Publication Date



Open access

Embargo Period


Degree Type


Degree Name

Doctor of Philosophy (PHD)


Chemistry (Arts and Sciences)

Date of Defense


First Committee Member

Roger. M. Leblanc

Second Committee Member

Jamie D. Walls

Third Committee Member

Burjor K. Captain

Fourth Committee Member

Kerim M. Gattás Asfura


Understanding of biomacromolecule interactions at the molecular level raises certain difficulties however with the Langmuir monolayer technique, interfacial interactions in two dimensions can be accurately measured to access and characterize physical, electrostatic and photophysical properties. Macromolecules such as serum albumin which comprises 50% of human blood plasma protein content is one focus of research in this thesis for its unique involvement in many biological functions. The important nature of this class of protein demands that it be studied in detail while modifying the experimental conditions in two dimensions to observe it in all types of environments. While different from bulk colloidal solution work, the two dimensional approach allows for the observation of the interaction between molecules and subphase at the air-water interface. Compiled in this thesis are studies which highlight the characterization of this protein using various surroundings and also observing the types of interactions it would have when at the biomembrane interface. Free-energy changes between molecules, packing status of the bulk analyte at the interface as well as phase transitions as the monolayer forms a more organized or aggregated state are just some of the characteristics which are observed through the Langmuir technique. This unique methodology demonstrates the chemical and physical behavior of this protein at the phase boundary throughout the compression of the monolayer. β-galactosidase (Escherichia coli is a naturally occurring, regenerating enzyme which specifically hydrolyzes the β-D-galactosyl linkage of lactose molecules into usable sources of carbon. Spectroscopic techniques were utilized to analyze β-galactosidase in its native state and in different environmental aqueous conditions as well as changes of interfacial properties and conformation investigated through surface chemistry and in situ spectroscopy. Characteristic for proteins consisting of tryptophan residues, UV-vis absorption was analyzed to observe changes in enzyme concentration revealing near its isoelectric point (4.6), β-galactosidase is most susceptible to monomer aggregation. Circular dichroism studies showed environmental aqueous alkaline conditions causes an increase in α-helix and a decrease in β-sheet content while the opposite effect was observed in acidic conditions, increasing the β-sheet content to a maximum of 43% and a minimum of α-helix content to 5%. Fluorescence assays showed that tryptophan emissions decreased over a short term of irradiation during experimentation leading to the conclusion that β-galactosidase fluorescence quenching is due to the non-radiative energy transfer of excited state donors to ground state acceptors while undergoing no major secondary structure changes. Substrate studies were conducted in order to calculate the Michaelis constant of X-gal, a glycoside substrate which undergoes hydrolysis in the presence of the enzyme. Analysis and comparison of the Michaelis constant of other known glycosides shows that X-gal has a high affinity for β-galactosidase. Conditions for an optimal Langmuir monolayer were firstly obtained by varying the subphase salt concentration and the surface pressure-area isotherm was used to extrapolate the limiting molecular area of the enzyme monolayer. Surface pressure stability measurements along with compression-decompression cycles revealed no aggregate formation. Consistent with the high content of tryptophan along with the data obtained from the isotherm, in situ UV-vis and fluorescence spectroscopy shows a steep rise in absorbance and photoluminescent intensity correlating to with a switch from a liquid-expanded to a liquid-condensed phase. The secondary structure, analyzed by infrared reflection-absorption spectroscopy of the monolayer at the air-water interface confirmed the stability of the enzyme evidenced by signal intensification as a function of increased surface pressure. A decrease in subphase pH increased the electrostatic repulsion as the enzyme was protonated leading to an expanded monolayer, increased the β-sheet content in the amide I region as well as increased the α-helix content in the amide II region. Recent years have produced many advances in quantum dot synthesis, application and analysis while their importance in the fields of chemistry, biology, engineering and physics has grown as well. Opposed to the bulk state properties, quantum dots are dependent upon the quantum confinement effect in all three spatial dimensions and so their applications have been broadened to optical switches and fluorescence labeling among others. Compiled here are extensive results in the characterization of CdSe(ZnS)-TOPO quantum dots using the Langmuir monolayer technique to approach these quantum dots in their two dimensional state as well as variation of ligands DHLA and MPS around the nanoparticles. Properties such as the limiting nanoparticle area, molar absorptivity, and self-assembly manipulation have shown the packing structure of the quantum dots at the air-water interface.


Langmuir monolayer; spectroscopy; human serum albumin; β-galactosidase; quantum dots