadır. Burada, Lycopodium clavatum sporlar saniye üretimi için, bu saniye hidrofilik bileşiklerin yüklenmesi için düzenlenmiş bir işlemi gösterilmektedir. Geçerli SEC izolasyon prosedürü son zamanlarda önemli ölçüde geleneksel SEC izolasyon kullanılan işleme gereksinimlerini azaltmak için, ve bozulmamış mikrokapsüllerin üretimini sağlamak için optimize edilmiştir. Doğal L. clavatum sporlar fosforik asit ile muamele edilmiş, aseton ile yağdan arındırılmış, ve büyük ölçüde sporoplasmi uzaklaştırmak için yıkanırc içeriği. aseton yağların yok sonra% 85 fosforik asit kullanılarak tek bir işlem adımı, tüm sporoplasmic içeriğini çıkarmak için gösterilmiştir. 30 saat asit işlem süresini sınırlayarak, temiz saniye izole etmek ve uzun süreli işlem süresi ile meydana gösterilmiştir SEC kırılma önlemek mümkündür. su ile iyice yıkamak, asitler seyreltik bazlar seyreltilmiş ve solvenler her sporoplasmic madde ve kimyasal artıklar yeterli kaldırılır sağlar. vakum yükleme tekniği Hidrofilik bileşik olarak bir model proteini (Sığır Serumu Albümini) yüklemek için kullanılır. Vakum yükleme genellikle diğer mikro kapsül protokollerinde gerekli olan sert solventler veya istenmeyen kimyasallar için gerek kalmadan çeşitli bileşikler yüklemek için basit bir teknik sağlar. Bu izolasyon ve yükleme iletişim kurallarına göre, SEC sn bir mikrokapsülleme gibi uygulamalar, terapötik, gıda, kozmetik çok çeşitli kullanım için umut verici bir madde, ve kişisel bakım eşya teminfuarları.

Sporopollenin ekzin Kapsülleri için Aerodinamik Ekstraksiyon Süreci L. clavatum SEC ekstraksiyon üç ana aşamalar ile elde edildi: (1) aseton kullanılarak yağlarını; (2) asidoliz fosforik asit% 85 ile (h / h); ve (3) Kapsamlı SEC yıkama solventler kullanılarak. Akıcı SEK ekstraksiyon işleminin akış Şekil 1, bir sunulmuştur -. Kısaca, işlem 50 ° C...

Watch this Scientific Journal Video about Extraction of Plant-based Capsules for Microencapsulation Applications at JoVE.com

Extraction of Plant-based Capsules for Microencapsulation Applications

Bu protokol / Eser Lycopodium clavatum sporlar Sporopollenin ekzin kapsüller (sn) üretimi için, bu saniye hidrofilik bileşiklerin yüklenmesi için düzenlenmiş bir işlem açıklanmaktadır.

Mikroenkapsülasyon Uygulamaları için Ekstraksiyon Tesisi tabanlı Kapsüller

Microcapsules derived from plant-based spores or pollen provide a robust platform for a diverse range of microencapsulation applications. Sporopollenin ...

extraction-of-plant-based-capsules-for-microencapsulation-applications

Research

JoVE Journal

Bioengineering

11.9K Görüntüleme.  Nanyang Technological University.  Bu protokol / Eser Lycopodium clavatum sporlar Sporopollenin ekzin kapsüller (sn) üretimi için, bu saniye hidrofilik bileşiklerin yüklenmesi için düzenlenmiş bir işlem açıklanmaktadır. 

Video: Extraction of Plant-based Capsules for Microencapsulation Applications

Preparation of Giant Vesicles Encapsulating Microspheres by Centrifugation of a Water-in-oil Emulsion

The constructive biology and the synthetic biology approach to creating artificial life involve the bottom-up assembly of biological or nonbiological materials. Such approaches have received considerable attention in research on the boundary between living and nonliving matter and have been used to construct artificial cells over the past two decades. In particular, Giant Vesicles (GVs) have often been used as artificial cell membranes. In this paper, we describe the preparation of GVs encapsulating highly packed microspheres as a model of cells containing highly condensed biomolecules. The GVs were prepared by means of a simple water-in-oil emulsion centrifugation method. Specifically, a homogenizer was used to emulsify an aqueous solution containing the materials to be encapsulated and an oil containing dissolved phospholipids, and the resulting emulsion was layered carefully on the surface of another aqueous solution. The layered system was then centrifuged to generate the GVs. This powerful method was used to encapsulate materials ranging from small molecules to microspheres.

Giant vesicles containing highly packed micrometer-sized components are useful cell models. The water-in-oil emulsion centrifugation method is a simple, powerful tool for the preparation of giant vesicles with encapsulated materials.

bir su-içinde-yağ emülsiyonu santrifüj Dev veziküller Kapsüllenen Mikrokürelerin hazırlanması

The constructive biology and the synthetic biology approach to creating artificial life involve the bottom-up assembly of biological or nonbiological ...

Biyokimya

Alginate Encapsulation of Pluripotent Stem Cells Using a Co-axial Nozzle

Pluripotent stem cells (PS cells) are the focus of intense research due to their role in regenerative medicine and drug screening. However, the development of a mass culture system would be required for using PS cells in these applications. Suspension culture is one promising culture method for the mass production of PS cells, although some issues such as controlling aggregation and limiting shear stress from the culture medium are still unsolved. In order to solve these problems, we developed a method of calcium alginate (Alg-Ca) encapsulation using a co-axial nozzle. This method can control the size of the capsules easily by co-flowing N2 gas. The controllable capsule diameter must be larger than 500 &#181;m because too high a flow rate of N2 gas causes the breakdown of droplets and thus heterogeneous-sized capsules. Moreover, a low concentration of Alg-Na and CaCl2 causes non-spherical capsules. Although an Alg-Ca capsule without a coating of Alg-PLL easily dissolves enabling the collection of cells, they can also potentially leak out from capsules lacking an Alg-PLL coating. Indeed, an alginate-PLL coating can prevent cellular leakage but is also hard to break. This technology can be used to research the stem cell niche as well as the mass production of PS cells because encapsulation can modify the micro-environment surrounding cells including the extracellular matrix and the concentration of secreted factors.

We established a method of encapsulating pluripotent stem cells (PS cells) into alginate hydrogel capsules using a co-axial nozzle. This prevents cells from aggregating excessively and limits the shear stress experienced by cells in suspension culture. The technique is applicable to the mass production of PS cells as well as research on stem cell niche.

Bir Koaksiyel Memesini kullanma Pluripotent Kök Hücre Aljinat Encapsulation

Pluripotent stem cells (PS cells) are the focus of intense research due to their role in regenerative medicine and drug screening.  However, the ...

Gelişim Biyolojisi

High Throughput Single-cell and Multiple-cell Micro-encapsulation

Microfluidic encapsulation methods have been previously utilized to capture cells in picoliter-scale aqueous, monodisperse drops, providing confinement from a bulk fluid environment with applications in high throughput screening, cytometry, and mass spectrometry. We describe a method to not only encapsulate single cells, but to repeatedly capture a set number of cells (here we demonstrate one- and two-cell encapsulation) to study both isolation and the interactions between cells in groups of controlled sizes. By combining drop generation techniques with cell and particle ordering, we demonstrate controlled encapsulation of cell-sized particles for efficient, continuous encapsulation. Using an aqueous particle suspension and immiscible fluorocarbon oil, we generate aqueous drops in oil with a flow focusing nozzle. The aqueous flow rate is sufficiently high to create ordering of particles which reach the nozzle at integer multiple frequencies of the drop generation frequency, encapsulating a controlled number of cells in each drop. For representative results, 9.9 &mu;m polystyrene particles are used as cell surrogates. This study shows a single-particle encapsulation efficiency Pk=1 of 83.7% and a double-particle encapsulation efficiency Pk=2 of 79.5% as compared to their respective Poisson efficiencies of 39.3% and 33.3%, respectively. The effect of consistent cell and particle concentration is demonstrated to be of major importance for efficient encapsulation, and dripping to jetting transitions are also addressed. 
Introduction 
Continuous media aqueous cell suspensions share a common fluid environment which allows cells to interact in parallel and also homogenizes the effects of specific cells in measurements from the media. High-throughput encapsulation of cells into picoliter-scale drops confines the samples to protect drops from cross-contamination, enable a measure of cellular diversity within samples, prevent dilution of reagents and expressed biomarkers, and amplify signals from bioreactor products. Drops also provide the ability to re-merge drops into larger aqueous samples or with other drops for intercellular signaling studies.1,2 The reduction in dilution implies stronger detection signals for higher accuracy measurements as well as the ability to reduce potentially costly sample and reagent volumes.3 Encapsulation of cells in drops has been utilized to improve detection of protein expression,4 antibodies,5,6 enzymes,7 and metabolic activity8 for high throughput screening, and could be used to improve high throughput cytometry.9 Additional studies present applications in bio-electrospraying of cell containing drops for mass spectrometry10 and targeted surface cell coatings.11 Some applications, however, have been limited by the lack of ability to control the number of cells encapsulated in drops. Here we present a method of ordered encapsulation12 which increases the demonstrated encapsulation efficiencies for one and two cells and may be extrapolated for encapsulation of a larger number of cells.
To achieve monodisperse drop generation, microfluidic "flow focusing" enables the creation of controllable-size drops of one fluid (an aqueous cell mixture) within another (a continuous oil phase) by using a nozzle at which the streams converge.13 For a given nozzle geometry, the drop generation frequency f and drop size can be altered by adjusting oil and aqueous flow rates Qoil and Qaq. As the flow rates increase, the flows may transition from drop generation to unstable jetting of aqueous fluid from the nozzle.14 
When the aqueous solution contains suspended particles, particles become encapsulated and isolated from one another at the nozzle. For drop generation using a randomly distributed aqueous cell suspension, the average fraction of drops Dk containing k cells is dictated by Poisson statistics, where Dk = &lambda;k exp(-&lambda;)/(k!) and &lambda; is the average number of cells per drop. The fraction of cells which end up in the "correctly" encapsulated drops is calculated using Pk = (k x Dk)/&Sigma;(k' x Dk'). The subtle difference between the two metrics is that Dk relates to the utilization of aqueous fluid and the amount of drop sorting that must be completed following encapsulation, and Pk relates to the utilization of the cell sample. As an example, one could use a dilute cell suspension (low &lambda;) to encapsulate drops where most drops containing cells would contain just one cell. While the efficiency metric Pk would be high, the majority of drops would be empty (low Dk), thus requiring a sorting mechanism to remove empty drops, also reducing throughput.15
Combining drop generation with inertial ordering provides the ability to encapsulate drops with more predictable numbers of cells per drop and higher throughputs than random encapsulation. Inertial focusing was first discovered by Segre and Silberberg16 and refers to the tendency of finite-sized particles to migrate to lateral equilibrium positions in channel flow. Inertial ordering refers to the tendency of the particles and cells to passively organize into equally spaced, staggered, constant velocity trains. Both focusing and ordering require sufficiently high flow rates (high Reynolds number) and particle sizes (high Particle Reynolds number).17,18 Here, the Reynolds number Re =uDh/&nu; and particle Reynolds number Rep =Re(a/Dh)2, where u is a characteristic flow velocity, Dh [=2wh/(w+h)] is the hydraulic diameter, &nu; is the kinematic viscosity, a is the particle diameter, w is the channel width, and h is the channel height. Empirically, the length required to achieve fully ordered trains decreases as Re and Rep increase. Note that the high Re and Rep requirements (for this study on the order of 5 and 0.5, respectively) may conflict with the need to keep aqueous flow rates low to avoid jetting at the drop generation nozzle. Additionally, high flow rates lead to higher shear stresses on cells, which are not addressed in this protocol. The previous ordered encapsulation study demonstrated that over 90% of singly encapsulated HL60 cells under similar flow conditions to those in this study maintained cell membrane integrity.12 However, the effect of the magnitude and time scales of shear stresses will need to be carefully considered when extrapolating to different cell types and flow parameters. The overlapping of the cell ordering, drop generation, and cell viability aqueous flow rate constraints provides an ideal operational regime for controlled encapsulation of single and multiple cells.
Because very few studies address inter-particle train spacing,19,20 determining the spacing is most easily done empirically and will depend on channel geometry, flow rate, particle size, and particle concentration. Nonetheless, the equal lateral spacing between trains implies that cells arrive at predictable, consistent time intervals. When drop generation occurs at the same rate at which ordered cells arrive at the nozzle, the cells become encapsulated within the drop in a controlled manner. This technique has been utilized to encapsulate single cells with throughputs on the order of 15 kHz,12 a significant improvement over previous studies reporting encapsulation rates on the order of 60-160 Hz.4,15 In the controlled encapsulation work, over 80% of drops contained one and only one cell, a significant efficiency improvement over Poisson (random) statistics, which predicts less than 40% efficiency on average.12
In previous controlled encapsulation work,12 the average number of particles per drop &lambda; was tuned to provide single-cell encapsulation. We hypothesize that through tuning of flow rates, we can efficiently encapsulate any number of cells per drop when &lambda; is equal or close to the number of desired cells per drop. While single-cell encapsulation is valuable in determining individual cell responses from stimuli, multiple-cell encapsulation provides information relating to the interaction of controlled numbers and types of cells. Here we present a protocol, representative results using polystyrene microspheres, and discussion for controlled encapsulation of multiple cells using a passive inertial ordering channel and drop generation nozzle.

Combining monodisperse drop generation with inertial ordering of cells and particles, we describe a method to encapsulate a desired number of cells or particles in a single drop at kHz rates. We demonstrate efficiencies twice exceeding those of unordered encapsulation for single- and double-particle drops.

Yüksek Verim Micro-encapsulation Tek hücreli ve Çoklu-hücre

Microfluidic encapsulation methods have been previously utilized to capture cells in picoliter-scale aqueous, monodisperse drops, providing confinement ...

Biyomühendislik

A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products

Diterpenoids form a diverse class of small molecule natural products that are widely distributed across the kingdoms of life and have critical biological functions in developmental processes, interorganismal interactions, and environmental adaptation. Due to these various bioactivities, many diterpenoids are also of economic importance as pharmaceuticals, food additives, biofuels, and other bioproducts. Advanced genomics and biochemical approaches have enabled a rapid increase in the knowledge of diterpenoid-metabolic genes, enzymes, and pathways. However, the structural complexity of diterpenoids and the narrow taxonomic distribution of individual compounds in often only a single species remain constraining factors for their efficient production. Availability of a broader range of metabolic enzymes now provide resources for producing diterpenoids in sufficient titers and purity to facilitate a deeper investigation of this important metabolite group. Drawing on established tools for microbial and plant-based enzyme co-expression, we present an easily operated and customizable protocol for the enzymatic production of diterpenoids in either Escherichia coli or Nicotiana benthamiana, and the purification of desired products via silica chromatography and semi-preparative HPLC. Using the group of maize (Zea mays) dolabralexin diterpenoids as an example, we highlight how modular combinations of diterpene synthase (diTPS) and cytochrome P450 monooxygenase (P450) enzymes can be used to generate different diterpenoid scaffolds. Purified compounds can be used in various downstream applications, such as metabolite structural analyses, enzyme structure-function studies, and in vitro and in planta bioactivity experiments.

Here we present easy to use protocols for producing and purifying diterpenoid metabolites through the combinatorial expression of biosynthetic enzymes in Escherichia coli or Nicotiana benthamiana, followed by chromatographic product purification. The resulting metabolites are suitable for various studies including molecular structure characterization, enzyme functional studies, and bioactivity assays.

Diterpenoid Doğal Ürünlerin Enzimatik Üretimi ve Saflaştırılması için Özelleştirilebilir Bir Yaklaşım

Diterpenoids form a diverse class of small molecule natural products that are widely distributed across the kingdoms of life and have critical biological ...

Bacterial Cellulose Spheres that Encapsulate Solid Materials

Bacterial cellulose (BC) spheres have been increasingly researched since the popularization of BC as a novel material. This protocol presents an affordable and simple method for BC sphere production. In addition to producing these spheres, an encapsulation method for solid particles has also been identified. To produce BC spheres, water, black tea, sugar, vinegar, and bacterial culture are combined in a baffled flask and the contents are agitated. After determining the proper culture conditions for BC sphere formation, their ability to encapsulate solid particles was tested using biochar, polymer beads, and mine waste. Spheres were characterized using ImageJ software and thermal gravimetric analysis (TGA). Results indicate that spheres with 7.5 mm diameters can be made in 7 days. Adding various particles increases the average size range of the BC capsules. The spheres encapsulated 10 - 20% of their dry mass. This method shows low-cost sphere production and encapsulation that is possible with easily obtainable materials. BC spheres may be used in the future as a contaminant removal aid, controlled release fertilizer coating, or soil amendment.

This protocol presents an easy, inexpensive method of forming bacterial cellulose (BC) spheres. This biomaterial can function as an encapsulation medium for solid materials, including biochar, polymer spheres, and mine waste.

Katı Malzemeleri Kapsülleyen Bakteriyel Selüloz Küreler

Bacterial cellulose (BC) spheres have been increasingly researched since the popularization of BC as a novel material. This protocol presents an ...

Mühendislik

Mammalian Cell Encapsulation in Alginate Beads Using a Simple Stirred Vessel

Cell encapsulation in alginate beads has been used for immobilized cell culture in vitro as well as for immunoisolation in vivo. Pancreatic islet encapsulation has been studied extensively as a means to increase islet survival in allogeneic or xenogeneic transplants. Alginate encapsulation is commonly achieved by nozzle extrusion and external gelation. Using this method, cell-containing alginate droplets formed at the tip of nozzles fall into a solution containing divalent cations that cause ionotropic alginate gelation as they diffuse into the droplets. The requirement for droplet formation at the nozzle tip limits the volumetric throughput and alginate concentration that can be achieved. This video describes a scalable emulsification method to encapsulate mammalian cells in 0.5% to 10% alginate with 70% to 90% cell survival. By this alternative method, alginate droplets containing cells and calcium carbonate are emulsified in mineral oil, followed by a decrease in pH leading to internal calcium release and ionotropic alginate gelation. The current method allows the production of alginate beads within 20 min of emulsification. The equipment required for the encapsulation step consists in simple stirred vessels available to most laboratories.

This video and manuscript describe an emulsion-based method to encapsulate mammalian cells in 0.5% to 10% alginate beads which can be produced in large batches using a simple stirred vessel. The encapsulated cells can be cultured in vitro or transplanted for cellular therapy applications.

Alginat Boncuklarda Memeli Hücre Kapsüllenmesinin Basit Bir Karıştırılmış Geminin Kullanımı

Cell encapsulation in alginate beads has been used for immobilized cell culture in vitro as well as for immunoisolation in vivo. Pancreatic islet ...

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular weight metabolites in tissues of the treated host to achieve long-term survival. The semipermeable membrane allows engrafted encapsulated cells to avoid rejection by the immune system. The encapsulation procedure was designed to enable a controlled release of bioactive compounds, such as insulin, other hormones, and cytokines. Here we describe a method for encapsulation of catabolic cells, which consume lipids for heat production and energy dissipation (thermogenesis) in the intra-abdominal adipose tissue of obese mice. Encapsulation of thermogenic catabolic cells may be potentially applicable to the prevention and treatment of obesity and type 2 diabetes. Another potential application of catabolic cells may include detoxification from alcohols or other toxic metabolites and environmental pollutants.

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

Yağ Dokusu Depolarında içine Nakli Enkapsülasyon termojenik Preadipositler

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular ...

Tıp

Synthesis of Black Seed Oil-loaded-, Alginate-based, pH-sensitive Beads Using Electrospraying Technique

Black seed oil (BSO), derived from the seeds of the Nigella sativa plant, has garnered attention for its potential anti-cancer properties, particularly in the context of colon cancer. Its active compound, thymoquinone, may help inhibit cancer cell growth and induce apoptosis in colon cancer cells. Additionally, black seed oil's anti-inflammatory and antioxidant effects could contribute to a healthier gut environment, potentially reducing cancer risk. Therefore, this study synthesized pH-sensitive alginate beads to deliver BSO into the colon in a controlled-release manner without releasing the drug at pH 1.2 (stomach), thus providing a well-defined release pattern at pH 6.8. The use of electrospray technology improves process performance by making it easier to formulate small, homogeneous beads with a higher rate of swelling and diffusion in the gastrointestinal medium. 
The formulated beads were characterized by an ex-vivo mucoadhesive strength test, bead size, sphericity factor (SF), encapsulation efficiency (EE), scanning electron microscope (SEM), in vitro swelling behavior (SB), and in vitro drug release in acidic and buffer media. All these manufactured beads demonstrated modest sizes of 0.58 &plusmn; 0.01 mm and spherical shape of 0.03 &plusmn; 0.00 mm in this testing. The formulation showed promising floating and releasing properties in vitro. With a very low cumulative percentage of beads, the oil EE of 90.13% &plusmn; 0.93% was high, and the release study demonstrated more than 90% in pH 6.8 with good floating nature in the stomach. Additionally, the beads were evenly spaced throughout the intestine. The electrospraying approach used in this protocol can be reproducible, yielding consistent outcomes. Therefore, this protocol can be used for large-scale production for commercialization purposes.

A technique employing high electric voltage and a targeted, active ingredient-loaded emulsion to fabricate pH-responsive, uniform microbeads is presented.

Çörek Otu Yağı Yüklü, Aljinat Esaslı, pH Duyarlı Boncukların Elektropüskürtme Tekniği Kullanılarak Sentezi

Black seed oil (BSO), derived from the seeds of the Nigella sativa plant, has garnered attention for its potential anti-cancer properties, particularly in ...

Kimya

Preparation and Characterization of Nanoliposomes for the Entrapment of Bioactive Hydrophilic Globular Proteins

Liposome nanocapsules have been applied for many purposes in the pharmaceutical, cosmetic, and food industries. Attributes of liposomes include their biocompatibility, biodegradability, non-immunogenicity, non-toxicity, and ability to entrap both hydrophilic and hydrophobic compounds. The classical hydration of thin lipid films in an organic solvent is applied herein as a technique to encapsulate tarin, a plant lectin, in nanoliposomes. Nanoliposome size, stability, entrapment efficiency, and morphological characterization are described in detail. The nanoliposomes are prepared using 1,2-dioleoyl-sn-glycerol-3-phosphoethanolamine (DOPE), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000] (ammonium salt; DSPE-MPEG 2000), and cholesterylhemisuccinate (CHEMS) as the main constituents. Lipids are first dissolved in chloroform to obtain a thin lipid film that is subsequently rehydrated in ammonium sulfate solution containing the protein to be entrapped and incubated overnight. Then, sonication and extrusion techniques are applied to generate nanosized unilamellar vesicles. The size and polydispersity index of the nanovesicles are determined by dynamic light scattering, while nanovesicle morphology is assessed by scanning electron microscopy. Entrapment efficiency is determined by the ratio of the amount of unencapsulated protein to original amount of initially loaded protein. Homogeneous liposomes are obtained with an average size of 155 nm and polydispersity index value of 0.168. A high entrapment efficiency of 83% is achieved.

This study describes classical hydration using the thin lipid film method for nanoliposome preparation followed by nanoparticle characterization. A 47 kDa-hydrophilic and globular protein, tarin, is successfully encapsulated as a strategy to improve stability, avoid fast clearance, and promote controlled release. The method can be adapted to hydrophobic molecules encapsulation.

Biyoaktif Hidrofilik Globüler Proteinlerin Tuzakları Için Nanoliposomes Hazırlanması ve Karakterizasyonu

Liposome nanocapsules have been applied for many purposes in the pharmaceutical, cosmetic, and food industries. Attributes of liposomes include their ...

Microfluidic Fabrication of Core-Shell Microcapsules carrying Human Pluripotent Stem Cell Spheroids

Three-dimensional (3D) or spheroid cultures of human pluripotent stem cells (hPSCs) offer the benefits of improved differentiation outcomes and scalability. In this paper, we describe a strategy for the robust and reproducible formation of hPSC spheroids where a co-axial flow focusing device is utilized to entrap hPSCs inside core-shell microcapsules. The core solution contained single cell suspension of hPSCs and was made viscous by the incorporation of high molecular weight poly(ethylene glycol) (PEG) and density gradient media. The shell stream comprised of PEG-4 arm-maleimide or PEG-4-Mal and flowed alongside the core stream toward two consecutive oil junctions. Droplet formation occurred at the first oil junction with shell solution wrapping itself around the core. Chemical crosslinking of the shell occurred at the second oil junction by introducing a di-thiol crosslinker (1,4-dithiothreitol or DTT) to these droplets. The crosslinker reacts with maleimide functional groups via click chemistry, resulting in the formation of a hydrogel shell around the microcapsules. Our encapsulation technology produced 400 &#181;m diameter capsules at a rate of 10 capsules per second. The resultant capsules had a hydrogel shell and an aqueous core that allowed single cells to rapidly assemble into aggregates and form spheroids. The process of encapsulation did not adversely affect the viability of hPSCs, with &gt;95% viability observed 3 days post-encapsulation. For comparison, hPSCs encapsulated in solid gel microparticles (without an aqueous core) did not form spheroids and had &lt;50% viability 3 days after encapsulation. Spheroid formation of hPSCs inside core-shell microcapsules occurred within 48 h after encapsulation, with the spheroid diameter being a function of cell inoculation density. Overall, the microfluidic encapsulation technology described in this protocol was well-suited for hPSCs encapsulation and spheroid formation.

This article describes encapsulation of human pluripotent stem cells (hPSCs) using a co-axial flow focusing device. We demonstrate that this microfluidic encapsulation technology enables efficient formation of hPSC spheroids.