Nitration of acetanilide (Lab report)

Introduction

Nitration is a type of chemical reaction which a nitro group is added to/substituted in a molecule. Basically it can be carried out by a mixture of concentrated nitric acid and sulphuric acid. Mixture is useful to obtain the active nitronium ion. Electrophilic aromatic substitution is a method used when a functional group is needed to be substituted on to an aromatic compound. In the nitration, nitronium ion acts as the electrophile that involves the attack of the electron-rich Benzene ring. In this experiment nitration is carried out using acetanilide.

Theory

Acetanilide displays moderately reactivity in electrophilic aromatic substitution. Also another advantage is, it’s not oxidized by nitric acid. Principally, acetanilide gives Ortho and Para mono nitroacetanilides. This position of nitronium ion is directed by the –NHCOCH₃ group attached to the benzene ring. This is due to the resonance delocalizing the benzene ring by nitrogen lone pair. Therefore Ortho and Para positions are more resonance stabilized than the Meta. Acetanilide undergoes ready nitration giving mainly the colourless P-nitroacetanilide, mixed with much smaller proportion of the yellow colour O-nitroacetanilide.

Procedure

·         About 2g of powdered acetanilide and 2mL of Glacial acetic acid were mixed well in a 100mL beaker.  4mL of con.H₂SO₄ was added to the mixture.

·         The beaker was placed in crushed ice until the temperature of the mixture was dropped down to 0-5⁰C.

·         4mL of con.H₂SO₄ was added drop wisely while stirring the viscous mixture continuously keeping the temperature below 10⁰C.

·         Afterwards the beaker was removed from the freezing conditions and allowed to stand 30 minutes at room temperature.

·         The mixture was poured onto about 20g of crushed ice and stirred to obtain crystals. The beaker was rinsed with 10mL of water containing few fragments of ice and the solution was added to the main bulk of the product.

·         It was allowed to stand for about 20minutes.

·         Later it was filtered at the pump and washed thoroughly with cold water.

·         Afterwards the crude product was recrystalized with water.

·         Finally the melting point of the product was determined.

Observations

·         Acetanilide powder was white in colour.

·         Glacial acitic acid, H₂SO₄ and acetanilide mixture was initially in pale yellow colour.

·         In cold condition, white colour crystals were formed.

·         Final purified crystals were also white.

·         The determined melting point range was 150-152⁰C.

Conclusion

The initial compounds used were known. Therefore according to those the final product should be nitroacetanilide. As the colour of the product was white, it should be Para nitroacetanilide.  

Discussion

Through the use of electrophilic aromatic substitution, acetanilide is nitrated to nitroacetanilide. There are several key steps involved in the nitration. The first step of the reaction involved in the donation of an electron pair by the acetanilide to the eletrophile, the nitronium ion. This nitronium ion was formed by the reaction of sulfuric and nitric acids. Basically the whole mechanism undergoes as below; (Benzene is shown here instead of acetanilide )

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To prevent dinitration of the acetanilide, the nitrating mixture of concentratred nitric acid and sulfuric acids were added in small portions to the acetanilide solution, so that the concentration of the nitrating agent is kept at minimum.

Also the addition of nitric acid is exothermic. Therefore the mixture would get too hot exceeding the temperature range suitable for the nitration. To avoid this, the addition of HNO₃ acid should be done very slowly, dropwisely.

Cold temperatures were used to slowdown the reaction rate and help to avoid over nitration.

Glacial acetic acid is used because it is a polar solvent capable of dissolving acetanilide and the acetate ion is a poor nucleophile, so no substitution is possible.

At the end, traces of acid should be removed because hydrogen ions catalyze the hydrolysis of the amide to p-nitroaniline. Acid is removed by pouring the mixture onto ice and water and filtering.

The melting point is determined to characterize the product. Theoretical melting point of the Para nitroacetanilide is found to be 214-216⁰C. The observed value was 150-152⁰C and it is much lower than the theoretical values and can be accounted for impurities in the product. Some impurities might be Ortho and Meta directing substances. Also there can be some experimental errors occurred during the experiment such as not controlling the exact temperatures mentioned for the reactions.

Controlling metabolism at cellular level: Compartmentation

Metabolism is composed of reaction sequences termed as metabolic pathways. The changes at each step in a metabolic pathway are regulated in order to maintain the stability and survival of the organism. Mainly there are four ways of controlling metabolism. Such one important way of controlling metabolism at cellular level is compartmentation.

When considering a eukaryotic cell; it’s composed of many different compartments known as sub cellular organelles in contrast to a prokaryotic cell. Each organelle contributes a specific metabolic function with different enzymatic activities under different conditions. These metabolic pathways needed to be interconnected to maintain the metabolism. Therefore compartmentation allows the distribution of metabolites and enzymes between the membranes bounded organelles. It’s the method that enables the cell to obtain different metabolic activities at the same time for more energy efficiency. For an example, glycolysis occurs in the cytoplasm and provides the mitochondria substrates such as pyruvates from carbohydrate oxidation. TCA cycle occurs within the mitochondrion matrix while oxidative phosphorylation occurs in mitochondria inner membrane. However, altogether all these three processes are included in one metabolism called ‘cell respiration’. Enzymes of each process are found where the processes take place.

Fat metabolism is another one good example for compartmentation. Fatty acids synthesis takes place in cytoplasm. Cytoplasm contains the required enzymes for the anabolism of fatty acid synthesis. But these fatty acids are degregaded in mitochondria supplying Acetyl Co A for TCA cycle. Mitochondria also contain the enzymes require for the catabolism of fat.

RNA polymerase allows RNA synthesis by using a template of DNA strand. It catalyzes the synthesis of RNA. RNA synthesis takes place in nucleus. DNA transcription also occurs in nucleus. Therefore both the RNA polymerase and DNA templates are available, so that the RNA synthesis is favorable. When considering protein synthesis, all the organelles used for it are close to one another. The nucleolus makes ribosome which synthesizes proteins. Endoplasmic reticulum is near the nucleus to receive Ribosomes and synthesized proteins. The Golgi apparatus is also near the endoplasmic reticulum for storing, packaging and distribution. Therefore all the activities within a cell are cooperated with one another.

Hence, compartmentation helps metabolic controlling by maintaining substrates, regulators and enzymes in separate locations with only favoured access between them.

General facts of parasitic nematodes:- Pin worm, giant round worm, whip worm, pork worm, hook worm

Enterobius sp.

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Common name: Pin worm/thread worm/ seat worm

Disease: Enterobiasis (symptoms:- itching in the preanal area)

Host: Human

Habitat: Small intestine

Infective stage: Eggs

Transmission: Ingesting eggs

Parasitic adaptations: Small, sticky, translucent eggs, cuticle

Enterobiu sp -Life cycle 

Trichuris trichuira

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Common name: Whip worm/tap wrom

Disease: Trichuriasis ( Symptoms: Diarrhea, anemia)

Host: Human

Habitat: Colon

Infective Stage: Embryonated eggs

Transmission: Ingesting eggs

Parasitic adaptations: Long slender tapered body, cuticle

Trichinella sp

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Common name: Pork worm

Disease: Trichinosis (Symptoms: nausea, diarrhea, vomiting, fever)

Host: Human, pig, bear, rat

Habitat: Intestine

Infective stage: Cysts (larva)

Transmission: Eating uncooked pork, 

Parasitic adaptations: Long slender body, cuticle

image via dpd.cdc.gov

Ascaris sp

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Common name: Giant round worm

Disease: Ascariasis (Symptoms: Diarrhea, cough, nausea)

Host: Human

Habitat: Intestine

Infective stage: Embryonated eggs ( containing larva)

Transmission: Ingesting eggs

Parasitic adaptations: Long slender body, cuticle, lipid cover around the eggs which is resistant to abrasion and chemicals

Ancylostoma sp

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Common name: Hook worm

Disease: Protein deficiency, anemia, cough

Host: Human, cat

Habitat: Intestine

Infective stage: Filari form larva

Transmission: Penetrating the intact skin of the host

Parasitic adaptations: Dental plates in mouth, slender body, cuticle

Plant senescence in brief

Senescence

Senescence is an important program in which the function of a cell/tissue/whole plant naturally leads to its death.

Senescence involves in plant aging.

When tissues senesce they produce enzymes which can recycle the “expensive” materials and reroute the sub-units to somewhere else that growth takes place.

Overall senescence

·         Occurs in the whole plant.

·         Death takes place right after flower and setting.

·         When the flower dies the whole plant dies too.

·         Ex: Asteraceae ( Sun flower)

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Top senescence

·         After the growth season the part above the ground dies remaining the underground part that can grow for several years.

·         Therefore a rhizome is present.

·         Ex: Weed, Grass

       

Deciduous senescence

·         In dry seasons like winter and summer, the leaves falls and trunk remains.

·         Leaves fall due to avoid the heavy transpiration occurs in the dry season.

·         Ex: Deciduous plants

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Progressive senescence

·         Older parts die due to the senescence and the new organs and tissues continue their development.

·         Ex: Green trees

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As above mentioned, senescence can occur in cells as well as tissues.

Cell senescence

·         Can be described according to the senescence in cell membrane and organelle level.

Senescence in cell membrane

Lipid phase change

·         Plasma membrane has the fluid mosaic structure.

·         This structure changes as the liquid crystalline state changes into the solid-gel state.

·         Therefore the membrane gets inflexible and hard.

Degradation and preoxidation of lipids

·         Lipid content gets decreased.

·         It occurs as the lipid synthesis decreases and lipase content increases.

·         Phospholipase, lipoxygenase, MDA and active O₂ content increase.

Increase in Phospholipase activity

·         Mainly the activity of Phospholipase D takes place in plants.

Biomembrane degradation and leakage

·         The equilibrium of ions breaks down.

·         Metabolisms get disordered.

Senescence in organelle level

·         Ribosomes and RER break down.

·         Chloroplasts collapse and mitochondrion cristae are swollen.

·           As the vacuoles break down, all the toxics inside them is released to the cell plasma.

·         Therefore the autophagy takes place and the senescence of organelles activates.