| 
							
      					 | 
  					 
  					
    					 | 
   					 
   										
    					Production of pectic extracts from sugar beet pulp with antiproliferative activity on a breast cancer cell line  | 
  					 
  					  										
						Jacqueline CONCHA1( ), Caroline WEINSTEIN1,3, María Elvira Zú?IGA2,3 | 
					 
															
						| 1. Department of Biochemistry, Pharmacy School, Universidad de Valparaíso, Gran Breta?a 1093, Playa Ancha, Valparaíso, Chile; 2. School of Biochemistry Engineering, Pontificia Universidad Católica de Valparaíso, General Cruz 34, Valparaíso, Chile; 3. Creas Conicyt-Regional Gore Región de Valparaíso R12C1001, Blanco 1623, of 1402, Valparaíso, Chile | 
					 
										
						 | 
					 
				 
				
				
					
						
							
								
									
		
		 
          
          
            
              
				
								                
													
													    | 
													    	
														 | 
													 
													
													
													
														
															
													
													    | 
													     		                            						                            																	    Abstract  In the last years, sugar beet pectins have been the subject of several investigations involving extraction methodologies, chemical composition and functional properties. The structure of pectins, which depends on the extraction method, is decisive in their capacity to induce apoptosis on several cancer cell lines like colon, prostate and breast. In this work, sugar beet pectin extraction was performed in the following steps: lipid extraction with hexane, removal of soluble complex carbohydrates and proteins, and enzymatic treatment with amyloglucosidase, protease, and pectinase. The enzymatic treatment was carried out with Rohapect DA6L under the following conditions: 50°C, pH 4.0, 2% enzyme/substrate (E/S) ratio, 15 h, and a solid to liquid ratio of 1 ∶ 10. The pectic extract showed a degree of polymerization (DP) profile of 55.8% with DP≥7; 4.9% with DP6; 5.8% between DP2 and DP6 ; 4.7% with DP2; and 28.8% with DP1. The pectic extract was examined for its antiproliferative activity on the MCF-7 breast cancer cell line. At a concentration range of 12.5–25 mg/mL the pectic extract killed 80.6% of the cells, exhibiting a higher antiproliferative activity than 4-hydroxytamoxifen (4-OHT), a classical anticancer drug, which killed 56.5% of the cells. 
																										     | 
														 
																												
												        														
															| Keywords 
																																																				pectic extracts  
																		  																																				antiproliferative activity  
																		  																																				breast cancer  
																		  																																				enzymatic treatment  
																																			  
															 | 
														 
																												
														 																											    														
															| 
																																Corresponding Author(s):
																CONCHA Jacqueline,Email:jacqueline.concha@uv.cl   
																													     		
													     	 | 
														 
																																										
															| 
																																																													Issue Date: 05 December 2013
																														 | 
														 
														 
                                                         | 
														 
														 
														
														
														
												 
												
												
                                                    
													
								             
                                             
            
					            
								            								            
								            								                                                        
								            
								                
																																												
															| 1 | 
															 
														     Oosterveld A, Beldman G, Voragen A G J. Enzymatic modification of pectic polysaccharides obtained from sugar beet pulp. Carbohydrate Polymers , 2002, 48(1): 73-81  doi: 10.1016/S0144-8617(01)00216-8
														     															 | 
																  
																														
															| 2 | 
															 
														     Oosterveld A, Beldman G, Schols H, Voragen A. Characterization of arabinose and ferulic acid rich pectic polysaccharides and hemicelluloses from sugar beet pulp. Carbohydrate Research , 2000, 328(2): 185-197  doi: 10.1016/S0008-6215(00)00095-1
														     															 | 
																  
																														
															| 3 | 
															 
														     Sun R, Hughes S. Fractional isolation and physico-chemical characterization of alkali-soluble polysaccharides from sugar beet pulp. Carbohydrate Polymers , 1999, 38(3): 273-281  doi: 10.1016/S0144-8617(98)00102-7
														     															 | 
																  
																														
															| 4 | 
															 
														     Schols H, Voragen G, Colquhoun I. Isolation and characterization of rhamnogalacturonan oligomers, liberated during degradation of pectic hairy regions by rhamnogalacturonase. Carbohydrate Research , 1994, 256(1): 97-111  doi: 10.1016/0008-6215(94)84230-2
														     															 | 
																  
																														
															| 5 | 
															 
														     Schols H, Vierhuis E, Bakx E, Voragen A. Different populations of pectic hairy regions occur in apple cell walls. Carbohydrate Research , 1995, 275(2): 343-360  doi: 10.1016/0008-6215(95)00155-M
														     															 | 
																  
																														
															| 6 | 
															 
														     Yapo B, Robert C, Etienne I, Wathelet B, Paquot M. Effect of extraction conditions on the yield, purity and surface properties of sugar beet pulp pectin extracts. Food Chemistry , 2007, 100(4): 1356-1364  doi: 10.1016/j.foodchem.2005.12.012
														     															 | 
																  
																														
															| 7 | 
															 
														     Yapo B. Pectin quantity, composition and physicochemical behaviour as influenced by the purification process. Food Research International , 2009, 42(8): 1197-1202  doi: 10.1016/j.foodres.2009.06.002
														     															 | 
																  
																														
															| 8 | 
															 
														     Yapo B. Pectic substances: From simple pectic polysaccharides to complex pectins—A new hypothetical model. Carbohydrate Polymers , 2011, 86(8): 373-385  doi: 10.1016/j.carbpol.2011.05.065
														     															 | 
																  
																														
															| 9 | 
															 
														     Yeoh S, Shi J, Langrish T. Comparisons between different techniques for water-based extraction of pectin from orange peels. Desalination , 2008, 218(1-3): 229-237  doi: 10.1016/j.desal.2007.02.018
														     															 | 
																  
																														
															| 10 | 
															 
														     Willats W, Knox J, Mikkelsen J. Pectin: New insights into an old polymer are starting to gel. Trends in Food Science & Technology , 2006, 17(3): 97-104  doi: 10.1016/j.tifs.2005.10.008
														     															 | 
																  
																														
															| 11 | 
															 
														     Micard V, Renard C M G C, Thibault J F. Enzymatic saccharification of sugar-beet pulp. Enzyme and Microbial Technology , 1996, 19(3): 162-170  doi: 10.1016/0141-0229(95)00224-3
														     															 | 
																  
																														
															| 12 | 
															 
														     Levigne S, Ralet M C, Thibault J F. Characterisation of pectins extracted from fresh sugar beet under different conditions using an experimental design. Carbohydrate Polymers , 2002, 49(2): 145-153  doi: 10.1016/S0144-8617(01)00314-9
														     															 | 
																  
																														
															| 13 | 
															 
														     Robert C, Emaga T, Wathelet B, Paquot M. Effect of variety and harvest date on pectin extracted from chicory roots (Cichorium intybus L.). Food Chemistry , 2008, 108(3): 1008-1018  doi: 10.1016/j.foodchem.2007.12.013
														     															 | 
																  
																														
															| 14 | 
															 
														     Schieber A, Hilt P, Streker P, Endre? H U, Rentschler C, Carle R. A new process for the combined recovery of pectin and phenolic compounds from apple pomace. Innovative Food Science & Emerging Technologies , 2003, 4(1): 99-107  doi: 10.1016/S1466-8564(02)00087-5
														     															 | 
																  
																														
															| 15 | 
															 
														     Westereng B, Michaelsen T, Samuelsen A, Knutsen S. Effects of extraction conditions on the chemical structure and biological activity of white cabbage pectin. Carbohydrate Polymers , 2008, 72(1): 32-42  doi: 10.1016/j.carbpol.2007.07.017
														     															 | 
																  
																														
															| 16 | 
															 
														     Koubala B, Kansci G, Mbome L, Crépeau M J, Thibault J F, Ralet M C. Effect of extraction conditions on some physicochemical characteristics of pectins from “Améliorée” and “Mango” mango peels. Food Hydrocolloids , 2008, 22(7): 1345-1351  doi: 10.1016/j.foodhyd.2007.07.005
														     															 | 
																  
																														
															| 17 | 
															 
														     Koubala B, Mbome L, Kansci G, Tchouanguep Mbiapo F, Crepeau M J, Thibault J F, Ralet M C. Physicochemical properties of pectins from ambarella peels (Spondias cytherea) obtained using different extraction conditions. Food Chemistry , 2008, 106(3): 1202-1207  doi: 10.1016/j.foodchem.2007.07.065
														     															 | 
																  
																														
															| 18 | 
															 
														     Liu Y, Shi J, Langrish T. Water-based extraction of pectin from flavedo and albedo of orange peels. Chemical Engineering Journal , 2006, 120(3): 203-209  doi: 10.1016/j.cej.2006.02.015
														     															 | 
																  
																														
															| 19 | 
															 
														     Rombouts F, Thibault J. Enzymic and chemical degradation and the fine structure of pectins from sugar-beet pulp. Carbohydrate Research , 1986, 154(1): 189-203  doi: 10.1016/S0008-6215(00)90032-6
														     															 | 
																  
																														
															| 20 | 
															 
														     Rombouts F, Thibault J. Feruloylated pectic substances from sugar-beet pulp. Carbohydrate Research , 1986, 154(1): 177-187  doi: 10.1016/S0008-6215(00)90031-4
														     															 | 
																  
																														
															| 21 | 
															 
														     Happi Emaga T, Ronkart S, Robert C, Wathelet B, Paquot M. Characterisation of pectins extracted from banana peels (Musa AAA) under different conditions using an experimental design. Food Chemistry , 2008, 108(2): 463-471  doi: 10.1016/j.foodchem.2007.10.078
														     															 | 
																  
																														
															| 22 | 
															 
														     Concha J, Zú?iga M. Enzymatic depolymerization of sugar beet pulp: Production and characterization of pectin and pectic-oligosaccharides as a potential source for functional carbohydrates. Chemical Engineering Journal , 2012, 192: 29-36  doi: 10.1016/j.cej.2012.03.085
														     															 | 
																  
																														
															| 23 | 
															 
														     Claye S, Idouraine A, Weber C. Extraction and fractionation of insoluble fiber from five fiber sources. Food Chemistry , 1996, 57(2): 305-310  doi: 10.1016/0308-8146(95)00250-2
														     															 | 
																  
																														
															| 24 | 
															 
														     Deytieux-Belleau C, Vallet A, Donèche B, Geny L. Pectin methylesterase and polygalacturonase in the developing grape skin. Plant Physiology and Biochemistry , 2008, 46(7): 638-646  doi: 10.1016/j.plaphy.2008.04.008
														     															 | 
																  
																														
															| 25 | 
															 
														     Iglesias M, Lozano J E. Extraction and characterization of sunflower pectin. Journal of Food Engineering , 2004, 62(3): 215-223  doi: 10.1016/S0260-8774(03)00234-6
														     															 | 
																  
																														
															| 26 | 
															 
														     Kulkarni S, Vijayanand P.Effect of extraction conditions on the quality characteristics of pectin from passion fruit peel (Passiflora edulis f. flavicarpa L.). LWT-Food Science and Technology , 2010, 43(7): 1026-1031 
														     															 | 
																  
																														
															| 27 | 
															 
														     Liu L, Cao J, Huang J, Cai Y, Yao J. Extraction of pectins with different degrees of esterification from mulberry branch bark. Bioresource Technology , 2010, 101(9): 3268-3273  doi: 10.1016/j.biortech.2009.12.062
														     															 | 
																  
																														
															| 28 | 
															 
														     Marry M, McCann M, Kolpak F, White A R, Stacey N J, Roberts K. Extraction of pectic polysaccharides from sugar-beet cell walls. Journal of the Science of Food and Agriculture , 2000, 80(1): 17-28  doi: 10.1002/(SICI)1097-0010(20000101)80:1<17::AID-JSFA491>3.0.CO;2-4
														     															 | 
																  
																														
															| 29 | 
															 
														     Sahari M, Akbarian M A, Hamedi M. Effect of variety and acid washing method on extraction yield and quality of sunflower head pectin. Food Chemistry , 2003, 83(1): 43-47  doi: 10.1016/S0308-8146(03)00034-7
														     															 | 
																  
																														
															| 30 | 
															 
														     Singthong J, Ningsanond S, Cui S, Goff H. Extraction and physicochemical characterization of Krueo Ma Noy pectin. Food Hydrocolloids , 2005, 19(5): 793-801  doi: 10.1016/j.foodhyd.2004.09.007
														     															 | 
																  
																														
															| 31 | 
															 
														     Zykwinska A, Boiffard M, Kontkanen H, Buchert J, Thibault J F, Bonnin E. Extraction of green labeled pectins and pectic oligosaccharides from plant byproducts. Journal of Agricultural and Food Chemistry , 2008, 56(19): 8926-8935  doi: 10.1021/jf801705a
														     															 | 
																  
																														
															| 32 | 
															 
														     Zykwinska A, Rondeau-Mouro C, Garnier C, Thibault J, Ralet M. Alkaline extractability of pectic arabinan and galactan and their mobility in sugar beet and potato cell walls. Carbohydrate Polymers , 2006, 65(4): 510-520  doi: 10.1016/j.carbpol.2006.02.012
														     															 | 
																  
																														
															| 33 | 
															 
														     Zykwinska A, Gaillard C, Boiffard M, Thibault J F, Bonnin E. “Green labelled” pectins with gelling and emulsifying properties can be extracted by enzymatic way from unexploited sources. Food Hydrocolloids , 2009, 23(8): 2468-2477  doi: 10.1016/j.foodhyd.2009.07.010
														     															 | 
																  
																														
															| 34 | 
															 
														     Sathisha U, Jayaram S, Harish Nayaka M, Dharmesh S. Inhibition of galectin-3 mediated cellular interactions by pectic polysaccharides from dietary sources. Glycoconjugate Journal , 2007, 24(8): 497-507  doi: 10.1007/s10719-007-9042-3
														     															 | 
																  
																														
															| 35 | 
															 
														     Jackson C, Dreaden T, Theobald L, Tran N M, Beal T L, Eid M, Gao M Y, Shirley R B, Stoffel M T, Kumar M V, Mohnen D. Pectin induces apoptosis in human prostate cancer cells: correlation of apoptotic function with pectin structure. Glycobiology , 2007, 17(8): 805-819  doi: 10.1093/glycob/cwm054
														     															 | 
																  
																														
															| 36 | 
															 
														     Maxwell E, Belshaw N, Waldron K, Morris V. Pectin—An emerging new bioactive food polysaccharide. Trends in Food Science & Technology , 2012, 24(2): 64-73  doi: 10.1016/j.tifs.2011.11.002
														     															 | 
																  
																														
															| 37 | 
															 
														     Kidd P. A new approach to metastatic cancer prevention: Modified citrus pectin (MCP), an unique pectin that blocks cell surface lectins. Alternative Medicine Review , 1996, 1(1): 4-10 
														     															 | 
																  
																														
															| 38 | 
															 
														     Davis C, Milner J. Gastrointestinal microflora, food components and colon cancer prevention. Journal of Nutritional Biochemistry , 2009, 20(10): 743-752  doi: 10.1016/j.jnutbio.2009.06.001
														     															 | 
																  
																														
															| 39 | 
															 
														     Glinskii O V, Huxley V H, Glinsky G V, Pienta K J, Raz A, Glinsky V V. Mechanical Entrapment is insufficient and intercellular adhesion is essential for metastasic cell arrest in distant organs. Neoplasia (New York, N.Y.) , 2005, 7(5): 522-527  doi: 10.1593/neo.04646
														     															 | 
																  
																														
															| 40 | 
															 
														     Bergman M, Djaldetti M, Salman H, Bessler H. Effect of citrus pectin on malignant cell proliferation. Biomedicine and Pharmacotherapy , 2010, 64(1): 44-47  doi: 10.1016/j.biopha.2009.03.004
														     															 | 
																  
																														
															| 41 | 
															 
														     Morra M, Cassinelli C, Cascardo G, Nagel M D, Della Volpe C, Siboni S, Maniglio D, Brugnara M, Ceccone G, Schols H A, Ulvskov P. Effects on interfacial properties and cell adhesion of surface modification by pectic hairy regions. Biomacromolecules , 2004, 5(6): 2094-2104  doi: 10.1021/bm049834q
														     															 | 
																  
																														
															| 42 | 
															 
														     Gulfi M, Arrigoni E, Amado R. Influence of structure on in vitro fermentability of commercial pectins and partially hydrolysed pectin preparations. Carbohydrate Polymers , 2005, 59(2): 247-255  doi: 10.1016/j.carbpol.2004.09.018
														     															 | 
																  
																														
															| 43 | 
															 
														     Bland E, Keshavarz T, Bucke C. The influence of small oligosaccharides on the immune system. Carbohydrate Research , 2004, 339(10): 1673-1678  doi: 10.1016/j.carres.2004.05.009
														     															 | 
																  
																														
															| 44 | 
															 
														     Garthoff J, Heemskerk S, Hempenius R. Safety evaluation of pectin-derived acidic oligosaccharides (pAOS): genotoxicity and sub-chronic studies. Regulatory toxicology and pharmacology. RTP , 2010, 57(1): 31-42 
														     															 | 
																  
																														
															| 45 | 
															 
														     Olano-Martin E, Gibson G, Rastell R. Comparison of the in vitro bifidogenic properties of pectins and pectic-oligosaccharides. Journal of Applied Microbiology , 2002, 93(3): 505-511  doi: 10.1046/j.1365-2672.2002.01719.x
														     															 | 
																  
																														
															| 46 | 
															 
														     Chauhan D, Catley L, Li G, Podar K, Hideshima T, Velankar M, Mitsiades C, Mitsiades N, Yasui H, Letai A, Ovaa H, Berkers C, Nicholson B, Chao T H, Neuteboom S T C, Richardson P, Palladino M A, Anderson K C. A novel orally active proteasome inhibitor induces apoptosis in multiple myeloma cells with mechanisms distinct from Bortezomib. Cancer Cell , 2005, 8(5): 407-419  doi: 10.1016/j.ccr.2005.10.013
														     															 | 
																  
																														
															| 47 | 
															 
														     Favela-Torres E, Volke-Sepúlveda T, Viniegra-González G. Production of Hydrolytic Depolymerising Pectinases. Production , 2006, 44(2): 221-227 
														     															 | 
																  
																														
															| 48 | 
															 
														     Miller G. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Analytical Chemistry , 1959, 31(3): 426-428  doi: 10.1021/ac60147a030
														     															 | 
																  
																														
															| 49 | 
															 
														     Dubois M, Gilles K, Hamilton J, Rebers P A, Smith F. Colorimetric method for determination of sugars and related substances. Analytical Chemistry , 1956, 28(3): 350-356 doi: 10.1021/ac60111a017
														     															 | 
																  
																														
															| 50 | 
															 
														     Skehan P, Storeng R, Scudiero D, Monks A, McMahon J, Vistica D, Warren J T, Bokesch H, Kenney S, Boyd M R. New colorimetric cytotoxicity assay for anticancer-drug screening. JNCI J Natl Cancer Inst , 1990, 82(13): 1107-1112  doi: 10.1093/jnci/82.13.1107
														     															 | 
																  
																														
															| 51 | 
															 
														     Holck J, Hjern? K, Lorentzen A, Vigsn?s L K, Hemmingsen L, Licht T R, Mikkelsen J D, Meyer A S. Tailored enzymatic production of oligosaccharides from sugar beet pectin and evidence of differential effects of a single DP chain length difference on human faecal microbiota composition after in vitro fermentation. Process Biochemistry , 2011, 46(5): 1039-1049 doi: 10.1016/j.procbio.2011.01.013
														     															 | 
																  
																														
															| 52 | 
															 
														     Gnanasambandam R. Determination of pectin degree of esterification by diffuse reflectance Fourier transforms infrared spectroscopy. Food Chemistry , 2000, 68(3): 327-332  doi: 10.1016/S0308-8146(99)00191-0
														     															 | 
																  
																														
															| 53 | 
															 
														     Bermudez G.Obtencion y caracterizacion de pectinas de alto y bajo metoxilo de la manzana, variedad Pachacamac. Revista-sociedad quimica del Perú , 2003, 69(3): 155-162 
														     															 | 
																  
																														
															| 54 | 
															 
														     Ovodov Y. Current views on pectin substances. Russian Journal of Bioorganic Chemistry , 2009, 35(3): 269-284  doi: 10.1134/S1068162009030017
														     															 | 
																  
																														
															| 55 | 
															 
														     Gullón B, Yá?ez R, Alonso J, Parajó J. Production of oligosaccharides and sugars from rye straw: A kinetic approach. Bioresource Technology , 2010, 101(17): 6676-6684  doi: 10.1016/j.biortech.2010.03.080
														     															 | 
																  
																														
															| 56 | 
															 
														     Sriamornsak P. Chemistry of pectin and its pharmaceutical uses: A review. Silpakorn University International Journal , 2003, 3(1-2): 206-228 
														     															 | 
																  
																														
															| 57 | 
															 
														     Nangia-Makker P, Hogan V, Honjo Y, Baccarini S, Tait L, Bresalier R, Raz A. Inhibition of human cancer cell growth and metastasis in nude mice by oral intake of modified citrus pectin. Journal of the National Cancer Institute , 2002, 94(24): 1854-1862  doi: 10.1093/jnci/94.24.1854
														     															 | 
																  
																														
															| 58 | 
															 
														     Sharma B, Naresh L, Dhuldhoya N.An overview on pectins. Time processing Journals , 2006, 4: 44-51
														     															 | 
																  
																														
															| 59 | 
															 
														     Ghazarian H, Idoni B, Oppenheimer S B. A glycobiology review: Carbohydrates, lectins and implications in cancer therapeutics. Acta Histochemica , 2011, 113(3): 236-247  doi: 10.1016/j.acthis.2010.02.004
														     															 | 
																  
																														
															| 60 | 
															 
														     Glinsky V, Raz A. Modified citrus pectin anti-metastatic properties: One bullet, multiple targets. Carbohydrate Research , 2009, 344(14): 1788-1791  doi: 10.1016/j.carres.2008.08.038
														     															 | 
																  
																														
															| 61 | 
															 
														     Nangia-Makker P, Conklin J, Hogan V, Raz A. Carbohydrate-binding proteins in cancer, and their ligands as therapeutic agents. Trends in Molecular Medicine , 2002, 8(4): 187-192  doi: 10.1016/S1471-4914(02)02295-5
														     															 | 
																  
																														
															| 62 | 
															 
														     Nakahara S, Oka N, Raz A. Raz. On the role of galectin-3 in cancer apoptosis. Apoptosis. An International Journal on Programmed Cell Death , 2005, 10(2): 267-275  doi: 10.1007/s10495-005-0801-y
														     															 | 
																  
																														
															| 63 | 
															 
														     Danguy A, Camby I, Kiss R. Galectins and cancer. Biochimica et Biophysica Acta , 2002, 1572(2-3): 285-293  doi: 10.1016/S0304-4165(02)00315-X
														     															 | 
																  
																														
															| 64 | 
															 
														     Nangia-Makker P, Balan V, Raz A. Regulation of tumor progression by extracellular galectin-3. Cancer Microenvironment. Official Journal of the International Cancer Microenvironment Society , 2008, 1(1): 43-51  doi: 10.1007/s12307-008-0003-6
														     															 | 
																  
																																										 
								             
                                             
								                                                        
                                            
                                            
								                                                        
                                            
                                            
                                            
								            
												
											    	
											        	 | 
											        	Viewed | 
											         
													
											        	 | 
											        	 | 
											         
											      	
												         | 
												        
												        	Full text 
												          	
												         | 
											        	
												        	
												        	 
												        	
												          	 
												          	
												          	
														 | 
													 
													
												         | 
												         | 
													 
													
												         | 
												        
												        	Abstract 
												          	
														 | 
												        
															
															 
															
															
												         | 
													 
													
												         | 
												         | 
													 
													
												         | 
												        Cited  | 
												        
												        	
												         | 
													 
													
												         | 
												         | 
												         | 
													 
													
													    |   | 
													    Shared | 
													       | 
												  	 
												  	
													     | 
													     | 
													     | 
											  		 
											  		
													    |   | 
													    Discussed | 
													       | 
												  	 
											 
											 
								         
                                        
  
									 | 
								 
							 
						 | 
					 
				 
			
		 |