Characterization of natural polymers for cosmetic applications

Botha, Carlo B. (2015-12)

Thesis

ENGLISH ABSTRACT: The characterization of the molar mass and the chemical composition distributions of hyaluronic acid (HA), a linear polysaccharide, is an important task for developing structure-property correlations and for the advancement of various industrial applications. Some of the current techniques to obtain these distributions exhibit problems related to the poor sample solubility of virgin and modified HA since chemical composition and molecular size separations are typically conducted in solution. Therefore, there is a need for characterization techniques enabling the analysis of virgin and modified hyaluronic acid that are accurate, robust and reproducible. In the first part of this work solubility studies were performed on unmodified HA as well as HA modified with acrylic moieties. The aim of this work was to obtain suitable solvent systems for both species that can be used for size and chemical composition analysis. The solubility tests provided useful insight into solvents applicable for the chromatographic fractionation of the HA samples. The results gave a guideline for which solvent system is the most suitable for dissolving all the HA’s, irrespective of their degree of modification (degree of substitution, DS). For a first overview, the HA’s were analysed by bulk methods such as proton nuclear magnetic resonance (1H-NMR) and Fourier transform infrared (FT-IR) spectroscopy. From the bulk analyses, the average degrees of substitution of the HA’s were quantitatively determined by 1H-NMR spectroscopy. FT-IR spectroscopy was shown to provide fast and reliable information on DS of chromatographic fractions and is, therefore, complementary to 1H-NMR. In the present work, different analytical approaches have been developed for the chemical composition and molar mass characterization of HA and its derivatives. The combination of size exclusion chromatography (SEC) and multi-angle laser light scattering (MALLS) detection provided accurate molar mass distributions. The chemical composition separation was conducted by gradient high performance liquid chromatography (HPLC). In further investigations, these fractionation techniques were hyphenated with an information-rich detector such as FT-IR to obtain information on the degree of acrylate substitution of HA as a function of either molar mass or chemical composition. The results of this research showed that carefully conducted solubility tests are an important prerequisite for developing accurate and robust fractionation techniques. For very polar polymers such as HA and its derivatives, solvent systems must be found that suppress aggregate formation and enable the macromolecules to adopt random coil conformations. To our knowledge, the first gradient HPLC separation of HA bearing acrylate functionalities was successfully achieved in this work. The hyphenation of gradient HPLC with FT-IR provided insight into the separation mechanism and the functional group distribution of these polymers.

AFRIKAANSE OPSOMMING: Die karakter analise van die verspreidings funksies, wat die beskrywing van grootte, en chemiese verwante parameters, van hialuroniese suur (HA) ‘n lineêre polisakaried insluit, in oplossing beklemtoon die noodsaaklikheid ten opsigte van struktuur-eienskap verhoudings, vir die vooruitgang van spesifieke industriele aanwendings. Sommige van die huidige tegnieke wat bogenoemde verspreidings aanbetref het veelvuldige probleme geassosieer daarmee in terme van swak monster ontbinding van beide ongemodifiëerde en gemodifiëerde HA, sienende dat chemiese komposisie en grootte skeidings tipies in oplossing plaasvind. Daarom is karakter analitiese tegnieke wat die analise van gemodifiëerde en ongemodifiëerde HA moontlik maak, van kardinale belang om informasie te verkry, wat dus ook akkuraat, duursaam en herproduseerbaar is. In die eerste gedeelte van die werk was oplosbaarheids studies uitgevoer met betrekking tot ongemodifiëerde HA asook gemodifiëerde HA gefunksioneer met akriel groepe. Die doel van die werk was om ‘n geskikte oplossings sisteem te vind vir beide spesies wat dus ook gebruik kon word vir grootte en chemise komposisie ontleding. Die in diepte ondersoek van die oplosbaarheid van HA het insiggewende informasie gebied vir oplosmiddels wat van toepassing is met betrekking tot die chromatografiese fraksionering van die HA monsters. Die resultate het ‘n duidelike indikasie aangedui met betrekking tot watter oplosmiddel sisteem die mees gepaste sou wees vir die HA’s ten spyte van die graad van substitusie (DS). Vir ‘n eerste oorsig was die HA’s dus ook ge-analiseer deur middel van grootmaat opsporing metodes soos: proton kern magnetise resonansie (1H-KMR), en Fourier transform infra-rooi (FT-IR) spektroskopie. Van die grootmaat analise was die gemiddelde graad van substitusie van die HA’s kwantitatief bepaal deur 1H-KMR spektroskopie. FT-IR spektroskopie het getoon om voorsiening te maak vir vinnige en betroubare inligting ten opsigte van die DS van chromatografiese fraksies en is dus aanvullend ten opsigte van 1H-KMR spektroskopie. In die huidige werk was verskeie analitiese benaderinges ook ontwikkel vir die chemiese komposisie en molêre massa karakterisering van HA en sy afleibares. Die gekombineerde gebruik van grootte uitsluiting chromatografie (SEC) met ‘n multi hoek laser lig verstrooiing (MALLS) detektor het akkurate molêre massa verspreidings voorsien. Die chemiese komposisie skeiding was uitgevoer deur middel van gradiënt hoë verrigting vloeistof chromatografie (HPLC). In verdere ondersoeke was die fraksionerings tegnieke ook gekoppel met ‘n informasie-reike detektor soos FT-IR met die doel om informasie te verkry ten opsigte van die graad van akriel substitusie van HA as ‘n funksie van óf molêre massa óf chemiese komposisie. Die resultate van die navorsing het getoon dat noukerig uitgevoerde oplosbaarheids studies ‘n belangrike voorvereiste is vir die ontwikkeling van akkurate en duursame fraksionerings tegnieke. In die geval van hoogs polêre polimere soos HA en sy afleibares moet oplosmiddels gevind word wat die vorming van konglomerasie onderdruk en die makromolekule in staat stel om lukrake spoel konformasies aan te neem. Tot ons kennis is die eerste gradiënt hoë verrigting vloeistof chromatografie (HPLC) skeidings van die HA’s gemodifiëer met akrilaat funksionele groepe suksesvol bereik in die navorsings werk. Die koppeling van gradiënt hoë verrigting vloeistof chromatografie met Fourier transform infra-rooi spektroskopie het insig verskaf op die skeidings meganisme en die graad van funksionele groep verspreiding van die polimere.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/97720
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