Inverted Test Tube Method

Methods often used for determining the gel point include the inverted test tube method, fallen ball method, viscoelasticity measurement, and differential scanning calorimetric (DSC) method. In what follows experimental studies on the gel point and melting point from the viewpoint of the lost fluidity of sols will be individually introduced, with an emphasis on experimental procedures. 

This method is the simplest one used to study the gel point of a solution or the melting point of a gel. This method is often used to learn the condition of gelation qualitatively [1-3]. However, it lacks generality in that one merely observes whether or not flie sample flows upon inversion of a test tube. Therefore, it is necessary to provide both qualitative and quantitative data, such as the amoimt of sample and the diameter of the test tube when the results are written and disseminated in research papers, etc. When the gel point and melting point of fliermoreversible gels are measured, for example, sufficient care must be paid to thermal history, such as the isothermal temperature and heating rate of the sample. 

The Eldridge-Ferry study is famous for the measurement of melting point by the inverted test tube method in which qualitative data are involved [4]. In this study, a quantitative evaluation of the relationship between the transition temperature and polymer concentration for the sol- 

Ferry [22] found linearity when log c and /T were plotted, where c was the concentration of gelatin and T was the melting point of the gel expressed in absolute temperature. Similarly, they also found linearity between logM and /T. The equation derived from the van t Hoff equation is used to express this linear relationship  

Other than this example, they also studied a wide variety of polymers, such as ethylene-butadiene copolymer, ethylene-vinylacrylate copolymer, and their branched polymers. They indicated that, for cases where the junction zone includes crystals, they could not verify functionality, which is equivalent to the number of polymers that form the junction zone. 

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The fallen ball method is the method used to judge gelation by placing a small ball in a sample and is often used as a qualitative method, as is the inverted test tube method shown in Fig. 4. It is necessary to clarify the diameter, material and density of the ball, and the diameter of the test tube. 

Sol-gel-sol phase transition diagrams of composite hydrogel were recorded using test tube-inverting method [18, 36]. The sol-gel-sol transition was visually observed by inverting the vials, and conditions of sol and gel were defined as flow liquid sol and no flow solid gel in one minute respectively. 

Figure 11.13a shows a microemulsion of the 0.1 M NaCI aqueous solution/Ci2E4/ decane system with 90% aqueous solution and an oil/surfactant ratio of 2.33, at 7°C. The HLB temperature of this system is approximately 18 C. When the sample is rapidly brought to a higher temperature, 40 C in the experiment shown in Figure 11.13b, the sample becomes milky in less than 40 s. If the test tube is inverted, no flow is observed, an indication that complete emulsification has been achieved. The emulsions produced by this emulsification method have finer and narrower droplet size distributions (Figure 11.14) than those obtained by the usual methods. 

Figure 1. An illustration of the Durham tube method. A Durham tube consists of a smaller test tube inverted inside a larger tube, both of which are sterile. To test for the presence of intracellular fermentable carbohydrate the larger tube is three-quarters filled with a suspension of cells, the smaller tube is inserted into the larger tube, and the cap is screwed tight The assembly is then inverted to fill the enclosed smaller tube with the suspension, righted, and incubated at an appropriate temperature (30 C is good for yeasts) as shown for tube A. If fermentable carbohydrates are present in the suspended cells, gas will accumulate in the top of the inverted smaller tube, causing it to float inside the larger tube, as shown for tube B...

Fig. 11 a Phase diagram of the poly(alanine-co-leucine)-poloxamer-poly(alanine-co-leucine) aqueous thermogelling solutions determined by the test tube inverting method, b Storage modulus (G ) and loss modulus (G") of the poly(alanine-co-leucine)-poloxamer-polyCalanine-co-leucine) aqueous thermogelling solutions as a function of temperature and concentration. The legends are the concentrations of the polymers in water. Reproduced with permission from [62]... 

Sharp increases in both the storage modulus (G ) and loss modulus (G") of PAL-PLX-PAL aqueous solutions were observed as the temperature increased (Fig. 11). G and G" are an elastic component and a viscous component of the complex modulus of a system, respectively. When G is greater than G", the system is considered to be a gel, and the crossover point was defined as the sol-to-gel transition temperature. The sol-to-gel transition temperatures defined by the test tube inverting method coincided with those defined by dynanfic mechanical analysis of G and G" within 2 to 3 °C. By varying the polymer concenlration, not only sol-to-gel transition temperature but also modulus of the gel could be controlled. The control of gel modulus (G ) has a significant effect on 3D cell culture as well as the differentiation of the stem cell. In the case of chondrocytes, the modulus of 300-2,500 Pa showed a cytocompatible microenvironment for proliferation of the cells. The gel prepared from 10.0 wt% aqueous solution of PAL-PLX-PAL formed a gel with a G of 380 Pa at 37 °C, thus being recommendable as a 3D culture matrix for chondrocytes. 

Two-Enzyme Micropolarization Method. Since little of the sample was available, this was tested by the semimicro method. 6.5 g. of sample and 0.5 ml. of lead solution were made up to 25 ml. 0.2 g. of ammonium hydrogen phosphate was used for de-leading. A semimicro polarization tube (200 mm.) holding 2 ml. was used for all polarizations. Reading of the de-leaded solution gave a direct polarization of - -85.88°S. 5 ml. of de-leaded solution was inverted overnight with 0.5 ml. of Sumasuco invertase and made up to 10 ml. ... 

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