Nirmal G's Blog

Normality for Solutions

Nirmal Kumar G — Wed, 07 Dec 2011 00:32:18 +0000

Find the Normality of a 70% pure O-Phosphoric acid (Specific Gravity – 1.54) of 100ml solution.

Ans:

Weight of O-Phosphoric acid is 70 x 1.54 = 107.8

Equivalent Weight is 98/3 = 32.6667

Normality = 107.8/(32.6667 x 0.1) = 33.0N

Other way:

1000 ml of solution contains 1540gm of phosphoric acid

Of this 70% mean 1078gm of phosphoric acid

Normality = 1078/32.6667 = 33.0N

Normality = Molarity x n (no. of hydrogen ions)

Nucleic Acids Estimation – Types, Sensitivity & Interferences

Nirmal Kumar G — Sat, 26 Mar 2011 05:40:00 +0000

Method	Sensitivity	Time	Principle	Interferences	Comments
Spectro photometric (A₂₆₀)	High 2.5µg	Rapid 5-10min	Absorption of 260nm light by aromatic bases	Proteins	Protein absorption may be corrected. A280/A260 indicates purity. Non-destructive. Measures both DNA and RNA.
Bisbenzimidazole	Very high 10ng	Rapid 5-10min	Enhancement of fluorescence of dye when bound to DNA	None known	Not reliable with plasmids or small DNA. Does not measure RNA. May be used with crude extracts. Non-destructive.
DNA dipstick	Very high 0.1-10ng	Moderate 30min	Proprietary	SDS, agarose	Gives no indication of purity. Measures single and double-stranded DNA and RNA.
EtBr-Agarose	Very high 1-10ng	Moderate 30-45min	Fluorescence of bound EtBr	None known	Measures DNA and RNA.

Protein Estimation – Types, Sensitivity & Effectors

Nirmal Kumar G — Sat, 26 Mar 2011 05:26:00 +0000

Method	Sensitivity	Time	Principle	Interferences	Comments
Biuret	Low (1-20mg)	Moderate 20-30min	Peptide bonds + alkaline Cu²⁺ gives purple complex	Zwitterionic buffers, some amino acids	Similar color with all proteins. Destructive to protein samples.
Lowry	High (~5µg)	Slow 40-60min	1.Biuret reaction 2.Reduction of phosphomolybdate-phosphotunstate by Tyr and Trp	Ammonium sulphate, glycine, zwitterionic buffers, mercaptans	Time-consuming. Color varies with proteins. Critical timing of procedure. Destructive to protein samples.
Bradford	High (~1µg)	Rapid 15min	λ_max of Coomassie dye shifts from 465nm to 595nm when protein-bound	Strongly basic buffers, detergents Triton X-100, SDS	Stable color which varies with proteins. Reagents commercially available. Destructive to protein samples. Discoloration to glassware.
BCA	High (1µg)	Slow 60min	1.Biuret reaction 2.Copper complex with BCA; λ_max = 562nm	EDTA, DTT, ammonium sulphate	Compatible with detergents. Reagents commercially available. Destructive to protein samples.
Spectrophotometric (A₂₈₀)	Moderate (50-1000µg)	Rapid 5-10min	Absorption of 280nm light by aromatic residues	Purines, pyrimidines, nucleic acids	Useful for monitoring column eluents. Nucleic acid absorption can be corrected. Non-destructive to protein samples. Varies with proteins.

Photophosphorylation

Nirmal Kumar G — Sat, 26 Mar 2011 05:18:00 +0000

Photosynthetic phosphorylation or photophosphorylation is the process of phosphate group transfer into ADP to synthesize energy rich ATP molecule making use of light as external energy source. According to chemi osmotic hypothesis (Mitchell 1961) the ATP is synthesized on ATPase complexes located on the non appressed portions of thylakoid membranes particularly towards margins.

During photosynthetic electron transport, hydrogen protons (H+) accumulate in the thylakoid space, due to splitting of water and transport between PQH2 to Cytf. Increase in the number of hydrogen protons in the thylakoid space results in increase of proton gradient. Down flow of protons from high to low concentration along H+ conc. gradient through ATPase complex provides the energy that allows an ATP synthase enzyme to produce ATP from ADP + Pi

Non – Cyclic Photophosphorylation

The electrons lost by P680 (PS-II) are taken up by P700 (PS-I) and do not get back to P680 i.e., unidirectional and hence it is called non- cyclic phosphorylation. The electrons pass through the primary acceptor, plastoquinone (PQ), cytochrome complex, plastocyanin (PC) and finally to P700.

Electron Transport flow and Non-cyclic Photophosphorylation

The electrons given out by P700 are taken up by primary acceptor and are ultimately passed on to NADP. The electrons combine with H+ and reduce NADP to NADPH2. The hydrogen ions also called protons are made available by splitting up of water. Non-cyclic photophosphorylation needs a constant supply of water molecules. The net result of non-cyclic phosphorylation is the formation of oxygen, NADPH and ATP molecules. Oxygen is produced as a waste product of photosynthesis.

Cyclic Photophosphorylation

The electrons released by P700 of PS-I in the presence of light are taken up by the primary acceptor and are then passed on to ferredoxin (Fd), plastoquinone (PQ), cytochrome complex, plastocyanin (PC) and finally back to P700 i.e., electrons come back to the same molecule after cyclic movement.

Cyclic Photophosphorylation

The cyclic photophosphorylation also results in the formation of ATP molecules just like in non – cyclic photo phosphorylation. As the electrons move downhill in the electron transport chain, they lose potential energy and ATP molecules are formed in the same way as in mitochondria during respiration.

During cyclic photophosphorylation, electrons from photosystem – I are not passed to NADP from the electron acceptor. Instead the electrons are transferred back to P700. This downhill movement of electrons from an electron acceptor to P700 results in the formation of ATP and this is termed as cyclic photophosphorylation. It is very important to note that oxygen and NADPH2 are not formed during cycle photophosphorylation.

Comparison of Non-cyclic and Cyclic PhotoPhosphorylation

Adapted From: Tutorvista.com

Chemical Compound and their Common Name

Nirmal Kumar G — Wed, 23 Mar 2011 02:15:00 +0000

S.No.	Common name	Chemical name
1	Aqua regia	Conc. HNO₃ + Conc. HCl
2	Aspirin	Acetylsalicylic acid
3	Bordeaux mixture	A solution of CuSO₄ + lime
4	Baking powder	NaHCO₃ + sodium potassium tartrate
5	Baeyer’s reagent	Alkaline KMnO₄ solution
6	Benedict solution	A solution of CuSO₄.5H₂O + NaOH + Sodium citrate
7	Coal gas	H₂ (47%) + CH₄ (32%) + CO (7%)
8	Fusion mixture	Na₂CO₃ + K₂CO₃ (laboratory reagent)
9	Gobar gas	CH₄ + CO₂ + H₂ (domestic fuel)
10	Fehling solution	CuSO₄.5H₂O + NaOH + Sodium potassium tartrate
11	Gun powder	KNO₃ (75%) + S (12%) + charcoal (9.3%)
12	Water gas	CO + H₂
13	Lucas reagent	Conc. HCl + anhyd. ZnCl₂
14	Molish reagent	α-naphthol (C₁₀H₈OH) + ethanol (C₂H₅OH)
15	Nitro lime	CaCN₂ + graphite
16	Purple of cassius	Colloidal solution of Au + SnCl₄
17	Power alcohol	(petrol + ethanol) (4:1) + little benzene (motor fuel)
18	Producer gas	N₂ (52-55%) + CO (22-30%) + H₂ (8-12%) + CO₂ (3%) (as a fuel)
19	Soda lime	NaOH + Ca(OH)₂
20	Schiff’s reagent	Colorless solution of fuchsin and sulphurous acid
21	Tincture of Iodine	I₂ + KI + ethanol + water (an antiseptic)
22	Alum	Ammonium aluminum sulphate [Al₂(SO₄)₃SO₄.(NH₄)₂.24H₂O
23	Oil of vitriol	Sulphuric acid (H₂SO₄)
24	Blue vitriol	Copper sulphate (CuSO₄.5H₂O)
25	Borax	Sodium tetraborate (Na₂B₄O₇.10H₂O)
26	Carborundum	Silicon carbide (SiC)
27	Dry ice	Carbon dioxide (solid) CO₂
28	Formalin	Formaldehyde (40% solution) HCHO
29	Gypsum	Calcium sulphate (CaSO₄.2H₂O)
30	Gammexane (BHC)	Benzene hexachloride (C₆H₆Cl₆)
31	Laughing gas	Nitrous oxide (N₂O)
32	Mohr’s salt	Ferrous ammonium sulphate (FeSO₄.(NH₄)₂SO₄.6H₂O)
33	Marsh gas (Damp-fire)	Methane (CH₄)
34	Oleum	Sulphuric acid (fuming) H₂S₂O₇
35	Plaster of Paris	Calcium sulphate hemihydrate (CaSO₄. ½ H₂O)
36	Philosopher’s wool	Zinc oxide (ZnO)
37	Tear gas	Chloropicrin (C₁₀H₅ClN₂)
38	Washing soda	Sodium carbonate (Na₂CO₃.10H₂O)
39	White vitriol	Zinc sulphate (ZnSO₄.7H₂O)
40	Paracetamol	N-(4-hydroxyphenyl) acetamide
41	RDX (cyclonite)	1,3,5-trinitroperhydro-1,3,5-triazine
42	Lindlar catalyst	Pd/BaSO₄ and S
43	Caustic soda	Sodium hydroxide (NaOH)
44	Caustic potash	Potassium hydroxide (KOH)

HeLa Cell Line

Nirmal Kumar G — Wed, 23 Mar 2011 01:59:00 +0000

HeLa term is used in conjunction with the word cell and used in the literature of the biological and medical topics. This line of cells that are derived from cancer cells taken from the patient named Henrietta Lacks, died from cancer in 1951. HeLa cell line are today used to study cancer.

HeLa cells are a human epithelial cervical cancer (cervical cancer), and the first human cells, from which a permanent cell line was established. On 9 February 1951, surgeon Lawrence Wharton Jr. removed the tissue from the patient Henrietta Lacks, a 31-year-old African American woman from Baltimore, in the Women’s Clinic of the Johns Hopkins Hospital. The cells were from the carcinoma of the cervix and were expected to be examined for malignancy. The patient died 8 months later from the disease.

A portion of cells from the biopsy were sent to George Gey, the then head of the cell culture laboratory at Johns Hopkins Hospital. The cells were cultivated and propagated in cell culture so well that since they are widely used in research. The HeLa cells were used in the establishment of the first polio vaccine by Jonas Salk. HeLa Cells are now available in many laboratories of the world. The worldwide sale of HeLa cells suggests that Ms. Lacks is probably the "most valuable" human individual had previously lived.

HeLa Cell Line used for….

Ten Popular Cell Lines used for Biological Research

HeLa isn’t the only cell line in use today. Thousands have found their way into labs worldwide. Here are some commonly used lines and the number of scientific papers they appear in.

Number of Scientific Papers

Will there be an effect of Temperature on pH scale ?

Nirmal Kumar G — Sun, 12 Sep 2010 17:03:00 +0000

Temperature and pH are two different parameters, that plays a major role in enzyme activity. Here, we will see the effect of Temperature upon the pH scale of water. We will use the concept of Ionic product of water to study this effect.

Water molecules can function as both acids and bases. One water molecule acting as a base can accept a hydrogen atom from the water molecule that acts as an acid and results in the formation of hydroxonium ion and hydroxide ion. The Ionic product of water, Kw, can be defined as the equilibrium constant for the reaction in which water undergoes an acid-base reaction with itself.

The pure water is supposed to have pH of 7 at 25^oC. At this pH and temperature, water will have a ionic product of 1 x 10^-14. The dissociation of water into hydrogen and hydoxyl ions is an endothermic process.

According to Le Chatelier’s Principle, if you make a change to the conditions of a reaction in dynamic equilibrium, the position of equilibrium moves to counter the change you have made. According to Le Chatelier, if you increase the temperature of the water, the equilibrium will move to lower the temperature again. It will do that by absorbing the extra heat.

That means that the forward reaction will be favoured, and more hydrogen ions and hydroxide ions will be formed. The effect of that is to increase the value of Kw as temperature increases.

T (^oC)	Kw (mol² dm^-6)	pH
0	0.114 x 10^-14	7.47
10	0.293 x 10^-14	7.27
20	0.681 x 10^-14	7.08
25	1.008 x 10^-14	7.00
30	1.471 x 10^-14	6.92
40	2.916 x 10^-14	6.77
50	5.476 x 10^-14	6.63
100	51.3 x 10^-14	6.14

The above table shows that, with increase in temperature, the Kw also increases, which results in the decrease of pH. But, it doesn’t mean that water becomes more acidic at higher temperatures.

At 100°C, the pH of pure water is 6.14. That is the neutral point on the pH scale at this higher temperature. A solution with a pH of 7 at this temperature is slightly alkaline because its pH is a bit higher than the neutral value of 6.14.

Similarly, you can argue that a solution with a pH of 7 at 0°C is slightly acidic, because its pH is a bit lower than the neutral value of 7.47 at this temperature.

Spectrophotometric concentrations of DNA and Protein

Nirmal Kumar G — Sat, 11 Sep 2010 18:37:00 +0000

We say this much concentration of biomolecule is present in a solution based on its optical density value. Let’s rewind about a basic concept of concentration & absorbance for DNA, RNA and Protein.

As we all know, proteins show maximum absorbance at 280nm. 1 OD of Proteins @ 280nm corresponds to 1mg/ml concentration. The ratio decreases with the interference of contaminants.

Nucleic acids show maximum absorbance at 260nm.

1 OD of dsDNA @ 260nm corresponds to 50ug/ml.

1 OD of ssDNA @ 260nm corresponds to 33ug/ml.

1 OD of ssRNA @ 260nm corresponds to 40ug/ml.

Plasma Desorption Mass Spectrometry (PDMS)

Nirmal Kumar G — Wed, 05 May 2010 03:31:03 +0000

Plasma desorption ionization mass spectrometry (PDMS; also called fission fragment ionization) is a mass spectrometry technique in which ionization of material in a solid sample by bombarding it with ionic or neutral atoms formed as a result of the nuclear fission of a suitable nuclide, typically the Californium isotope ²⁵²Cf.

Courtesy: Californium-252 Plasma Desorption Mass Spectroscopy. Science 1976, 191, 920-925.

Californium-252 plasma desorption mass spectrometry (PDMS) hasbeen employed for the characterization of a series of humaninsulin derivatives in order to evaluate the performance ofthis technique as an analytical tool in protein engineering.Several of the characterized modifications result in a 1 a.m.u.mass change. The precision in mass determination obtainableby PDMS analysis is not sufficient for unambiguous verificationof such modifications based on the molecular weight alone. Itis, however, possible to carry out in situ enzymatic digestionof the sample. Subsequent PDMS analysis will in most cases revealif the modification has been introduced as intended.

Courtesy: Protein Engineering vol. 2 no. 6 pp. 449-457, 1989

Nucleic Acids – What are they?

Nirmal Kumar G — Fri, 29 Jan 2010 09:39:00 +0000

Johann Friedrich Miescher isolated a weakly acidic substance what we now refer to as DNA in 1870 from human white blood cells. He called it as NUCLEIN. He then separated it into protein and nucleic acids. In the 1920′s nucleic acids were found to be major components of chromosomes, small gene-carrying bodies in the nuclei of complex cells. The backbone of a nucleic acid is made of alternating sugar and phosphate molecules bonded together in a long chain. The acidic character of the nucleic acids was attributed to the phosphoric acid moiety. Each of the sugar groups in the backbone is attached (via the bond shown in red) to a third type of molecule called a nucleotide base – Adenine, Guanine, Cytosine, Thymine, Uracil.

To reflect the unusual sugar component (2-deoxyribose), chromosomal nucleic acids are called deoxyribonucleic acids, abbreviated DNA. Analogous nucleic acids in which the sugar component is ribose are termed ribonucleic acids, abbreviated RNA.

Deoxyribonucleic acid (DNA):

In most living organisms (except for viruses), genetic information is stored in deoxyribonucleic acid, or DNA residing in the nucleus of living cells. DNA gets its name from the sugar molecule contained in its backbone(deoxyribose); however, it gets its significance from its unique structure. Four different nucleotide bases occur in DNA: adenine (A), cytosine (C), guanine (G), and thymine (T).

Ribonucleic Acid (RNA):

Like DNA, RNA contains the bases adenine (A), cytosine (C), and guanine (G); however, RNA does not contain thymine, instead, RNA’s fourth nucleotide is the base uracil (U). Unlike the double-stranded DNA molecule, RNA is a single-stranded molecule. RNA is the main genetic material used in the organisms called viruses, and RNA is also important in the production of proteins in other living organisms. RNA can move around the cells of living organisms and thus serves as a sort of genetic messenger, relaying the information stored in the cell’s DNA out from the nucleus to other parts of the cell where it is used to help make proteins. Three kinds of RNA are identified, the largest subgroup (85 to 90%) being ribosomal RNA, rRNA, the major component of ribosomes, together with proteins. The size of rRNA molecules varies, but is generally less than a thousandth the size of DNA. The other forms of RNA are messenger RNA , mRNA, and transfer RNA , tRNA. Both have a more transient existence and are smaller than rRNA. All these RNA’s have similar constitutions, and differ from DNA in two important respects, the sugar component of RNA is ribose, and the pyrimidine base uracil replaces the thymine base of DNA.

Adapted from: Virtual textbook of Organic Chemistry; Vision Learning.