Electronegativity

Yuri Anthony D. Sucgang
BMLS-IA

Electrophoresis Machine
Gel Electrophoresis Apparatus(Aragose Gel)
Electrophoresis is the motion of dispersed particles relative to a fluid under the influence of a spatially uniform electric field.

Gel electrophoresis is a laboratory procedure used to separate biological molecules with an electrical current. In this lesson, we'll review how agarose gel electrophoresis works and introduce the equipment necessary to perform an electrophoresis experiment.

Separation of DNA molecules of different sizes can be achieved by using an agarose gel. Recall that agarose is a polysaccharide that can be used to form a gel to separate molecules based on size. Because of the gelatin-like nature of agarose, a solution of agarose can be heated and cooled to form a gel in a casting tray.
Think of casting the agarose gel like pouring hot gelatin into a mold. The hot agarose liquid is poured into a casting tray. Once the mixture cools, a thin agarose brick will form. To ensure there's a place to put the DNA in the gel, a comb is placed in the agarose liquid before it cools. Each tooth in the comb will become a hole, or 'well,' in the solidified agarose gel.
Once cast, this gel is placed inside a piece of equipment called a gel box. An electrode - one positive and one negative - resides at each end of the gel box. The wells are always oriented, so they're farther from the positive electrode. This ensures that the DNA molecules in the well must travel through the majority of the agarose gel, thus providing sufficient time for separation.
Air isn't a great conductor of electricity, so we cover the gel with electrophoresis buffer. Electrophoresis buffer is a salt solution. It isn't table salt, but the salt ions can carry an electrical charge just like salt water can. The salt in the electrophoresis buffer completes the circuit between the positive and negative electrodes.
When the electrodes of the gel box are connected to a power supply, electricity flows through the electrical circuit, causing the negatively charged DNA molecules to move into the agarose gel. The DNA molecules continue to travel through the agarose toward the positive electrode as long as an electrical current is present. Recall that shorter DNA molecules travel through agarose faster than longer DNA molecules. In this way, agarose gel electrophoresis separates different DNA fragments based on size.
Once the samples are loaded, the electrical current supplied by the power supply not only moves the DNA samples through the gel but the dye molecules as well. Note the colored lines that appear. These lines do not represent the DNA fragments. These lines represent the dye in the loading buffer that was used to visualize the samples during the loading step.
Once the gel run is complete, the agarose gel can be removed from the gel box and soaked in an ethidium bromide solution. Recall that ethidium bromide is used to visualize DNA. Ethidium bromide molecules intercalate, or insert, between the nitrogenous bases in a DNA molecule.
In summary, gel electrophoresis is a laboratory procedure used to separate biological molecules with an electrical current. Together with a gel box and a power supply, an agarose gel can be used to separate DNA molecules based on size. Loading buffer enables scientists to insert DNA samples into the wells of the agarose gel.
Once the electrophoresis procedure is initiated, the dye in the loading buffer forms a dye front that is used to determine when the procedure is complete. When the electrophoresis procedure is complete, the agarose gel can be soaked in an ethidium bromide solution to visualize the DNA bands on a UV box.

Yuri Anthony D. Sucgang
BMLS-IA

Chromatography Equipments
Column Chromatography

Chromatography is the collective term for a set of laboratory techniques for the separation of mixtures. The mixture is dissolved in a fluid called the mobile phase, which carries it through a structure holding another material called the stationary phase.

Column chromatography is basically a type of adsorption chromatography techniques. Here the separation of components depends upon the extent of adsorption to stationary phase. Here the stationary phase is a solid material packed in a vertical column made of glass or metal.
When a mixture of mobile phase and sample to be separated are introduced from top of the column, the individual components of mixture move with different rates. Those with lower affinity and adsorption to stationary phase move faster and eluted out first while those with greater adsorption affinity move or travel slower and get eluted out last.
The solute molecules adsorb to the column in a reversible manner. The rate of the movement of the components is given as follows
R= Rate of movement of a component / Rate of movement of mobile phase. i.e. it is the ratio of distance moved by solute to the distance moved by solvent.
Planar Chromatography
Planar chromatography is a separation technique in which the stationary phase is present as or on a plane. The plane can be a paper, serving as such or impregnated by a substance as the stationary bed (paper chromatography) or a layer of solid particles spread on a support such as a glass plate (thin layer chromatography). Different compounds in the sample mixture travel different distances according to how strongly they interact with the stationary phase as compared to the mobile phase.

Paper chromatography is a technique that involves placing a small dot or line of sample solution onto a strip of chromatography paper. The paper is placed in a jar containing a shallow layer of solvent and sealed. As the solvent rises through the paper, it meets the sample mixture, which starts to travel up the paper with the solvent. This paper is made of cellulose, a polar substance, and the compounds within the mixture travel farther if they are non-polar. More polar substances bond with the cellulose paper more quickly, and therefore do not travel as far.
Paper is made of cellulose fibres, and cellulose is a polymer of the simple sugar, glucose. The key point about cellulose is that the polymer chains have -OH groups sticking out all around them. To that extent, it presents the same sort of surface as silica gel or alumina in thin layer chromatography. It would be tempting to try to explain paper chromatography in terms of the way that different compounds are adsorbed to different extents on to the paper surface. In other words, it would be nice to be able to use the same explanation for both thin layer and paper chromatography. Unfortunately, it is more complicated than that! The complication arises because the cellulose fibres attract water vapour from the atmosphere as well as any water that was present when the paper was made. You can therefore think of paper as being cellulose fibres with a very thin layer of water molecules bound to the surface. It is the interaction with this water which is the most important effect during paper chromatography.

Yuri Anthony D. Sucgang
BMLS-IA

Electrochemical Cell Set-up(experiment)

a) The simplest experimental set up for performing electrochemical experiments. b) A schematic representation of a three electrode potentiostat .
Up to now we discussed shortly the principles of semiconductor-electrolyte interfaces. However, it was said nothing about the practical implementation of electrochemical techniques. Let us start by discussing the most simple experimental setup for performing electrochemical measurements. The following elements are necessary: three electrodes immersed into an electrolyte;
A battery;
A voltmeter.
An amperemeter.
The electrode, which has to be studied or where the electrochemical reaction should take place (in our case it will be the electrode in which we intend to introduce pores), is called the working electrode (WE). The second electrode, which is closing the circuit, is called the counter electrode (CE). The third one, used to measure the voltage between the electrolyte and WE, is called the reference electrode (RE).
If a current is flowing thought a solid-electrolyte interface, chemical reactions will occur at the interface. Electrons leaving the solid will reduce species in solution, whereas electrons moving into the solid lead to the oxidation of species in solution. Due to the chemical reactions at the interface the composition of the electrolyte near the surface of the solid will change continuously and thus the distribution of the voltage across the semiconductor-electrolyte interface will vary as well. This is especially true at high current densities. For this reason, in order to maintain a constant composition of the solution near the surface of the sample, i.e. the working electrode, continuous pumping of the electrolyte is necessary.

a) A schematic representation of a setup with a three electrode potentiostat. b) A schematic representation of a set up with a four electrode potentiostat.
As long as the contact between the sample and the working electrode is good enough. However, if the ohmic resistance between the WE and the sample is not sufficiently small, e.g. when using In/Ga alloy, a four electrode potentiostat must be used. The fourth electrode is called the sense electrode (SE ) and is connected to the sample. Now, the potentiostat can be regarded as being composed of two 'independent' subsystems. One containing the WE and CE electrodes through which the current is flowing. Correspondingly, the second subsystem contains SE and RE electrodes and measures the potential. A feedback interaction between the two subsystems results in an ideal tool for controlling electrochemical processes. In this configuration the contact between the sense electrode and the sample is not critical because no current is flowing through it. Now the desired potential will be exactly applied on the sample/electrolyte junction.
In spite of the fact that a four electrode potentiostat diminishes the importance of the contact quality between WE and the sample, during the pore formation process the quality of the contact between the sample and the working electrode is still very important. It determines how uniform the current is distributed across the whole surface of the sample. If the contact is not uniform the distribution of the current and consequently the porous layer will not be uniform as well.
A significant improvement of the uniformity of the backside contact can be achieved by a liquid contact, i.e. the sample has two electrolyte junctions. The first junction (the front side) will be the one of interest and where the pores will grow. The second junction will play the role of an uniform backside contact. On both junctions electrochemical reactions will take place. If at the front junction an anodic reaction takes place, then at the back contact a cathodic reaction will occur. This idea can be realized by the so called double electrochemical cell.

Yuri Anthony D. Sucgang
BMLS-IA

Separation of Cations and Anions
Classification of the Cations and Anions
The five groups of cations and the characteristics of these groups are as follows:
*Group 1 Cations of this group form precipitates with dilute hydrochloric acid. Ions of this group are lead(II), mercury(I), and silver(I).

*Group 2 The cations of this group do not react with hydrochloric acid, but form precipitates with hydrogen sulphide in dilute mineral acid medium. Ions of this group are mercury(II),copper(II), bismuth(III), cadmium (II), tin(II), tin(IV), arsenic(III), arsenic(V), antimony(III), and antimony(V).
The first four form the sub-group 2/a and the last six the sub-group 2/b.
While sulphides of cations in Group
2/a are insoluble in ammonium polysulphide, those of cations in
Group 2/b are soluble.

*Group 3 Cations of this group do not react either with dilute hydrochloric acid, or with hydrogen in dilute mineral acid medium.However they form precipitates with ammonium sulphide
SrCO3, BaCO3 (s)in neutral or ammoniacal medium.
Cations of this group are iron(II), iron(III),cobalt(II),nickel(II), manganese(II),chromium(III), aluminium(III), and zinc(II).

*Group 4 Cations of this group do not react with the reagents of Groups 1, 2, and 3.
They form precipitates with ammonium carbonate in the presence of ammonium chloride in neutral medium. Cations of this group are calcium(II), strontium(II), and barium(II). *Group 5 Common cations, which do not react with reagents of the previous groups, form the last group of cations, which includes magnesium(II), lithium(I), sodium(I), potassium(I), and ammonium(I) ions.

The methods available for the detection of anions are not as systematic as those which have been described above for cations. No really satisfactory scheme has yet been proposed which permits the separation of the common anions into major groups, and the subsequent unequivocal separation of each group into its independent constituents; however, it is possible to detect anions individually in most cases, after perhaps a 1-2 stage separation. It is advantageous to remove all heavy metals from the sample by extracting the anions through boiling with sodium carbonate solution; heavy metal ions are precipitated out in the form of carbonates, while the anions remain in solution accompanied by sodium ions.
The following scheme of classification of anions has been found to work well in practice; anions are divided into four groups on the basis of their reactions with dilute hydrochloric acid and of the differences of solubilities of their barium and silver salts.
The four groups of anions and the characteristics of these groups are as follows:
Group 1 Visible change, gas evolution and/or formation of a precipitate, with dilute hydrochloric acid. Ions of this group are carbonate, silicate, sulphide, sulphite, and thiosulphate. Group 2 The anions of this group do not react with hydrochloric acid, but form precipitates with barium ions in neutral medium. Ions of this group are sulphate, phosphate, fluoride, and borate.
Group 3 Anions of this group do not react either with dilute hydrochloric acid, or with barium ions in neutral medium. However, they form precipitates with silver ions in dilute nitric acid medium. Anions of this group are chloride, bromide, iodide, and thiocyanate. Group 4 Common anions, which do not react with reagents of the previous groups, form the last group of anions, which includes nitrite, nitrate and chlorate ions. The identification of a single cation in solution is a fairly simple and straightforward process, although without a good identification scheme it may require so many experiments as the number of potential cations, if we know at least one specific reaction for each cation. In order to reduce the number of tests required for the identification, it is important to develop a good identification scheme, which reduces the number of potential cations step by step placing them into groups. There are several possibilities and anyone could develop his/her own identification scheme.
The scheme you find below follows the classification of cations into groups, as described in the Fresenius' system. In case of a solid sample it is assumed that the sample is soluble in water or dilute nitric acid.
Once the cation is found, its presence should be verified by other, characteristic reactions. TESTING FOR A SINGLE CATION IN SOLUTION

(1) Group I cations Add to the solution an excess of dilute HCl. If there is no change, follow (2a).
A white precipitate may contain Pb2+, Hg22+ or Ag+.
Filter and wash the precipitate and then add NH3 solution to the precipitate.
If the precipitate does not change:Pb2+ present turns black: Hg22+ present dissolves:Ag+ present

(2a) Group IIA cations Acidify the solution and add H2S in excess. If there is no change follow (3).
A precipitate may result if Hg2+, Bi3+, Cu2+, Cd2+, As3+, As5+, Sb3+, Sb5+, Sn2+,
Sn4+ were originally present. (Check the colour of the precipitate !)
Filter the precipitate, wash with dilute HCl, and treat with an excess of (NH4)2Sx. If the precipitate dissolves, follow (2b).

If the remaining precipitate is yellow: Cd2+ present

Take a fresh sample and add dilute NaOH. If the precipitate is blue:Cu2+ present yellow:Hg2+ present white:Bi3+ present

(2b) Group IIB cations Add dilute HCl to the (NH4)2Sx filtrate in excess, when the precipitate reappears. Take a fresh sample, acidify, and precipitate the sulphide.
Examine its colour: brown precipitate:Sn2+ present
An orange precipitate indicates Sb. To identify its oxidation state, take a fresh sample, acidify with 1:1 HCl and add KI: no colouration:Sb3+ present brown colourationSb5+ present
A yellow precipitate indicates As or Sn4+. Add (NH4)2CO3 in excess.
If the precipitate remains undissolved:Sn4+ present Perform the luminescence test for identifying Sn4+ ions.

If the precipitate dissoves:As3+ or As5+ present.
To identify the oxidation state of As present in the solution, take a fresh sample, acidify with 1:1 HCl and add KI: no coloration:As3+ present brown coloration:As5+ present

(3) Group III cations Neutralise the solution with NH3 solution and add (NH4)2S in excess. If there is no change, follow (4). Examine the precipitate.
A green precipitate indicates Cr3+. To a fresh sample, add NaOH: green precipitate which dissolves in an excess of the reagent:Cr3+ present

A pink (flesh-like) precipitate indicates Mn2+. To a fresh sample, add NaOH: white precipitate, which turns darker on standing:Mn2+ present
A white precipitate may be caused by Al3+ or Zn2+. To a fresh sample add NH3, first in moderate amounts, then in excess: white precipitate, which dissolves in excess NH3 soln.: Zn2+ present white precipitate, which remaines unchanged if excess NH3 is added: Al3+ present
A black precipitate occurs if Co2+, Ni2+, Fe2+ or Fe3+ were present originally.
Filter, wash and mix the precipitate with 1:1 HCl. The precipitate dissolves if Fe2+ or
Fe3+ were present, otherwise it remains unchanged.
To a fresh sample add NaOH in excess: green precipitate, turning dark on standing: dark brown precipitate: blue precipitate, turning pink if excess NaOH is added: green precipitate, which remains unchanged on standing: Fe2+ present
Fe3+ present

Co2+ present

Ni2+ present

(4) Group IV cations To the solution add (NH4)2CO3 in excess, in the presence of NH4Cl. If there is no precipitation, follow (5).
A white precipitate indicates the presence of Ba2+, Sr2+ or Ca2+.
To a fresh sample add a four fold (in volume) of saturated CaSO4 solution: immediate white precipitate:Ba2+ present a white precipitate is slowly formed: Sr2+ present no precipitation occurs:Ca2+ present

(5) Group V cations Heat a fresh sample gently with some dilute NaOH: characteristic odour of ammonia: NH4+ present
Carry out a flame test with the original sample: red coloration: yellow coloration: pale violet coloration:
To the solution add NaOH in excess: white precipitate, which turns red by adding a few drops of titan yellow reagent:Mg2+ present
+
Li present
Na+ present K+ present

TESTING FOR A SINGLE ANION IN SOLUTION The identification of a single anion in solution is a fairly simple and straightforward process, and anyone could develop his/her own identification scheme.
The scheme below follows the classification of anions into four groups, as described on page 131. It is assumed that the heavy metals are removed from the solution. In case of a solid sample it is assumed that the sample is soluble in water.
Once the anion is found, its presence should be verified by other, characteristic reactions. (1) Group I anions Add to the solution an excess of dilute HCl. If there is no change, follow (2).
If a white precipitate or/and gas liberation is observed, one of the following anions 2−2− 2−2−2− may present: CO3 , SiO3 , S , SO3 , S2O3 .
White, gelatinous precipitate without the liberation of any gas:
SiO3
2− present White precipitate with the liberation of SO2. The gas is tested with a filter paper moistened with potassium iodate and starch solution. 2− blue coloration:S2O3 present

No precipitate, only gas liberation is observed. Test the gas with filter paper moistened with lead acetate solution. 2− black coloration:S present Test the gas with filter paper moistened with potassium iodate and starch solution. 2− blue coloration:SO3 present Introduce the gas into baryta or lime water: 2− white precipitate:CO3 present

(2) Group II anions Neutralise the solution and add BaCl2 solution. If there is no change follow (3). 2−3− −3−
A white precipitate may result if SO4 , PO4 , F , or BO3 was originally present.

Filter the precipitate, and add HCl solution. precipitate is not soluble:
Add concentrated sulphuric acid to the precipitate or to the original solid sample, and warm the test tube: − test-tube acquires a greasy appearance:F present
SO4 present
2−

Add concentrated sulphuric acid and ethanol to the precipitate or to the original sample in a porcelain basin and ignite the alcohol: 3− green-edged flame:BO3 present
−
−
−

Take a fresh sample and add ammonium molybdate reagent. 3− yellow, crystalline precipitate:PO4 present
(3) Group III anions Acidify the solution with dilute nitric acid and add AgNO3 solution. If there is no change, follow (4).
Examine the precipitate. −−
A yellow and yellowish white precipitate indicates I and Br , respectively.
To a fresh sample, add 1-2 ml carbon tetrachloride and chlorine water dropweise, and shake it intensively: violet organic layer, which turns − colourless with excess chlorine water:I present reddish-brown organic layer, which turns − yellow with excess chlorine water:Br present
A white precipitate may be caused by Cl or SCN .
To a fresh sample add FeCl3 solution: − blood-red coloration:SCN present
Apply the chromyl chloride test: positive test:
Cl present
(4) Group IV anions −−− One of the following ions may present: NO2 , NO3 , or ClO3 .
Acidify a fresh sample with acetic acid, add sulphanilic acid and α-naphthylamine reagents: − red coloration:NO2 present
Acidify a fresh sample with acetic acid, add sulphanilic acid and α-naphthylamine reagents and zinc chips: − red coloration:NO3 present
Add zinc chips to a fresh sample, filter the solution after a couple of minutes, acidify with dilute nitric or sulphuric acid and add AgNO3 solution to the filtrate: − white precipitate:ClO3 present