Digitoxin, a “cardiac glycoside”, is used to treat congestive heart failure andcardiac arrhythmia

Derived from cholesterol, with shortening and lactonization of side chain

Digitoxin competes with K+ for a binding site on enzyme potassium-ATPase(potassium blocker/antagonist) inhibiting Na-K ATPase pump

By blocking this activity, it causes Ca2+ to be transported out more slowly;so heart muscle is exposed to Ca2+ for a longer period of time

As a result, the heart contracts more forcefully (“inotropic effect”)

It also increases cholinergic stimulation to the heart, which slows it down(steadier, stronger heartbeat with more rest between beats)

Too high a dose of Digitalis can be toxic, but the therapeutic dose is about 50% of the toxic dose so it must be taken with care.Similar compounds have been used as arrow poisons!

Tetraterpene precursorphytoene is made the sameway by joining 2 GGPPs

Triterpenes: Precursor squalene is made by tail-to-tail joining of two FPPs

precursor to cholesterol& other animal steroids

precursor to manyplant sterols

How folding and cyclizations lead to sterols & triterpenes

Folding pattern leading to protosteryl cation, precursor of cycloartenol & lanosterol

Precursor to ginsenosidesfound in Panax ginseng (p.224)

Pentacyclictriterpenes

Folding pattern leading to most pentacyclic triterpenoids in plants

Skeletal numbering system of pentacyclic triterpenoids

Some bioactive pentacyclic triterpenoids

Triterpenoidsaponins have sugars attached to 3-OH giving them soapy properties.

Some triterps are esterified at the 3-OH

Betulinic acidUrsolic acid Oleanolic acidR = H = Ursolic acidR = OH = Pomolic acid

R

Cancer fighter from Cranberry Waste???

• Ursolic acid in cranberries first reported 1934

• Waxy component of peel

• 1950’s: triterpenoids isolated from cranberry pulp (Laakso)

• Considered “wastes” from the processing of berries to make juice

• What do we know about ursolic acid today?

Ursolic acid affects numerous pathways associated with inflammation and carcinogenesis

in vitro Inhibits proliferation of many tumor cell lines Induces apoptosis in liver, leukemia, human prostate cancer models,

arrests cells in G1 phase; caspase activation, c-IAP down-regulation observed

Decreases MMP-9 expression in human fibrosarcoma (may decrease tumor metastasis)

Decreases COX-2 expression (anti-inflammation) Inhibits activation of NF-kB (anti-inflammation)in vivo Decreases inflammation in mouse-paw edema model Decreases FSaII murine fibrosarcoma growth in mouse model (IV

administration) Diet containing 0.10% UA delayed tumor appearance, significantly

decreased tumor volume in mice (DeAngel, et al, 2010)

Ursolic acid (UA)

Ursolic acid & its hydroxycinnamoyl esters are plentiful in cranberry peels

cis-3-O-p-hydroxycinnamoyl ursolic acid (cis-HC-UA)

trans-3-O-p-hydroxycinnamoyl ursolic acid (trans-HC-UA)

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COOH Content in cranberry products, other Vaccinium berries, inhibition of matrix

metalloproteinasesKondo, et al, JFSA, 2011

Cytotoxicity in tumor cells and structures elucidation

by 1H & 13C NMR;Murphy, et al, JAFC, 2003

NCI screening in tumor cell lines showsHC-UA esters antiproliferative at M concentrations

Cell line Tumor type GI50 (M) GI50 (M)

cis HC-UA trans HC-UA

MCF-7 Breast 1.4 1.9

MDA-MB-435 Breast 4.3 8

DU-145 Prostate 4.1 5.3

NCI-H460 Lung 2.2 2.6

NCI-H322M Lung 1.2 10

HT-29 Colon 5 19

HCT-116 Colon 1.5 3.5

RPMI-8226 Leukemia 1.8 6

SF-295 CNS 3.0 2.7

RXF 393 Renal 1.5 1.8

TK-10 Renal 1.2 11

SN12C Renal 1.7 2.4

* M. Kondo, M.S. Thesis, UMass Dartmouth, 2006

Ursolic acid dramatically reduced formation of colon tumor colonies in a 14-day soft agar colony formation bioassay

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Liberty, et al, Acta Horticulturae, 2009

Ursolic acid also increased apoptosis in colon tumor cellsLiberty, et al, Acta Horticulturae, 2009

Steroids: numbering system and possible ring conformations

Ginseng (Panax ginseng)Traditional Asian medicinal herbRoots are dried & peeled (white) or steamed (red)Contains triterpenoid saponins (“ginsenosides”) with multiple sugars attached

Taken as a general restorative, “adaptogen” to help body cope with stressTraditional uses: anemia, diabetes, gastrointestinal, insomnia, impotenceEffects on CNS—stimulate or relax? Better concentration? Studies indicate ginseng may modulate blood pressure, enhancecerebral blood flow and increase mitochondrial energy metabolism.

CholesterolMain functions and properties• Cell membrane structure & fluidity• Precursor of steroid hormones and bile acids• Carried by lipoproteins in esterified form• Found in eggs, dairy, meat• Molecule is flat due to the trans-fused configuration of the B, C & D rings• 8 chiral carbons = 256 possible stereoisomers (only one occurs in nature!)

Diet, medicine and cholesterol levels• Most cholesterol in the body does not come directly from food sources but is biosynthesized in response to body’s needs• However, excess dietary cholesterol can inhibit uptake by cells and dietary fats affect cholesterol transport & metabolism• High HDL levels reduce cholesterol level in bloodstream• Many cholesterol-lowering drugs (statins) work by interfering with cholesterol biosynthesis• Statin structure mimics that of HMG Co-A - competitively inhibits HMG Co-A reductase, a key enzyme in IPP biosynthesis

Sterol skeletons: variation is mainly in the side chain on the D ring,additional methylation on other rings

Some common plant and fungal sterols

Medicinal applications of sterols (Dewick, Ch. 5) Dietary plant sterols have been known to reduce cholesterol levels in laboratory animals for many years, which has led to the introduction of plant sterol esters as food additives, particularly in margarines, as an aid to reducing blood levels of low density lipoprotein (LDL) cholesterol (risk factor for heart disease) Plant sterol esters are usually obtained by esterifying sitosterol with fatty acids to produce a fat-soluble product. Regular consumption of this material (1.3 g per day) is shown to reduce blood LDL cholesterol levels by 10–15%. The average diet will normally include small amounts of plant sterol esters. How it works: The plant sterols are more hydrophobic than cholesterol and have a higher affinity for micelles involved in fat digestion, effectively decreasing intestinal cholesterol absorption. Plant sterols themselves are not absorbed from the GI tract. Related materials used in a similar way are plant stanol esters. Stanols are obtained by hydrogenation of plant sterols: mainly sitostanol and campestanol. These are then esterified with fatty acids. Regular consumption of plant stanol esters (3.4 g per day) is shown to reduce blood LDL cholesterol levels by an average of 14%. Stanols are usually transesterified with rapeseed oil, which is rich in unsaturated fatty acids. Foods fortified with plant sterols include spreads (such as Benecol and Take Control), beverages (such as Minute Maid Heart Wise) and snacks (such as Nature Valley Healthy Heart Chewy Granola Bars).

Biosynthetic route to an animal steroid: progesterone

A semi-synthetic route to progesterone used in production of birth control pills

Some key steroids in human physiology

Estrogens

Testosterone and estrogens produced through same pathway

Cholesterol

Natural andsynthetic“anabolic” steroids

Vitamin D• Cholecalciferol is a sterol (isoprenoid) formed photochemically

in animal skin from 7-dehydrocholesterol by sun’s irradiation• Also found in yeasts and some plants as D2 (ergocalciferol)

which only differs in side chain structure • Dietary sources include liver and dairy products, fish liver oils• Vitamin D3 is hydroxylated in the body to its active forms, first

calcidiol and then to calcitriol (Figure 5.106, p58 Dewick).• Key role: Ca metabolism• Calcitriol stimulates the absorption of calcium and phosphate in

the intestine and the mobilization of calcium from bone• Other physiological functions include a role in immune

modulation, hormone secretion, and cell differentiation • Vitamin D deficiency is common in Northern latitudes due to

reduced exposure to sunlight• Rickets = inability to calcify the collagen matrix of growing bone,

causing lack of rigidity in the bones (children) or osteoporosis• In most countries, milk and cereals are fortified with vitamin D3

Vitamin D is a product of the mevalonate pathway, made from a cholesterol derivative.B ring opens up allowing for rotation and isomerization

Studies in human populations

• Epidemiological studies follow a selected population over a defined period of time, monitor intake of the test substance as well as other health factors, habits, lifestyle and assess the effect of these parameters on the number and type of disease-related incidents.

• Clinical trials usually use a randomized, placebo-controlled, double-blind design to administer treatment over a defined period, then assess the difference in specific measurable endpoints between the populations

Recent findings on Vitamin D and health

• Cancer: Vitamin D thought to improve cancer survival – high levels associated with 40% less colon cancer mortality (Dana Farber study)

• Incidence of certain cancers higher in Northern latitudes• CVD: patients age 50+ with Vit. D deficiency were 45-

78% more likely to develop cardiovascular diseases & mortality (Utah study)

• Diabetes: Vitamin D supplementation reduced insulin resistance in a New Zealand study

• Crohn’s disease: Vit.D induced expression of NOD2 gene for defense against inflammatory bacteria

• Bone health: a study of n = 70,000 showed Vit D + Ca supplement reduced fractures by < 16% (BMJ study);Vit.D (600 IU) + Ca (1200 mg) with weight-bearing exercise recommended to prevent osteoporosis.

Vitamin D and immunity• An 2009 study published by researchers at Oregon State University

reveals that Vitamin D is so crucial to the functioning of your immune system that the ability of vitamin D to boost immune function and destroy invading microorganisms has been conserved in the genome for over 60 million years.

• In primates, this action of "turning on" an optimal response to microbial attack only works properly in the presence of adequate vitamin D, which is produced as a result of sun exposure, and is available in smaller amounts from dietary sources.

• Vitamin D prevents the "adaptive" immune response from over-reacting, thereby reducing inflammation, and appears to suppress that immune response. At the same time vitamin D boosts the innate immune response by turning on production of antimicrobial peptides. The overall effect may help to prevent the immune system from overreacting.

• http://www.naturalnews.com/027231_Vitamin_D_immune_system_vaccines.html

• Also check out “The miracle of Vitamin D” at http://www.westonaprice.org/The-Miracle-of-Vitamin-D.html

in vitro study: Vitamin D2 induces apoptosis in human leukemia cells

• Chen, W-J et al, Induction of Apoptosis by Vitamin D2, Ergocalciferol, via Reactive Oxygen Species Generation, Glutathione Depletion, and Caspase Activation in Human Leukemia Cells. J. Agric. Food Chem. 2008, 56, 2996–3005

• Investigated the ability of D2 to induce apoptosis (programmed cell death) in HL-60 leukemia cells and the mechanistic pathways involved

• Found that D2 induced apoptosis primarily through an oxidative pathway involving reactions in the mitochondria (caspase activation and modulation of regulatory proteins)