1、Designation: F2103 11Standard Guide forCharacterization and Testing of Chitosan Salts as StartingMaterials Intended for Use in Biomedical and Tissue-Engineered Medical Product Applications1This standard is issued under the fixed designation F2103; the number immediately following the designation ind
2、icates the year oforiginal adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.INTRODUCTIONBiopolymers from marine sources have been
3、 studied and used in commercial applications andproduct development for a number of years. Chitosan, a linear polysaccharide consisting ofglucosamine and N-acetyl glucosamine derived mainly from crustacean shells, has been used in manytechnical applications such as water purification (as a flocculan
4、t), in cosmetics, and recently as aproposed fat-binding weight control product. In solution, the cationic nature of chitosan gives thispolymer a mucoadhesive property. Chitosan salts can be used as a matrix or scaffold material as wellas in non-parenteral delivery systems for challenging drugs. Chit
5、osan salts have been shown toincrease the transport of polar drugs across the nasal epithelial surface. The purpose of this guide isto identify key parameters relevant for the functionality and characterization of chitosan salts for thedevelopment of new commercial applications of chitosan salts for
6、 the biomedical and pharmaceuticalindustries.1. Scope1.1 This guide covers the evaluation of chitosan saltssuitable for use in biomedical or pharmaceutical applications,or both, including, but not limited to, tissue-engineered medi-cal products (TEMPS).1.2 This guide addresses key parameters relevan
7、t for thefunctionality, characterization, and purity of chitosan salts.1.3 As with any material, some characteristics of chitosanmay be altered by processing techniques (such as molding,extrusion, machining, assembly, sterilization, and so forth)required for the production of a specific part or devi
8、ce.Therefore, properties of fabricated forms of this polymershould be evaluated using test methods that are appropriate toensure safety and efficacy.1.4 WarningMercury has been designated by EPA andmany state agencies as a hazardous material that can causecentral nervous system, kidney, and liver da
9、mage. Mercury, orits vapor, may be hazardous to health and corrosive tomaterials. Caution should be taken when handling mercury andmercury-containing products. See the applicable product Ma-terial Safety Data Sheet (MSDS) for details and EPAs website(http:/www.epa.gov/mercury/faq.htm) for additional
10、 informa-tion. Users should be aware that selling mercury or mercury-containing products, or both, in your state may be prohibited bystate law.1.5 The values stated in SI units are to be regarded asstandard. No other units of measurement are included in thisstandard.1.6 This standard does not purpor
11、t to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:2D2196
12、 Test Methods for Rheological Properties of Non-Newtonian Materials by Rotational (Brookfield type) Vis-cometerF619 Practice for Extraction of Medical PlasticsF748 Practice for Selecting Generic Biological Test Meth-ods for Materials and DevicesF749 Practice for Evaluating Material Extracts by Intra
13、cu-taneous Injection in the Rabbit1This guide is under the jurisdiction of ASTM Committee F04 on Medical andSurgical Materials and Devices and is the direct responsibility of SubcommitteeF04.42 on Biomaterials and Biomolecules for TEMPs.Current edition approved March 1, 2011. Published March 2011. O
14、riginallyapproved in 2001. Last previous edition approved in 2007 as F2103 01(2007)2.DOI: 10.1520/F2103-11.2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the stand
15、ards Document Summary page onthe ASTM website.1Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.F756 Practice for Assessment of Hemolytic Properties ofMaterialsF763 Practice for Short-Term Screening of Implant Materi-alsF813 Practice f
16、or Direct Contact Cell Culture Evaluation ofMaterials for Medical DevicesF895 Test Method for Agar Diffusion Cell Culture Screen-ing for CytotoxicityF981 Practice for Assessment of Compatibility of Biomate-rials for Surgical Implants with Respect to Effect ofMaterials on Muscle and BoneF1251 Termino
17、logy Relating to Polymeric Biomaterials inMedical and Surgical DevicesF1439 Guide for Performance of Lifetime Bioassay for theTumorigenic Potential of Implant MaterialsF1903 Practice for Testing For Biological Responses toParticles In VitroF1904 Practice for Testing the Biological Responses toPartic
18、les in vivoF1905 Practice For Selecting Tests for Determining thePropensity of Materials to Cause Immunotoxicity3F1906 Practice for Evaluation of Immune Responses InBiocompatibility Testing Using ELISA Tests, LymphocyteProliferation, and Cell Migration32.2 Ph. Eur. Document:Ph. Eur. Monograph Chitos
19、an Chloride, Nov. 200042.3 ISO Documents:ISO 10993 Biological Evaluation of Medical Devices5ISO 10993-1 Biological Evaluation of Medical DevicesPart 1: Evaluation and Testing5ISO 10993-3Part 3: Tests for Genotoxicity, Carcinogenic-ity and Reproductive Toxicity5ISO 10993-9Part 9: Framework for Identi
20、fication andQuantification of Potential Degradation Products5ISO 10993-17Part 17: Methods for Establishment ofAllowable Limits for Leachable Substances Using Health-Based Risk Assessment5ISO 13408-1: 1998: Aseptic Processing of Health CareProductsPart 1: General Requirements52.4 ICH Documents:Intern
21、ational Conference on Harmonization (1997) Guid-ance for Industry M3 Nonclinical Safety Studies for theConduct of Human Clinical Trials for Pharmaceuticals 62FR 629226International Conference on Harmonization (1996) Guide-line for Industry S2A Specific Aspects of RegulatoryGenotoxicity Tests for Pha
22、rmaceuticals 61 FR 181996International Conference on Harmonization (1997) Guid-ance for Industry S2B Genotoxicity: A Standard Batteryfor Genotoxicity Testing of Pharmaceuticals 62 FR624726International Conference on Harmonization (1994) Guide-line for Industry S5A Detection of Toxicity to Reproduc-t
23、ion for Medicinal Products 59 FR 487466International Conference on Harmonization (1996) Guid-ance for Industry S5B Detection of Toxicity to Reproduc-tion for Medicinal Products: Addendum on Toxicity toMale Fertility 61 FR 153606International Conference on Harmonization (1996) Guide-line for Industry
24、 S1A The Need for Long-term RodentCarcinogenicity Studies of Pharmaceuticals 61 FR 81536International Conference on Harmonization (1998) Guid-ance for Industry S1B Testing for Carcinogenicity ofPharmaceuticals 63 FR 89836International Conference on Harmonization (1995) Guide-line for Industry S1C Do
25、se Selection for CarcinogenicityStudies of Pharmaceuticals 60 FR 112786International Conference on Harmonization (1997) S1CRGuidance for Industry Addendum to Dose Selection forCarcinogenicity Studies of Pharmaceuticals: Addition of aLimit Dose and Related Notes 62 FR 642596International Conference o
26、n Harmonization (ICH) Q1A ICHHarmonized Tripartite Guidance for Stability Testing ofNew Drug Substances and Products (September 23,1994)62.5 FDA Documents:FDA Guideline on Validation of the Limulus AmebocyteTest as an End-Product Endotoxin Test for Human andAnimal Parenteral Drugs, Biological Produc
27、ts and Health-care Products DHHS, December 19877FDA Interim Guidance for Human and Veterinary DrugProducts and Biologicals. Kinetic LAL Techniques-DHHS, July 15, 199172.6 ANSI Documents:ANSI/AAMI/ISO 11737-1: 1995 Sterilization of MedicalDevicesMicrobiological MethodsPart 1: Estimationof Bioburden o
28、n Product5ANSI/AAMI/ISO 11737-2: 1998 Sterilization of MedicalDevicesMicrobiological MethodsPart 2: Tests of Ste-rility Performed in the Validation of a Sterilization Pro-cess52.7 AAMI Documents:AAMI TIR No. 191998: Guidance for ANSI/AAMI/ISO109937: 1995, Biological Evaluation of MedicalDevicesPart
29、7: Ethylene Oxide Sterilization Residuals8AAMI/ISO 141601998: Sterilization of Single-UseMedical Devices Incorporating Materials of AnimalOriginValidation and Routine Control of Sterilizationby Liquid Chemical Sterilants8AAMI ST67/CDV-2: 1999: Sterilization of MedicalDevicesRequirements for Products
30、 Labeled “Sterile”82.8 EN Documents:EN 12442-1 Animal Tissues and Their Derivative Utilizedin the Manufacture of Medical DevicesPart 1: Analysis3Withdrawn. The last approved version of this historical standard is referencedon www.astm.org.4Available from EDQM, Publications and Services European Phar
31、macopoeia,BP 907 226, avenue de Colmar, F-67029 Strasbourg Cedex 1, France.5Available from American National Standards Institute (ANSI), 25 W. 43rd St.,4th Floor, New York, NY 10036, http:/www.ansi.org.6Available from ICH Secretariat, c/o IFPMA, 30 rue de St-Jean, PO Box 758,1211 Geneva 13, Switzerl
32、and.7Available from Food and Drug Administration (FDA), 5600 Fishers Ln.,Rockville, MD 20857, http:/www.fda.gov.8Association for the Advancement of Medical Instrumentation, 111 N. GlebeRd., Suite 220, Arlington, VA 222014795.F2103 112and Management of Risk9EN 12442-Part 3: Validation of the Eliminat
33、ion and/orInactivation of Virus and Transmissible Agents93. Terminology3.1 Definitions:3.1.1 chitosan, na linear polysaccharide consisting ofb(14) linked 2-acetamido-2-deoxy-D-glucopyranose(GlcNAc) and 2-amino-2-deoxy-D-glucopyranose (GlcN).3.1.1.1 DiscussionChitosan is a polysaccharide derivedby N-
34、deacetylation of chitin.3.1.2 decomposition, nstructural changes of chitosans as aresult of exposure to environmental, chemical, or thermalfactors, such as temperatures greater than 200C.3.1.2.1 DiscussionDecomposition can result in deleteri-ous changes to the chitosan.3.1.3 degradation, nchange in
35、the chemical structure,physical properties, or appearance of a material.3.1.3.1 DiscussionDegradation of polysaccharides occursby means of cleavage of the glycosidic bonds, usually by acidcatalyzed hydrolysis. Degradation can also occur thermally.Note that degradation is not synonymous with decompos
36、ition.Degradation is often used as a synonym for depolymerizationwhen referring to polymers.3.1.4 degree of deacetylation, nthe fraction or percentageof glucosamine units (deacetylated monomers) in a chitosanpolymer molecule.3.1.5 depolymerization, nreduction in length of a polymerchain to form shor
37、ter polymeric units.3.1.5.1 DiscussionDepolymerization may reduce thepolymer chain to oligomeric or monomeric units, or both. Inchitosan, hydrolysis of the glycosidic bonds is the primarymechanism.3.1.6 endotoxin, npyrogenic high molar mass li-popolysaccharide (LPS) complex associated with the cell
38、wallof gram-negative bacteria.3.1.6.1 DiscussionThough endotoxins are pyrogens, notall pyrogens are endotoxins. Endotoxins are specifically de-tected through a Limulus Amebocyte Lysate (LAL) test.3.1.7 molecular mass average (molecular weight average),nthe given molecular weight (Mw) of a chitosan w
39、ill alwaysrepresent an average of all of the molecules in the population.The most common ways to express the Mw are as the numberaverage (Mn) and the weight average (Mw). The two averagesare defined by the following equations:Mn5(iNiMi(iNiandMw5(iWiMi(iWi5(iNiMi2(iNiMiwhere:Ni= number of molecules h
40、aving a specific molecularweight Miandwi= weight of molecules having a specific molecularweight Mi. In a polydisperse molecular population therelation Mw Mnis always valid. The coefficient Mw/Mnis referred to as the polydispersity index, and willtypically be in the range 1.5 to 3.0 for commercialchi
41、tosans.3.1.8 pyrogen, nany substance that produces fever whenadministered parenterally.4. Significance and Use4.1 This guide contains a listing of those characterizationparameters that are directly related to the functionality ofchitosan. This guide can be used as an aid in the selection andcharacte
42、rization of the appropriate chitosan or chitosan salt fora particular application. This standard is intended to giveguidance in the methods and types of testing necessary toproperly characterize, assess, and ensure consistency in theperformance of a particular chitosan. It may have use in theregulat
43、ion of devices containing chitosan by appropriate au-thorities.4.2 The chitosan salts covered by this guide may be gelled,extruded, or otherwise formulated into biomedical devices foruse as tissue-engineered medical products or drug deliverydevices for implantation as determined to be appropriate, b
44、asedon supporting biocompatibility and physical test data. Recom-mendations in this guide should not be interpreted as aguarantee of clinical success in any tissue-engineered medicalproduct or drug delivery application.4.3 To ensure that the material supplied satisfies require-ments for use in TEMPs
45、, several general areas of characteriza-tion should be considered. These include identity of chitosan,physical and chemical characterization and testing, impuritiesprofile, and performance-related tests.5. Chemical and Physical Test Methods5.1 Identity of ChitosanThe identity of chitosan andchitosan
46、 salts can be established by several methods including,but not limited to the following:5.1.1 Chitosan chloride monograph Ph. Eur.9Available from European Committee for Standardization, CEN ManagementCentre, 36 rue de Stassart, B-1050 Brussels, Belgium.F2103 1135.1.2 Fourier Transform Infrared Spect
47、roscopy (FT-IR)Almost all organic chemical compounds absorb infrared radia-tion at frequencies characteristic for the functional groups inthe compound. A FT-IR spectrum will show absorption bandsrelating to bond stretching and bending and can therefore serveas a unique fingerprint of a specific comp
48、ound. Cast a chitosanfilm from a 0.25 % (w/v) solution of chitosan (in 1 % aceticacid) or chitosan salt (dissolved in water) by drying approxi-mately 500 L of the sample onto a disposable IR card10for 3to4hat60C. Record a background spectrum between 4000and 400 cm-1 using 128 scans at a resolution o
49、f 4 cm-1. Recordthe IR spectrum of a dried blank IR card, then record the IRspectrum of the sample using 128 scans at a resolution of 4cm-1, percent transmission mode. Label the peaks. Typicalfrequencies (cm-1) for chitosan are as follows:Chitosan Base(as Acetate)Chitosan Chloride Chitosan Glutamate3362b 3344b 1555b1556 1605 13961406 1513 11541153 1379 1085s1083s 11541086sThe peak designators are: sh: sharp; s: strong; m: medium;w: weak; and b: broad.5.2 Physical and Chemical Characterization of Chitosan:5.2.1 The composition an
