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Biochemistry

Ampholytes, Polyampholytes, pI and Zwitterion

Many substances in nature contain both acidic and basic groups as well as many different types of these groups in the same molecule. (e.g. proteins). These are called ampholytes (one acidic and one basic group) or polyampholytes (many acidic and basic groups). Proteins contains many different amino acids some of which contain ionizable side groups, both acidic and basic. Therefore, a useful term for dealing with the titration of ampholytes and polyampholytes (e.g. proteins) is the isoelectric point, pI. This is described as the pH at which the effective net charge on a molecule is zero.

For the case of a simple ampholyte like the amino acid glycine the pI, when calculated from the Henderson-Hasselbalch equation, is shown to be the average of the pK for the a-COOH group and the pK for the a-NH2 group:

pI = [pKa-(COOH) + pKa-(NH3+)]/2

For more complex molecules such as polyampholytes the pI is the average of the pKa values that represent the boundaries of the zwitterionic form of the molecule. The pI value, like that of pK, is very informative as to the nature of different molecules. A molecule with a low pI would contain a predominance of acidic groups, whereas a high pI indicates predominance of basic groups.

 Ketogenic Only

Amino Acid
Leucine (L)
Lysine (K)

 Mnemonic: "LL is Keto" (Leucine, Lysine)

Glucogenic Only

Amino Acids
Most others except those listed below as ketogenic or both

Both Ketogenic & Glucogenic

Amino Acid
Isoleucine (I)
Phenylalanine (F)
Threonine (T)
Tryptophan (W)
Tyrosine (Y)

Mnemonic: "I Prefer To Try Tasty Yogurt" (I, Phe, T, Trp, Tyr)

Enzyme assays measure the activity or concentration of specific enzymes in blood or tissue samples. Elevated or reduced levels often indicate organ dysfunction or cellular damage.

Enzyme Groups & Their Clinical Significance

Enzyme Group Function Clinical Relevance
Oxidoreductases Catalyze oxidation-reduction reactions Liver, cardiac, and muscle injury markers
Transferases Transfer functional groups between molecules Liver and muscle enzymes (e.g., AST, ALT)
Hydrolases Break chemical bonds using water Pancreatic enzymes (e.g., amylase, lipase)
Lyases Break bonds without hydrolysis or oxidation Less commonly used in diagnostics
Isomerases Rearrange molecular structures Rarely used clinically
Ligases Join molecules using ATP Mostly research-based, not routine diagnostics

PHOSPHOLIPIDS

These are complex or compound lipids containing phosphoric acid, in addition to fatty acids, nitrogenous base and alcohol 

There are two  classes of phospholipids

1. Glycerophospholipids (or phosphoglycerides) that contain glycerol as the alcohol.

2. Sphingophospholipids (or sphingomyelins) that contain sphingosine as the alcohol

Glycerophospholipids

Glycerophospholipids are the major lipids that occur in biological membranes. They consist of glycerol 3-phosphate esterified at its C1 and C2 with fatty acids. Usually, C1 contains a saturated fatty acid while C2 contains an unsaturated fatty acid.

In glycerophospholipids, we refer to the glycerol residue (highlighted red above) as the "glycerol backbone."

Glycerophospholipids are Amphipathic

Glycerophospholipids are sub classified as

1. Phosphatidylethanolamine or cephalin also abbreviated as PE is found in biological membranes and composed of ethanolamine bonded to phosphate group on diglyceride.

 

2. Phosphatidylcholine or lecithin or PC which has chloline bonded with phosphate group and glycerophosphoric acid with different fatty acids like palmitic or hexadecanoic acid, margaric acid, oleic acid. It is a major component of cell membrane and mainly present in egg yolk and soy beans.

3. Phosphatidic acid (phosphatidate) (PA)

It consists of a glycerol with one saturated fatty acid bonded to carbon-1 of glycerol and an unsaturated fatty acid bonded to carbon-2 with a phosphate group bonded to carbon-3.

4.Phosphatidylserine (PS)

This phospholipid contains serine as an organic compound with other main components of phospholipids. Generally it found on the cytosolic side of cell membranes.

5. Phosphoinositides

It is a group of phospholipids which are negatively charged and act as a a minor component in the cytosolic side of eukaryotic cell membranes. On the basis of different number of phosphate groups they can be different types like phosphatidylinositol phosphate (PIP), phosphatidylinositol bisphosphate(PIP2) and phosphatidylinositol trisphosphate (PIP3). PIP, PIP2 and PIP3 and collectively termed as phosphoinositide.

6. Cardiolipin :

lt is so named as it was first isolated from heart muscle. Structurally, a cardiolipin consists of two molecules of phosphatidic acid held by an additional glycerol through phosphate groups. lt is an important component of inner mitochondrial membrane. Cardiolipin is the only phosphoglyceride that possesses antigenic properties.

Titration of a weak acid with a strong base

• A weak acid is mostly in its conjugate acid form

• When strong base is added, it removes protons from the solution, more and more acid is in the conjugate base form, and the pH increases

• When the moles of base added equals half the total moles of acid, the weak acid and its conjugate base are in equal amounts. The ratio of CB / WA = 1 and according to the HH equation, pH = pKa + log(1) or pH = pKa.

• If more base is added, the conjugate base form becomes greater till the equivalance point when all of the acid is in the conjugate base form.

PROPERTIES OF TRIACYLGTYCEROLS

1. Hydrolysis : Triacylglycerols undergo stepwise enzymatic hydrolysis to finally liberate free fatty acids and glycerol.

The process of hydrolysis, catalysed by lipases is important for digestion of fat in the gastrointestinal tract and fat mobilization from the adipose tissues.

2. Saponification : The hydrolysis of triacylglycerols by alkali to produce glycerol and soaps is known as saponification.

3.Rancidity: Rancidity is the term used to represent the deterioration of fats and oils resulting in an unpleasant taste. Fats containing unsaturated fatty acids are more susceptible to rancidity.

Hydrolytic rancidity occurs due to partial hydrolysis of triacylglycerols by bacterial enzymes.

Oxidative rancidity is due to oxidation of unsaturated fatty acids.

This results in the formation of unpleasant products such as dicarboxylic acids, aldehydes, ketones etc.

 

Antioxidants : The substances which can prevent the occurrence of oxidative rancidity are known as antioxidants.

Trace amounts of antioxidants such as tocopherols  (vitamin E), hydroquinone, gallic acid and c,-naphthol are added to the commercial preparations of fats and oils to prevent rancidity. Propylgallate, butylatedhydroxyanisole (BHA)  and butylated hydroxytoluene (BHT) are the antioxidants used in food preservation.

Lipid peroxidation in vivo: In the living cells, lipids undergo oxidation to produce peroxides and free radicals which can damage the tissue. .

The free radicals are believed to cause inflammatory diseases, ageing, cancer , atherosclerosis etc

Iodine number : lt is defined as the grams (number)  of iodine absorbed by 100 g of fat or oil. lodine number is useful to know the relative

unsaturation of fats, and is directly proportional to the content of unsaturated fatty acids

Determination of iodine number will help to know the degree of adulteration of a given oil

Saponification number : lt is defined as the mg  (number) of KOH required to hydrolyse (saponify) one gram of fat or oiL

Reichert-Meissl (RM)  number: lt is defined as the number of ml 0.1 N KOH required to completely neutralize the soluble volatile fatty acids distilled from 5 g fat. RM number is useful in testing the purity of butter since it contains a good concentration of volatile fatty acids (butyric acid, caproic acid and caprylic acid).

Acid number : lt is defined as the number of mg of KOH required to completely neutralize free fatty acids present in one gram fat or oil. In normal circumstances, refined oils should be free from any free fatty acids.

Enzyme Clinical Use Indicates
ALT (SGPT) Specific to liver injury Hepatitis, liver necrosis
AST (SGOT) Found in liver, heart, muscle Liver disease, myocardial infarction
ALP Bile duct obstruction, bone disease Cholestasis, bone metastases
GGT Alcoholic liver disease, biliary obstruction Alcohol abuse, hepatobiliary disease
LDH Non-specific, elevated in many conditions Liver, cardiac, or hemolytic disorders

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