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Steroids: cholesterol,

Bile acids and steroid hormones

Vitamin D

1. Structure and stereochemistry of steroids

2. Cholesterol as a precursor of bile acids, steroid hormones andvitamin D3

3. Structure of the primary and secondary bile acids, conjugation withglycine and taurine

4. Steroid hormones - progestins, glucocorticoids, mineralocorticoids,androgens, estrogens

5. Vitamin D and its active forms

Structure and stereochemistry of steroids

Steroids are derivatives of theperhydrocyclopentanophenanthrene ringsystem.

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STEROID NUMBERING SYSTEM

A B

C D1

2

3

45

6

7

89

10

1112

13

14 15

16

1718

19

Steroid ring

• The steroid ring is notplanar, but flattenedin away that it extends as muchas possible.

• All the rings are in as muchof a chair conformation aspossible.

• There is no rotation possiblearound bonds in the rings,which makes sterols prettyrigid, except for the chain atC17.

Conformation

• There are four rings in a steroid skeleton and hence there are three fusion points. A/B, B/C and C/D ringsshare two carbons each (fusion).

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An A-B cis steroid

The methyl groups are attached at points of ring junction and are called angular methyl groups.

Other groups on the same side of the steroid plane as the angular methyl groups are said to be β substitutents

Groups below the plane of the steroid ring system are said to be α substituents

The main sterol found in the membranes of eukaryotic cells is cholesterol. The polar head in this lipids is the hydroxyl in carbon C3.

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Cholesterol

One of the most widely occuring steroids, was first isolatedin 1770.

Contains 8 chiral C atoms, this means that 28 or 256 stereoisomers arepossible, but only one of them is cholesterol.

Contains 27 carbons

Cholesterol sources, biosynthesis and degradation

• Slightly less than half of the cholesterolin the body derives from biosynthesis de novo.

• Biosynthesis in the liver accounts for approximately 10%, and in the intestine approximately 15%, of the amount produced each day.

• Cholesterol synthesis occurs in the cytoplasmand microsomes from the two-carbon acetate group of acetyl-CoA.

Biosynthesis of Cholesterol

The process has five major steps: • 1.Acetyl-CoAs are converted to 3-hydroxy-3-methylglutaryl-

CoA (HMG-CoA)

• 2. HMG-CoA is converted to mevalonate

• 3. Mevalonate is converted to the isoprene based

molecule, isopentenyl pyrophosphate (IPP), with

the concomitant loss of CO2

• 4. IPP is converted to squalene

• 5. Squalene is converted to cholesterol.

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Acetyl CoA is the source of all carbon atoms in cholesterolAcetyl CoA is the source of all carbon atoms in cholesterol

Squalene

β -hydroxy- β- methylglutaryl CoA

Mevalonate

Farmesyl pyrophosphate

Acetyl CoA

CoA

Acetoacetyl CoA

CoA

Acetyl CoA

HMG-CoA

reductase

Cyclization

HMG-CoA Reductase

HMG-CoA reductase

1. integral membrane protein in the ER

2. carries out an irreversible reaction

3. is an important regulatory enzyme in cholesterol synthesis

Fate of Cholesterol

• Incorporated into hepatocyte membranes

• Exported– Bile acids

– Cholesterol esters

– Free cholesterol

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Cholesterol Esters

• Acyl-CoA:cholesterolacyl transferase(ACAT)is an ER membraneprotein

• ACAT transfers fatty acidof CoA to C3 hydroxylgroup of cholesterol

• Excess cholesterol isstored as cholesterolesters in cytosolic lipiddroplets

Fig. 8

The physiological roles of cholesterol include:

a) an important lipid component of biological membranes, b) precursor of steroid hormones and c) source of bile acids.

ROLES OF CHOLESTEROL AND BILE ACIDS (SALTS)

Bile acids are polar derivatives of cholesterol and aid in:

a) lipid digestionb) lipid absorptionc) cholesterol excretion

Bile Acids Synthesis and Utilization

• The end products of cholesterolutilization are the bile acids, synthesized in the liver.

• Synthesis of bile acids is one of the predominant mechanisms for the excretion of excess cholesterol.

• However, the excretion of cholesterol in the form of bile acids is insufficient to compensate for an excess dietary intake of cholesterol.

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Bile

• Bile is formed in the liver andsecreted down the bile duct intothe gallbladder, from where itpasses into the intestine

• The gallbladder stores bileproduced by the liver betweenmeals.

• During digestion, the gallbladdercontracts and supplies bilerapidly to the duodenum by wayof the common bile duct

Bile

• Bile is a complex solution of salts and protein, containing micelles composed of cholesterol, phospholipids and bile salts.

• The composition of thesemicelles is critical, as imbalancemay lead to crystalization ofcholesterol in the gallbladder,leading to the formation ofgallstones.

GALLSTONES

• Cholesterol gallstones can form as a result of excessivesecretion of cholesterol or of insufficient amounts of bile acidsand lecithin relative to cholesterol in bile

• If the amount of cholesterol in bile acids getabove 15%, thengallstones may result.

• If cholesterol gets too high, then they may crystallize out ofsolution in the gall bladder, where bile is concentrated tenfoldbefore secretion.

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Functions of Bile Salts� Important for cholesterol excretion:

1. As metabolic products of cholesterol

2. Solubilizer of cholesterol in bile

� Emulsifying factors for dietary lipids,a prerequisite step for efficient lipid digestion

� Cofactor for pancreatic lipase and PLA2

� Facilitate intestinal lipid absorption byformation of mixed micelle

Bile salts• detergent character of bile salts is due to the hydrophobic-hydrophilic

nature of the molecules• the presence of hydroxyl (or sulfate) and the terminal carboxyl group on the

tail gives the molecule its hydrophilic face• the steroid ring with its puckered plane provides the hydrophobic face

Emulsification of Dietary Lipidsin Duodenum: Role of Bile Salts

• Emulsification increases the surface area of lipid droplets,therefore the digestive enzymes can effectively act.

• Mechanisms:1. Mechanical mixing by peristalsis

2. Detergent effect of bile salts:Bile salts interact with lipid particles and aqueousduodenal contents, stabilizing the particles as theybecome smaller, and preventing them from coalescing.

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Absorption of Lipids by Intestinal Mucosal Cells: Role of Bile salts

Mixed micelles:�Disc-shaped clusters of amphipathic lipids. �Arranged with their hydrophobic groups on the inside and their hydrophilic groups on the outside.

�Micelle includes end products of lipid digestion, bile salts and fat-soluble vitamins

Note: Short- and medium-chain fatty acids do not requiremixed micelle for absorption by intestinal cells

The bile acids are amphipathic, with detergent properties.

They emulsify fat globules into smaller micelles, increasing the surface area accessible to lipid-hydrolyzing enzymes.

Synthesis of Bile Salts

• Rate-limiting step performed by the 7α-hydroxylase (CYP7A1) and isregulated by bile salt concentration

• End product: Cholic acid series & Chenocholic acid series• Bile salts can be conjugated & become better detergents

Fig. 9 Fig. 10

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Synthesis of bile salts

Primary bile acids

Bile Salts

• Bile acids & salts are effective detergents• Synthesized in the liver• Stored & concentrated in the gallbladder• Discharged into gut and aides in absorption of

intraluminal lipids, cholesterol, & fat soluble vitamines• Bile acid refers to the protonated form while bile salts

refers to the ionized form– The pH of the intestine is 7 and the pKa of bile salts is 6,

which means that 50% are protonated

• These terms are sometimes used interchangeably

SYNTHESIS OF BILE SALTS

• Bile salts are just bile acids with the carboxylatefunction modified by adding an amine to it via anamide linkage.

• Bile Salts are even more polar than theircorresponding acids.

• Glycocholate: Cholate with glycine added as anamide function.

• Taurocholate: Cholate with Taurine added as anamide function. Taurine is a modified cysteine.

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Primary and secondary bile acids

• Chenodeoxycholic acid (45%) and cholic acid (31%)are referred to as the primary bile acids.

• Within the intestines the primary bile acids are acted upon by bacteria and converted to the secondary bile acids, identified as deoxycholate (from cholate) and lithocholate (from chenodeoxycholate).

• Both primary and secondary bile acids are reabsorbed by the intestines and delivered back to the liver via the portal circulation.

Types of Bile Acids/Salts

• Primary bile acids– Good emulsifying agents

• All OH groups on same side• pKa = 6 (partially ionized)

• Conjugated bile salts– Amide bonds with glycine

or taurine– Very good emulsifier

• pKa lower than bile acids

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Primary Bile Acids and Salts

Chenodeoxycholic acid

Bile salts (Conjugated bile acids):amide-linked with glycine or taurine

The ratio of glycine to taurine forms in the bile is3:1

GlycocholicTaurocholic

GlycochenodeoxycholicTaurochenodeoxycholic

Cholic acid BILE ACIDS

BILE SALTS

Bile acids are excreted in conjugated form, as taurocholic acid and glycocholic acid

Bile acids are conjugated with glycine or taurine before being secreted into bile, where the ratio of glycine to taurine-conjugated acid is about 3:1

Fate of the bile salts and the synthesis of secondary bile acids

�Intestinal bacteria deconjugate and dehydroxylate the bile salts removingtheglycine and taurine residues and the hydroxyl group at position 7.The secondary bilesalts are less soluble and less readily resorbed from the intestinal lumen.

�Lithocholic acid has a hydroxyl group only at position 3, is the least soluble bileacid, its major fate is excretion.

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Secondary Bile Acids

Bile salts Glyco- or Tauro-cholate -Chenodeoxycholate

Bile acids Cholic acid Chenodeoxycholic

2° Bile acids Deoxycholic acid Lithocholic

Intestinal bacteria

Intestinal bacteria

GlycineTaurine

OH

Primary bile acids Secondary bile acids

•Within the intestines the primary bile acids are convertedby bacteria into thesecondary bile acids, identified asdeoxycholate(from cholate) andlithocholate(from chenodeoxycholate).

Steroids

Grups –OH localized at:

C-3 C-7 C-12

Primary bile acids:

cholic acid OH OH OH

chenodeoxycholic acid OH OH H

Secondary bile acids:

deoxycholic acid OH H OH

lithocholic acid OH H H

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The pK of the bile acids is about 6.

�Therefore, in the contents of the intestinal lumen, which normally have a pH of 6,about half of the molecules are present in the protonated form and half are ionized,forming bile salts.

�The carbonyl group at the end of the side chain is activated bya reaction thatrequires ATP and coenzyme A.

�The acyl CoA derivatives can react with eitherglycine or taurine, formingamides that are known as the conjugated bile salts.

�The bile acids conjugated with glycine have a pK of about 4, so a higher percentage of the molecules is present in the ionized form at the pH of the intestine.

�The taurine conjugates have a pK of about 2, and even greater percentage of the molecules of these conjugates are ionized in the lumen of the gut.

�Taurine is synthesized from the amino acid cysteine.

Clinical Significance of Bile Acid Synthesis

• Bile acids perform four physiologically significant functions: • 1.Their synthesis and subsequent excretion in the feces

represent the only significant mechanism for the elimination of excess cholesterol.

• 2. Bile acids and phospholipids solubilize cholesterol in the bile, thereby preventing the precipitation of cholesterol in the gallbladder.

• 3.They facilitate the digestion of dietary triacylglycerols by acting as emulsifying agents that render fats accessible to pancreatic lipases.

• 4.They facilitate the intestinal absorption of fat-soluble vitamins.

•the primary and secondary bile acids are reabsorbed almost exclusively in the ileum returning to the liver by way of the portal circulation (98 to 99%)•the entherohepatic circulation

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Bile salts are synthesized in theliver, stored in the gallbladder,secreted into the small intestine,resorbed in the ileum, andreturned to the liver.

Only 5% are excreted.

Recycling of bile salts

Cholesterol

�An amphipathic lipid , an essential structural component of membranes and theouther layer of plasma lipoproteins,

�Present in tissues and lipoproteins either asa free cholesterol or cholesteryl ester,

�Lipoproteins transport free cholesterol in the circulation, where it readily equili-brates with cholesterol in other lipoproteins and in membranes,

�Cholesteryl ester is a storage formof cholesterol in most tissues,

�Low density lipoprotein (LDL) is the mediator of cholesterol and cholesteryl esteruptake into many tissues,

�Free cholesterol is removed from tissues by high density lipoprotein (HDL) andtransported to the liver for conversion to bile acids,

�Excess cholesterol is excreted from the liver in the bile as cholesterol or bile salts;a large proportion of bile salts is absorbed into the portal vein and returned tothe liver as part of the entherohepatic circulation,

�Elevated levels of cholesterol are associated withatherosclerosis

Cholesterol in the cell membrane

Cholesterol is also present in the membrane. It maintains the fluidity and increases the

stability of the membrane. Without cholesterol the membrane would easily split apart

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The Utilization of Cholesterol• Cholesterol is transported in the plasma predominantly as cholesteryl esters

associated withlipoproteins.

• Dietary cholesterol is transported from the small intestine to the liver withinchylomicrons.

• Cholesterol synthesized by the liver, as well as any dietarycholesterol inthe liver that exceeds hepatic needs, is transported in the serum within

LDLs (Low density lipoproteins).

The Acyl-Co: cholesterol acyltransferase (ACAT) reaction (Synthesis of Cholesterol Esters by ACAT)

Located mainly in the endoplasmic reticulum ACAT catalyzes the covalent joining of cholesterol and long-chain fatty acyl-CoA to form cholesterol esters.

Cholesterol ester products of the ACAT reaction are either stored in cytosolic droplets or secreted from cells as components of apo-B lipoproteins.

HDL

�High-density lipoprotein (HDL) begins in the liver and smallintestine as small protein rich particles containing relatively littlecholesterol and cholesterol ester

�HDLs contain apoA-I and apoA-II, as well as the enzymelecithin-cholesterol acyltransferase (LCAT) which catalyzes theformation of cholesterol ester

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Functions of HDL

• Transfers proteins to other lipoproteins

• Picks up lipids from other lipoproteins

• Picks up cholesterol from cell membranes

• Converts cholesterol to cholesterol esters via theLCATreaction

• Transfers cholesterol esters to other lipoproteins, whichtransport them to the liver (referred to as “reverse cholesteroltransport)

Reverse Cholesterol Transport

• An enzyme present in our circulation,Lecithin CholesterolAcyl Transferase (LCAT), removes one fatty acid fromPhosphatidylcholine (PC) and attaches it to cholesterol,converting cholesterol into a cholesterol ester, which is thentaken up by a HDL lipoprotein and transported back to theliver, from where the cholesterol is excreted from the body inthe form of a bile salt

Acyltransferase lecithin : cholesterol (LCAT)

The reaction catalyzed by LCAT. This enzyme is present on the surface of HDL, and stimulated by the HDL component of apoA-1.

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Lipoproteins and Lipid Transport

Steroid hormones

Steroid hormones are grouped into five categories:

- progestins,- glucocorticoids,- mineralocorticoids,- androgens,- estrogens.

�All contain the four-ring structure of the steroid nucleus and areremarkably similar in structure, considering the enormous differencesin their physiological effects.

�They mediate a wide variety of vital physiological functions.

FUNCTIONS: Steroids Hormones

• Cholesterol: precursor of the five major

classes of steroid hormones

• Glucocorticoid : carbohydrate metabolism

• Mineralocorticoid: Sodium retention in

renal tubules

• Androgen, Estrogen, Progestagens:

reproductive system

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Steroid hormones

�The primary organs involved in their biosynthesis arethe gonadsand the adrenal cortex, andthe placentain pregnant females.

�In steroid hormones, the side chain of cholesterol is eithershortened or nonexistent.

�Steroid hormones (SH) are not stored for release after synthesis.

�The level of circulating SH is controlled by its rate of synthesis,which is controlled by signals from the brain.

�Activation of SH synthesis involves stimulation of bothhydrolysis of cholesterol esters and uptake cholesterol intomitochondria of cells in the target organ.

SYNTHESIS OF STEROID HORMONESCHOLESTEROL ------> PREGNENOLONE

• Removal of 6 of the side-chain carbons.

This step is common to all steroid hormone synthesis.

• This step occurs on the mitochondrial membrane.

• This leaves a C21 compound,pregnenolone, which may accurately bedescribed as the mother of all steroids(the C21 steroid carbon skeleton is giventhe namepregnane).

CHOLESTEROL ------> PREGNENOLONE

Steroids

There is an enzyme complex called cholesterol desmolase that hydroxylates C-20 and C-22 and cleaves it to yield pregnolone

Desmolase (in mitochondria) forms pregnenolone, precursor to all others

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activated to turn on pathways

Cholesterol Pregnenolone Progesterone

11-Deoxy-cortisone

21-hydroxylase

Progesterone Aldosterone

General pathways for the synthesis of aldosterone and cortisol in the adrenal cortex

11-Deoxy-cortisol

Progesterone Cortisol

21-hydroxylase

Estrone(produced in both male and

female adipose cells)

Pathways for the synthesis of testosterone (testes) and the estrogens estradiol (ovaries) and estrone (adipose cells)

Cholesterol Pregnenolone Progesterone

Progesterone Androstenedione Testosterone(pathway ends here in testes)

Estradiol(pathway continues to

here in ovaries)

In obese men, overproduction of estrogen in fat cells can cause gynecomastia = excessive male breast development

Transformations of Cholesterol: Steroid Hormones

O

O

O

OH

OHHO

O

CH3

HO

CH3

Cholesterol

Estradiol

Progesterone

Cortisol

O

OH

TestosteroneHO

OH

CH2

HO

OH

OHVitamin D

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Groups of steroid hormones

1. Progestins – regulate events during pregnancy,progesterone is themajor hormone involved in sustaining pregnancy

2. Glucocorticoids –affect basal metabolism, host defense mechanisms,blood pressure, response to stress, promote gluconeogenesis and supressinflammation reactions (cortisol, corticosterone)

3. Mineralocorticoids – affect electrolyte balance and ion transport,regulate ion balance by promoting reabsorption in the kidney of Na+ ,Cl-, HCO3

- (aldosterone)

4. Androgens – promote male sexual differentiation, spermatogenesis, andmaintain male characteristics (androstenedione, testosterone),

5. Estrogens– female sex hormones, stimulate the development of tissuesinvolved in reproduction, support femal characteristics (estrone,estradiol)

Table 1. Functions of Hormones Derived from Cholesterol

Product Functions

Progesterone prepares uterus lining for implantation of ovum

Glucocorticoids (cortisol)(produced in adrenal cortex)(catabolic steroid)

promote gluconeogenesis; favor breakdown of fat and protein (fuel mobilization); anti-inflammatory

Mineralocorticoids(aldosterone) (produced in adrenal glands)

maintains blood volume and blood pressure by increasing sodium reabsorption by kidney

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Androgens (strongest = testosterone)(produced in testes primarily but weak androgens in adrenal cortex) (anabolic steroid)

development of male secondary sex characteristics; prevents bone resorption

Estrogen(produced in ovaries primarily but also in adipose cells of males and females)

development of female secondary sex characteristics; prevents bone resorption

Vitamin D (not a steroid hormone)(produced in the skin in response to UV light and processed to active form in kidney)

intestinal calcium absorption; promotes bone formation; prevents phosphate loss by kidneys

Product Functions

Table 1. Functions of Hormones Derived from Cholesterol

Steroid hormones act in the nucleus to change gene expression

�Steroid hormones are too hydrophobic to dissolve readily inthe blood, are carried on specific carrier proteins from the pointof their release to their target tissues

�In the target tissue, these hormones pass through the plasmamembrane by simple diffusion and bind to specific receptorproteins in the nucleus

�The hormone-receptor complexes act by binding to highlyspecific DNA sequences and altering gene expression

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Vitamin D• Vitamin D is a steroid prohormone, by various metabolic

changes in the body it gives rise to a hormone known as calcitriol, which play a central role in calcium and phosphate

metabolism.

Formation of provitamin D(7-dehydrocholesterol)

�D Vitamins are generated fromthe provitamins ergosterol and 7-dehydrocholesterolby the action of sunlight.

�Ergosterol occurs in plants and 7-dehydrocholesterol in animals.

�UV light from sunlight cleaves the B ring of both compounds.

�Ergocalciferol (vitamin D2) is formed in plants.

�Cholecalciferol (vitamin D3) is formed in exposed skin.

A specific transport protein called the vitamin D-binding protein binds vitamin D3 and moves its from the skin or intestine to the liver, where it undergoes 25-hydroxylation

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HO

Vitamin D3

Diet

HO

OH

25(OH) D3

Liver

25-hydroxylase

OHHO

OH

1,25(OH)2 D3(active hormone form)

Kidney1-hydroxylase

HO 7

Provitamin D3(7-dehydrocholesterol: Intermediate in cholesterol synthesis)

UV fromsunlight

Skin

Figure 7. Photobiosynthesis of vitamin D3 and its metabolism

Specific receptors in intestine, bone, kidney

Ca:Intestinal absorptionRenal reabsorption

PO4:Intestinal absorption Renal reabsorption

1,25-dihydroxycholecalcyferol (active form of D3)

• Active VITAMIN D3

– B-Ring opens

– 1-Hydroxylation

– 25-Hydroxylation

The product (1α,25 dihydroxyD3) is the most potent vitamin D metabolite.

Its production is regulated by itsown concentration, parathyroidhormone, and serum phosphate.

vitamin D-deficiency diseases

�Deficiency of vitamin Dcauses rickets andosteomalacia.