Evaluation of Monolayers

The ability to image lateral heterogeneity in Langmuir monolayers dates back to Zocher and Stiebel s 1930 study with divergent light illumination [166]. More recently the focus shifted toward the use of fluorescence microscopy of mono-layers containing a small amount of fluorescent dye [167]. Even in single-corn- 

A newer and perhaps more useful application of ellipsometry to Langmuir films is their lateral characterization via ellipsometric microscopy [146], A simple modification of the nuU ellipsometer allows one to image features down to 10-/im resolution. Working with a fixed polarizer and analyzer, some domains are at extinction while others are not and appear bright. This approach requires no fluorescent label and can be applied to systems on reflective supports. 

Brewster angle microscopy takes advantage of the reflectivity behavior of light at an interface. This method relies on the fact that light passing from a material of lower refractive index, no into a medium of higher index i will have 

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One must conclude therefore that the BET procedure for evaluation of monolayer capacity is not applicable to a Type III (nor by implication, to a Type V) isotherm. 

Force of Adhesion of a Powder Layer. Let us examine further the methods used in evaluating the adhesion of a layer of powder as distinguished from the methods used in evaluation of monolayer adhesion. First, let us note certain inaccuracies in the methods being used, as reported in the work of Cremer, Buzagh, and other investigators. 

The external reflection of infrared radiation can be used to characterize the thickness and orientation of adsorbates on metal surfaces. Buontempo and Rice [153-155] have recently extended this technique to molecules at dielectric surfaces, including Langmuir monolayers at the air-water interface. Analysis of the dichroic ratio, the ratio of reflectivity parallel to the plane of incidence (p-polarization) to that perpendicular to it (.r-polarization) allows evaluation of the molecular orientation in terms of a tilt angle and rotation around the backbone [153]. An example of the p-polarized reflection spectrum for stearyl alcohol is shown in Fig. IV-13. Unfortunately, quantitative analysis of the experimental measurements of the antisymmetric CH2 stretch for heneicosanol [153,155] stearly alcohol [154] and tetracosanoic [156] monolayers is made difflcult by the scatter in the IR peak heights. 

An essential component of cell membranes are the lipids, lecithins, or phosphatidylcholines (PC). The typical ir-a behavior shown in Fig. XV-6 is similar to that for the simple fatty-acid monolayers (see Fig. IV-16) and has been modeled theoretically [36]. Branched hydrocarbons tails tend to expand the mono-layer [38], but generally the phase behavior is described by a fluid-gel transition at the plateau [39] and a semicrystalline phase at low a. As illustrated in Fig. XV-7, the areas of the dense phase may initially be highly branched, but they anneal to a circular shape on recompression [40]. The theoretical evaluation of these shape transitions is discussed in Section IV-4F. 

Evaluation of the monolayer capacity from a stepped isotherm raises... 

The BET method for calculation of specific surface A involves two steps evaluation of the monolayer capacity n from the isotherm, and conversion of n into A by means of the molecular area a . 

If the swelling effects in the polymeric phase are considered, the evaluation of the adsorbed macromolecules as a monolayer seems to be too approximate. The mean-square end-to-end distance of the polystyrene coil (M = 10000) in... 

The evaluation of the apparent ionization constants (i) can indicate in partition experiments the extent to which a charged form of the drug partitions into the octanol or liposome bilayer domains, (ii) can indicate in solubility measurements, the presence of aggregates in saturated solutions and whether the aggregates are ionized or neutral and the extent to which salts of dmgs form, and (iii) can indicate in permeability measurements, whether the aqueous boundary layer adjacent to the membrane barrier, Umits the transport of drugs across artificial phospholipid membranes [parallel artificial membrane permeation assay (PAMPA)] or across monolayers of cultured cells [Caco-2, Madin-Darby canine kidney (MDCK), etc.]. 

Electron microscopic evaluation of infected Caco-2 monolayers grown on membranes, under conditions that induced cellular polarization, prompted the conclusion that the larvae occupy the cytoplasm of cells they invade (ManWarren et al., 1997). Apical and basal plasma membranes appeared to be preserved in infected cells (Fig. 6.3). These findings reproduced the observations of Wright (1979) in his examination of intestinal tissues from infected mice. In contrast, when Li et al. (1998) performed similar experiments in HT29 monolayers grown on plastic, they concluded... 

Fig. 5.2. Two-step process for evaluation of intestinal drug absorption. The first step represents the prediction of intestinal permeability (e.g., over Caco-2 monolayers) from in-silico models or from physico-chemical...

Very few published data exist on the evaluation of automated systems, though one report has been made of an automated absorption assay using Caco-2 cells cultured on both sides of polycarbonate membranes [93], The concept of culturing cells on the lower sides of the membranes was investigated as a means of improving the opportunity to study transport in the secretory basolateral to apical direction. However, this approach resulted in increased variability and impaired active transport properties of the cell monolayers, and was therefore not recommended. 

EVALUATION OF MELTING AND CRYSTALLINE RELAXATION TEMPERATURES OF FATTY ACID MONOLAYERS ON THE WATER SURFACE... 

According to this equation, a plot of dependence of [1/(,P0//J 1 )]/q(P/P0) on P/P0 should have a linear form allowing evaluation of qm and CBET. The surface area is calculated by Equation 9.18 and the corresponding value is frequently designated by, 4BET. One can also determine the pressure, PJP(p under which a monolayer is being formed ... 

Evaluation of the epithelial integrity can be performed by measuring the transepithelial electrical resistance (TEER). TEER values ranging from 150 ohms.cm2 up to 600 ohms.cm2 have been reported. An alternative method for assessing the monolayer integrity is to monitor the flux of hydrophilic marker molecules that pass the monolayers by the paracellular route (e.g., mannitol, Na-fluorescein, or atenolol). 

There are several cell monolayer models that are frequently used for the evaluation of drug permeability and absorption potential (Table 18.1). For a more detailed discussion, please refer to Chap. 8. Caco-2 cells (adenocarcinoma cells derived from colon) are the most extensively characterized and frequently used of the available cell lines [5-9], A unique feature of Caco-2 cells is that they undergo spontaneous enterocyte differentiation in cell culture. Unlike intestinal enterocytes, Caco-2 cells are immortalized and replicate rapidly into confluent monolayers. When the cells reach confluency during culture on a semi-porous membrane, they start to polarize and form tight junctions, creating an ideal system for permeability and transport studies. During the past decade, use of... 

When a surfactant is injected into the liquid beneath an insoluble monolayer, surfactant molecules may adsorb at the surface, penetrating between the monolayer molecules. However it is difficult to determine the extent of this penetration. In principle, equilibrium penetration is described by the Gibbs equation, but the practical application of this equation is complicated by the need to evaluate the dependence of the activity of monolayer substance on surface pressure. There have been several approaches to this problem. In this paper, previously published surface pressure-area Isotherms for cholesterol monolayers on solutions of hexadecy1-trimethyl-ammonium bromide have been analysed by three different methods and the results compared. For this system there is no significant difference between the adsorption calculated by the equation of Pethica and that from the procedure of Alexander and Barnes, but analysis by the method of Motomura, et al. gives results which differ considerably. These differences indicate that an independent experimental measurement of the adsorption should be capable of discriminating between the Motomura method and the other two. 

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