Friday, November 22, 2019

Determination of pH using glass electrode

The pH is the measure of the hydrogen ion concentration in a sample solution. It expresses the degree of acidity or basicity (alkalinity) of a solution. There are different methods to determine pH paper or indicator paper, litmus paper (Blue, Red), and indicators, etc. However, a pH meter is one of the most accurate types of measurement and is widely used to determine the pH value in the pH scale ranges from 0 to 14.
The pH can be determined more accurately with a pH meter that includes a probe, an electrode, and a display. A variety of pH electrodes are available in which glass electrodes are commonly used. Various glass electrodes are also available in the market. Each glass electrode has special properties so they must be carefully selected to suit the measurement application. Because the process of taking the pH measurement changes the glass membrane of a pH glass electrode over time. Due to exposure to water, alkali ions dissolve from the glass surface and the oxide groups of silicates become OH groups that cause a source layer. The use of a special pH glass electrode membrane is a reproducible equilibrium between the solution and the glass surface, dependent only on the hydrogen ion concentration in the solution and source layer.
In the glass electrode method of pH determination, the recognized pH of a reference solution is measured by two electrodes, a glass electrode, and a reference electrode, and determining the voltage generated amongst these electrodes. The variation in pH between the solution outside and inside the membrane generates an electromotive force proportional to this pH difference.

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Column chromatography solvent selection

The selection of the best solvent system for column chromatography requires a balancing act between solvent and compound polarities. For most separations, the solvent should be less-polar than the compounds and compounds must also be soluble in the solvent so they are not permanently adsorbed.

The principle of column chromatography is the adsorption of the molecules by the stationary phase. There are four different types of column chromatography as per the working mechanism such as adsorption chromatography, ion-exchange chromatography, partition chromatography, gel chromatography. The applications of column chromatography consist of pharmaceuticals as well as it is used for the purification of solutes.
The convenience and versatility of column chromatography have made it one of the most commonly used separation techniques for components. Column chromatography is also able to separate a number of analytes from a sample mixture. One more advantage of column chromatography is that using this purification technique does not require much information about the physical properties of the compound, making this method extremely valuable when separating novel analytes.
If the chemical characteristics of your sample mixture are more complex, then there are some options that are available to separate pure analytes. The travel rate of a compound through the column extremely depends on the use of the mobile phase. Typically, the more polar solvent can make the compound travel quickly through the column. For the solid phase, polar solvents include better affinity, permit the compounds to elute more quickly. Care should be taken to confirm that the solvent system selected for column chromatography has the suitable polarity to make separation among the compounds in the sample mixture. Solvent or mobile phase selection is significant for separated by column chromatography.
To recognize a better solvent system for column chromatography, a sequence of thin-layer chromatography (TLC) trials must be performed prior to conducting experiments. In order to use a binary solvent system, it may be necessary in some cases. The solvent or mobile system used relies on the behavior of the sample component on the silica gel. Generally, we select a solvent or mobile system in which the compound we wish to separate has an Rf value of about 0.3, although, many analysts run gradient systems. It starts with hexane, and then gradually increases the solvent polarity until the compound is eluted. It is also the method of HPLC.


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Why are salt plates used in IR rather than glass?

Sodium chloride (NaCl) cells are transparent to IR and do not absorb the infrared radiation in the region between 4000-450 cm-1; hence it is used to analyze the sample in IR. When analyzing the IR spectra the glass or quartz cells cannot be used because it absorbs IR radiation. Liquid samples are often cleaned between NaCl salt plates. As solvents absorb IR radiation, IR cells typically have narrow path lengths to maintain solvent absorption to adequate levels. Solid samples are often prepared by potassium bromide (KBr) in pellet form in the hydraulic pellet press. Nowadays Attenuated total reflectance spectroscopy (ATR) is another characteristic application for determining the functional group of molecules. It doesn’t need to prepare samples. This facilitates samples to be analyzed directly in solid, liquid, gel, or powder state without any preparation.


 
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Why use NaCl plates in IR
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Determination of pH using a glass electrode
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Why use NaCl plates in IR

The IR plates that we use in the IR spectroscopy are made of polished sodium chloride (NaCl). Since it is transparent to infrared radiation it is used to take the IR spectra of liquid samples, these plates work similarly to potassium bromide (KBr) for solid samples. The major difficulty of using a liquid sample is to decide a solvent with which to dilute the sample. No solvent is ideal, however, if some information about the molecule is known, a solvent can be selected accordingly. We can use for CHCl3 or DCM liquid for dissolving the sample since it is suitable for plates. Water is not a suitable solvent as NaCl is water-soluble. If there is water in the sample the plates will be ruined.
NaCl plates are delicate and very sensitive to moisture; hence nowadays the attenuated total reflectance system is used to take IR spectra. In ATR we can use a wide range of sample types such as solid, liquid, gel and powder and does not need to prepare samples. The diamond ATR is most popular because it is very rugged, simple to use and gives high-quality spectra in the region 4000–400 cm-1.

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What are the advantages of ATR?

Nowadays Attenuated Total Reflectance (ATR) accessories are the most commonly used technique in FTIR analysis. Conventional, time-consuming sample preparation methods such as KBr pellet and Nujol mull have now beyond in favor of using diamond ATR that do not need preparation of the sample, and which are used in a wide variety of sample types.
The Advantages Attenuated Total Reflection Technique (ATR) are as follows:
  • There is no required for preparation of samples, this is a major advantage of ATR. The sample has to be placed directly on the ATR sample point.
  • The minimum amount of sample is required.
  • It is a time-saving technique.
  • The simple and fast cleaning-just remove the sample and clean the crystal surface
  • No pellet pressing or grinding is required to collect spectrum
  • A wide range of sample types such as solids, liquids, gels, powders, and pastes in the entire pH range can be analyzed.
  • This gives high-quality spectra over the entire range of the infrared region (4000–400 cm-1).
  • Diamond ATR is chemically inert, very strong, and high resistance to scratches.

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Why diamond crystal is the best choice for ATR

IR spectroscopy deals with the interaction between a component and radiation in the infrared region (4000-400 cm-1), which is light with longer wavelengths and lower frequency than visible light. For solid sample pellet press technique is commonly used in infrared spectroscopy; it is prepared in a hydraulic pellet press containing potassium bromide (KBr) and sample (1:100 ratio). Liquid samples are analyzed by rectangular cells of KBr or NaCl, but both solid and liquid techniques have some limitations. Hence, Attenuated total reflectance (ATR) is the most commonly used sampling methods in Fourier transform infrared spectroscopy (FTIR). Since, it is time-saving, simple to use and gives a spectrum without the need for sample preparation.

Diamond ATR is one of the most commonly used technique to determine the functional groups of the sample components since its robustness make it appropriate for a broad range of applications. It is best for analysis of solids, liquids, gels, and pastes (either soft or hard material) and diamond ATR is not affected by the pH, color, and temperature of the sample. Diamond is chemically inert, very strong, high resistance to scratches and its complete insolubility making it a perfect crystal material for general measurements over a wide range of samples.


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Why Potassium Bromide is used in Infrared Spectroscopy

IR spectroscopy is the measurement of the intensity and wavelength of absorption of mid-infrared lights by a sample either solid/liquid/gas. IR is carried out to determine the functional group of the sample component. Nowadays the FTIR can be done by directly putting samples on Attenuated total reflectance (ATR) or by using the potassium bromide (KBr) pellet technique.
Potassium bromide is commonly used in sample preparation as it is robust, chemically inert and it is transparent in near UV as well as long-wave IR wavelength. It has no optical absorption lines in its high transmission region and due to its wide spectral range, it is broadly used as infrared optical windows.

 
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Why KBr used in IR spectroscopy

Infrared spectroscopy is a useful method to identify the functional groups of sample materials. A sample can be in solid, liquid, and gaseous states and can be analyzed by IR or FTIR spectroscopy with the sample as a transmission technique from radiant light interaction. A solid sample type is a typical sampling technique for the study of IR transmission spectroscopy. This involves preparing a solid sample in the form of a potassium bromide (KBr) disk or pellet (7 mm or 13 mm) to properly mount the sampling area of ​​an infrared spectrophotometer.
Pellet preparation is a significant factor for IR spectroscopy it has to be transparent so that it allows to pass of IR radiations through them. So to get the precise IR spectrum of a sample with good intensity, sharp peaks with high resolution, salts such as potassium bromide (KBr), sodium chloride (NaCl), and Silver chloride (AgCl) are used for the mixing of the sample.
KBr is commonly used as a window material or in pellet preparation as a carrier of the sample in the ratio of 1:100 since it is optically transparent for the light in IR between 4000–400 cm-1. KBr is commonly used in infrared spectroscopy because there is an alkyl halide and no alkyl halide shows any absorption in the region of infrared and is chemically inert and robust.


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Difference between normal and reversed-phase chromatography

High-performance liquid chromatography is a technique to separate the mixtures of sample components and is qualitative and quantitatively used in analytical chemistry. HPLC has different modes of separation such as reversed-phase chromatography, normal phase chromatography, affinity chromatography, and size exclusion chromatography, etc.

The main difference between normal phase chromatography and reversed-phase chromatography is that the NP-HPLC uses the polar mobile phase and a non-polar stationary phase while RP-HPLC uses a low polar mobile phase and a polar stationary phase.

The normal phase occurs when the mobile phase is non-polar while the stationary-phase that is sorbent is polar, which means that the more hydrophilic molecules the more it will interact with the stationary phase, hence its retention time increases inside the HPLC column.

The reversed-phase chromatography is mostly used the analytical technique to separate the analytes mobile phase is polar while the stationary phase is non-polar it means if the molecule is hydrophilic, it will have less interaction with the stationary phase results in polar analytes elutes quickly and non-polarity analytes elute slowly.
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