This document summarizes information about cardiac glycosides, flavonoids, and coumarins. It describes the historical and modern uses of cardiac glycosides from plants to treat heart conditions. It provides details on the sources, active compounds, extraction, and assay of key cardiac glycosides like digoxin. It also gives examples of flavonoids like rutin and hesperidin, and notes their isolation and properties. Finally, it introduces coumarins and their plant sources and mentions their anticoagulant activity.

1. Cardiac glycosides
Flavonoids
Anna Drew
with grateful acknowledgement for inspirational teaching received at
The School of Pharmacy, University of London

2. Cardiac glycosides
• Plant glycosides with specific action on heart
• Historical use:
– to assasinate people, arrow poisons
• Historical sources:
– South American toad skins, African plant extracts
• Modern use:
– to treat congestive heart failure (dropsy)
– aglycone structure important for activity

3. Sources
• Scrophulariaceae
• Digitalis purpurea leaves (foxglove)
• Digitalis lanata leaves – white flowers
• Apocyanaceae
• Strophanthus vine seeds – Africa
• Liliceae
• Urginea bulbs (squill) – Europe, India
• Convallaria leaves (lily of the valley)
– also produces a volatile oil perfume

4. Active compounds
• steroid nucleus
• AB cis-junction
• CD cis-junction
– not planar
• C14 = 3y -OH
• C3 = 2y –OH
– sugars attached
• C17 = lactone ring
– classified into 2 groups
Cardenolides
more common
opens in alkali
Bufanolides

5. • sugars:
– 1-4 β-linked at C3 in various combinations
– glucose, rhamnose, deoxy-sugars
• eg digitoxose, digitalose
Strophanidiol
Strophanthus
Scillarenin
(squill)

6. Extraction
• large molecular weight molecules with sugars -> polar
– soluble in water and alcohol
• expensive – long process, solvents

7. Digitalis
• Scrophulariaceae family
– foxglove - biennial flowering plants
• cases of poisoning rare
• natural emetic if eaten in excess
– Digitalis purpurea leaf – purple, British
• -> Digitalis Tablets B.P.
• -> Tincture of Digitalis B.P.
• commercially grown Holland, E. Europe
• NB no extraction for these products
– Digitalis lanata leaf – white,
Mediterranean
• used for manufacture of pure glycosides
• ie digoxin, lanatoside C
• commericially produced Holland, Equador,
USA

8. Chemistry of D.lanata
• compounds belong to
cardenolide series
– 5 membered lactone ring
– approx 96 compounds
• [1930-1950 Stroll
worked on structures]
R1 R2 Names 1y 2y
H H digitoxigenin A A digitoxin
H OH gitoxigenin B B gitoxin
OH H digoxigenin C C digoxin
OH OH diginatigenin D D diginatin
H formylester gitaloxigenin E E gitaloxin
*
* Acetyl group
confers crystalline
properties - makes
compounds more
easily isolated

9. [i] Digitoxose
• sugar found on primary glycosides of D.lanata
• glucose on the end of a chain of O-linked digitoxose sugars at C3
• during harvesting and drying enzymes can remove acetyl groups and the
end glucose
– hence drying method needs to be followed or glycosides degrade further
– after collection dried as rapidly as possible at 60oC, stored in airtight containers
protected from light (contain no more than 6% moisture)
• expect about 10 compounds from D.lanata
– important ones:
• Digoxin “Lanoxin” – Wellcome – 0.25 μg white tablet
• Digitoxin “Digitalin” 0.25 μg small pink tablet
• Lanatoside “Cedilanid” 0.10 μg – less well absorbed but used for rapid digitalisation
• Others not marketed, used experimentally

10. Some cardioactive glycosides from D.lanata:
Ref: Trease & Evans

11. Chemistry of D.purpurea
• Steroid cardenolides
– contains 30 glycosides, 6 main ones
– only has 3 aglycones
• Purpurea 1y glycosides
– do NOT have acetylated digitoxose third sugar
• but these are found in smaller quantities
– called ABE series
Aglycones 1y 2y
digitoxigenin A digitoxin
gitoxigenin B gitoxin
gitaloxigenin E gitaloxin

12. [ii] Digitalose
• found in both species
• only strospeside important as emergency injection for heart attacks –
quickest acting cardiac glycoside

13. Assay of Digitalis B.P.
• required to contain not less than 0.3% total
cardenolides calculated as digitoxin
• important to guarantee reproducibility of
products (drug dosage)
• narrow therapeutic index
• can cause cardiac arrest
• slowly excreted, bound to serum proteins
• long term therapy for patients
• patients tend to be older, weak

14. • Digitalis B.P. tablets
– crushed dried leaves -> green tablet
– contain 30 glycosides each with different
onset, action and excretion profiles
– in different amounts
• influenced by growing conditions
• (temp, water, sun, drying process)
– assay for each glycoside as accurately as
possible – dilute effects by adding grass
• Two ways:

15. [1] Biological assay
• British method - inaccurate but safer
• Tincture of extract of leaves or tablets
• diluted with saline so alcohol <6%v/v
• guinea pigs (6 test, 6 control) x 3 =36
– expensive but can average results
• measure volume injected into vein of leg/foot before
heart stops beating
• monitor heart rate via ECG
– or open chest wall and watch inserted needle with flag on move
– Better to watch ECG – have to differentiate from death from too
large an injected volume
• trained staff required, can calculate potency
• assay acceptable within 80-120% error margin (not that
accurate)

16. • Disadvantages
– inaccurate, expensive
– injecting material IV (avoiding absorption,
excretion)
– end point is death
– toxicity test not therapeutic assessment
• Advantages
– assessing some biological activity
– safety mechanism

17. [2] Chemical assay
• Problem: 30 different glycosides – can measure them
accurately but may not correlate with therapeutic activity
of drug
• Make a tincture (with alcohol)
• decolourise with lead subacetate
• extract glycosides by partition with CHCl3
• evaporate to give residue (containing cardiac glycosides)
• hydrolyse with HCl to remove sugars leaving aglycone
– residue contains gitoxigenin and digitoxigenin (AB series)
– gitaloxigenin -> gitoxigenin when acid hydrolysed

18. • Colourimetric assay to separately determine material amounts
(i) total aglycone
• purple colour with dinitrobenzoic acid and alkali
(ii) digitoxigenin only
• green colour with FeCl3 + acetic acid
– can substract answers to work out
• Digitoxigenin (A series) content
• Gitoxigenin (B series) content
• Advantages:
– precise method (reproducible 2%, standard error 5%)
– unqualified staff, quicker
• Disadvantages:
– doesn’t correlate with biological activity
– only estimating approx 60% therapeutic material
• BOTH methods used in industry

19. Flavonoids
• mainly O-linked glycosides
• occur in plants, lichens, moss
• those in free state and glycosides largest
naturally occurring group of phenols
• aromatic, based on γ-pyrone moiety
• can get several forms of flavonoids depending
on nuclei
flavone isoflavone flavonol

20. • often yellow (flavus Latin – yellow)
• known for a long time
• interest in them for
– anti-inflammatory (and analgesic) properties
– anti-allergic effects
– antithrombotic, vasoprotective properties
• decrease capillary fragility
• phlebitis – changes in vessel walls in extremities
-> plasma leakage -> oedema
– mainly due to high oestrogen, sometimes in males
– tumour inhibition promotion
– protective for gastric mucosa

21. • sugars:
– glucose, rhamnose, arabinose, xylose
– 2-3 attached to phenolic groups in middle of
structures

22. Examples
(a) Rutin (Vitamin P)
– from Fagopyrum esculentum
(buckwheat)
= rhamnoglucoside of quercetin
(b) Hesperidin (‘citrin’)
– from citrus industry
= hesperetin (methyl eriodictyol),
rhamnose, glucose

23. Isolation
• easy
• water and alcohol soluble
• give brightly coloured
solutions
• crystallise easily
• may give colour reactions
eg
– MgCl2 -> violet -> orange
– alkali KOH -> orange
• easy to detect

24. Coumarins
– aromatics based on α-pyrone
– widely distributed in plants
• Leguminosae
• Rubiaceae
• Umbelliferae
• Solanaceae
– first medicinal compounds from
clover
• certain types toxic to animals in
summer
• anticoagulant activity found
• dicoumarols produced clinically