History of Biochemistry
Noel G Coley, The Open University, Milton Keynes, UK
A history of biochemistry from the eighteenth to the late twentieth century, revealing some
of the problems faced by biochemists seeking chemical explanations for biological
phenomena and introducing some origins of modern biochemistry.
Introduction
Biochemistry aims to explain biological phenomena in
chemical terms. The problems are highly complex and until
the twentieth century progress was slow and unreliable.
During the eighteenth century chemistry was dominated
by the phlogiston theory and the traditional method of
organic analysis by destructive distillation provided no
information about elementary composition. Both in
theory and techniques therefore, chemistry was wholly
inadequate to unravel the mysteries of the vital functions.
Yet the importance of fermentation was recognized,
photosynthesis was discovered and studies of animal
respiration and digestion made useful contributions to
early biochemistry. The complex chemical transforma-
tions occurring at moderate temperatures during the vital
functions led to the belief that in living matter the ordinary
laws of chemistry were modied by unknown vital forces.
These controlled the special conditions pertaining to living
matter, but when life was extinguished ordinary chemical
forces once more came into play, resulting in decomposi-
tion and decay. Despite growing experimental evidence to
the contrary, vitalism persisted among physiologists and
chemists into the nineteenth century.
The displacement of phlogiston by oxygen from the late
eighteenth century greatly facilitated progress. In addition,
Antoine Lavoisiers experimental denition of the chemi-
cal element was accompanied by a more consistent use of
the balance and a rational chemical nomenclature.
Combustion analysis, rst introduced by Joseph Louis
Gay-Lussac in 1811, allowed the empirical formulae of
organic compounds to be determined, although more
accurate atomic weights were required for reliable results.
Chemists analysed plant and animal substances from the
beginning of the nineteenth century, while physiologists
and some physicians applied themselves to studies of the
vital functions in health and disease. However, the
development of biochemistry would also require later
nineteenth-century advances in physical and organic
chemistry. These included structural organic chemistry,
electrochemistry, osmosis, colloid chemistry, stereochem-
ENCYCLOPEDIA OF LIFE SCIENCES 2001, John Wiley & Sons, Ltd. www.els.net
Introductory article
Article Contents
. Introduction
. Animal Chemistry
. Cell Theory
. Biological Oxidation
. Structural Organic Chemistry
. Hormones and Endocrinology
. Vitamins
. Blood Chemistry
. Molecular Biology and the Nucleic Acids
istry and molecular energy studies together with physical
techniques such as X-ray diraction, electrophoresis,
chromatography and labelling with radioisotopes.
Animal Chemistry
In the 1780s Lavoisier proposed a mechanism for photo-
synthesis by which plants take in carbon dioxide and
release oxygen; he also began to investigate animal
respiration. With Pierre Laplace he measured the heat
evolved by small mammals, and similarities between
combustion and respiration led him to locate the produc-
tion of animal heat entirely in the lungs. William Prout,
better known for his atomic hypothesis, also studied the
vital processes. He thought that digestion and assimilation
proceeded in stages involving the action of saliva, gastric
juice, pancreatic juice and bile, followed by oxidation in the
lungs, where the products of digestion were nally
converted into blood. In 1824 Prout discovered hydro-
chloric acid in gastric juice and he later classied foods as
saccharinous, albuminous and oleaginous (cf. carbohy-
drates, proteins and fats).
Michel Euge`ne Chevreul investigated the nature of
animal fats from about 1811. He showed that the fatty
acids were analogous to inorganic acids and were capable
of yielding salts with bases. Separating them using solvents,
he identied each fatty acid by its melting point. The use of
a physical constant as a criterion of purity and means of
identication was an important step in the development of
organic analysis. His results, published in 1823, showed
that animal fats, although products of the vital functions,
were normal chemical compounds.
Jons Jacob Berzelius and Justus Liebig were among the
leading nineteenth-century chemists who advanced bio-
chemistry. Berzelius analysed animal solids and uids;
Liebig endeavoured to extend Berzeliuss work by creating
a comprehensive metabolic theory. Beginning with the
oxygen theory and his own organic analyses, Liebig
1
 History of Biochemistry
described the assimilation of foods and the oxidation of
muscle tissues releasing energy and excretory products like
urea and uric acid. For three decades after its publication in
1842, Liebigs Animal Chemistry, or organic chemistry in its
application to physiology and pathology stimulated chemi-
cal and physiological research. Unfortunately, Liebig was
no physiologist and most of those who tried to nd
supporting evidence for his theories failed. By about 1870
Liebigs theories had been almost entirely superseded, but
the research they engendered helped to advance biochem-
istry and some nineteenth-century chemists even applied
Liebigs ideas successfully. For example, Henry Bence
Jones discovered proteins now known to be part of the
autoimmune system and Ludwig Thudichum identied
some important biochemical substances, including hae-
matoporphyrins in the blood and the main chemical
constituents of the brain.
Cell Theory
In 1838 Theodor Schwann, originator of the cell theory in
biology, suggested that fermentation occurred only in living
yeast cells. Liebig refused to accept this and proposed an
alternative chemical theory of fermentation but Louis
Pasteur, opposing Liebigs chemical theory, showed that
fermentation depends on the vital functions of living yeast
cells and bacteria. The controversy was resolved in 1872
when Pasteurs work received general recognition.
Close links between plant and animal life have been
identied. Animal metabolism is ultimately dependent on
the assimilation of plant material and consequently
photosynthesis studies have made important contributions
to the elucidation of animal metabolism. In 1817 J. Pelletier
and J. B. Caventou suggested the name chlorophyll for
the green substance common to all plants. The connection
between chlorophyll and starch in growing plants was
recognized in1862 when Julius von Sachs, a German plant
physiologist, suggested that starch in green plants is
produced from carbon dioxide and water. In the early
years of the twentieth century chromatography revealed
two forms of chlorophyll and in 1914, Richard Willstatter
and Arthur Stoll showed that both are esters of dibasic
acids with methyl alcohol and phytol, a hitherto unknown
unsaturated, aliphatic alcohol (C20H39OH). Further in-
vestigation of photosynthesis showed that it involved a
chemical and a photochemical step. The chemical step is
associated with the formation of a chlorophyllcarbon
dioxide compound and the photochemical step involves
the formation of a peroxide which is then decomposed by
an enzyme to yield oxygen, formaldehyde and reconsti-
tuted chlorophyll.
About 1860 Moritz Traube suggested that to understand
the chemistry of life required a correct theory of fermenta-
tion. His ferments were oxygen-rich agents derived from
2
proteins. In 1878 Willy Kuhne coined the term enzyme for
Traubes ferments. Enzyme action had rst been observed
in 1833 when Anselme Payen and Jean Francois Persoz
isolated a compound (diastase) which converted starch
into sugar. Three years later pepsin was extracted from the
stomach wall by Schwann. These discoveries preceded
Berzeliuss notion of catalysis (1837) which, he predicted,
would prove important in plants and animals where
complex reactions took place at very moderate tempera-
tures. Further investigation of fermentation resulted in a
distinction between organized ferments such as yeast and
unorganized ferments including the enzymes, the biolo-
gical catalysts, but in 1877 Felix Hoppe-Seyler suggested
that there was no fundamental dierence between them.
Twenty years later Eduard Buchner fermented a sugar
solution with a cell-free extract of yeast. This discovery
showed that the contents of the cells, rather than their life
processes, caused fermentation. This led to a theory of
fermentation based on enzyme action and provided the key
to the study of cell chemistry.
Liebigs idea that animals are incapable of biosynthesis
was challenged in the 1850s by Claude Bernards discovery
of the glycogenic function of the liver. In 1867 Carl Voit
tried to revive Liebigs theory of the direct assimilation of
proteins, but Eduard Puger, Professor of Physiology at
Bonn, argued that there were constitutional dierences
between food proteins and tissue proteins. In the 1870s he
demonstrated the importance of intracellular respiration, a
discovery embraced by Bernard, and suggested a theory of
indirect nutrition, whereby animal cells synthesize complex
substances from simpler nutrient molecules derived from
food. Bernard postulated an internal environment within
the cells, where chemical degradation and synthesis takes
place. According to Bernard all the principles necessary for
the maintenance of animal life are released into the blood.
Respiration introduces oxygen, digestion introduces the
necessary nutrients together with the secretions of the
various organs. The blood is the carrier of all these
substances and the cells absorb from it only what they
require to maintain the vital functions occurring inside
them. The circulation and secretions together ensure,
besides the renewal of the internal environment, the
removal of waste products, and all these changes are
regulated and harmonized by the nervous system, main-
taining a steady state (homeostasis).
Bernards theory of the internal environment marks a
watershed in the history of biochemistry. It put an end to
theories of direct assimilation, replacing them with the
breakdown of complex food molecules into smaller nutrient
constituents from which new compounds specic to the
needs of each cell were synthesized. This theory pregured
one of the most important discoveries of modern molecular
biology the coded programme of protein synthesis but
Bernards ideas of indirect nutrition and the internal
environment were too advanced for many of his contem-
poraries. Much detailed research was needed to establish

them and it was due to British and American scientists
including W. M. Bayliss, E. H. Starling, C. S. Sherrington, J.
S. Haldane, J. Barcroft, W. B. Cannon and L. J. Henderson
that the concept of the internal environment was elaborated
from the beginning of the twentieth century.
Biological Oxidation
It has been recognized from the beginning of the nineteenth
century that energy is released during biological oxidation,
though the mechanisms by which this was brought about
remained in doubt. When C. F. Schonbein discovered
ozone in 1840 he suggested that the rst step in biological
oxidation was the conversion of oxygen to ozone. This idea
led to the popular nineteenth-century ozone craze. Later,
in 1903, A. N. Bach and R. Chodat discovered the enzyme
peroxidase in plant cells and it was thought that peroxides
caused biological oxidation in a two-stage process. In the
rst stage the enzyme oxygenase formed peroxides; in the
second stage peroxidase used these peroxides to oxidize
other organic compounds. This theory dominated ideas on
biological oxidation until about 1920. Although based on
plants, it was widely thought that the oxygenase
peroxidase system also accounted for respiration in animal
tissues. It was later found that oxygenase is itself an enzyme
(catechol oxidase) capable of oxidizing catechol and
similar compounds to quinones and that peroxidase
cannot use these as oxidizing agents.
From 1920 another theory of biological oxidation was
proposed, based on the respiratory systems in yeast,
bacteria and animal tissues. According to this theory,
oxygen atoms were rst activated by combination with
iron atoms in a haemoprotein enzyme. Otto Warburg
called this the respiratory ferment; it would later be
identied as cytochrome oxidase. A rival school considered
that the organic molecules were activated rather than
oxygen and that most biological oxidations were dehy-
drogenations brought about by a new group of enzymes,
the dehydrogenases. These have turned out to be the most
important factors in biological oxidations; there are now
well over 150 known dehydrogenases. By studying these
two opposing systems using spectroscopic techniques
David Keilin followed the course of respiratory changes
in the mitochondria of cells. He discovered the oxidative
cytochromes in 1925 and identied many intermediate
steps in cell respiration.
In 1906 Arthur Harden and W. J. Young observed that
the addition of a soluble phosphate to a fermenting sugar
solution caused the rate of fermentation to increase. The
additional carbon dioxide and alcohol formed was
proportional to the quantity of phosphate added and the
phosphate was converted into a hexosephosphoric acid.
These observations were important for metabolic studies in
the mid-twentieth century, which have shown that most
History of Biochemistry
vital functions depend on enzyme action. The hydrolytic
processes catalysed by enzymes break down starch, fats
and proteins into simpler molecules (monosaccharides,
fatty acids and amino acids) which then enter complex
metabolic pathways also controlled by enzymes. During
these processes energy is released, providing animal heat or
to be stored for later use in muscular exertion. One of the
principal aims of twentieth-century biochemistry has been
to discover the details of these metabolic pathways.
Considerable progress has been made, though many
details still require further elucidation.
In 1907 Walter Fletcher and Frederick Gowland
Hopkins published the rst reliable quantitative data on
the proportions of lactic acid in muscle tissue. Lactic acid
was known to take part in the biochemical transformations
of carbohydrates, linking them with proteins and fats, but
these two workers now recognized that its role in the
metabolism of animal tissues is pivotal. However, they did
not discover that lactic acid holds an intermediate position
between sugar and alcohol in the metabolism of muscle
tissue.
In the early years of the twentieth century Gustav
Embden had isolated several intermediate metabolic
products from muscle tissues, including adenyl phosphoric
acid, also found in the liver. Embden was the rst to
discover and link together all the steps in the conversion of
glycogen to lactic acid. Between 1919 and 1921 Otto
Meyerhof, recognizing that muscle is the only tissue in
which it is possible to compare the chemical changes
occurring with the work done or heat energy evolved,
investigated lactic acid formation in muscle tissue as a
measure of work done. This led to the glycogenlactic acid
cycle, and made a fundamental contribution to the
understanding of muscular action. In all but the briefest,
most intense muscular contractions additional adenosine
triphosphate (ATP), an important energy-carrying coen-
zyme, is supplied by the chemical reactions of the
glycolytic, or EmbdenMeyerhofParnas, pathway, par-
ticularly applicable to rapid muscular action. In this, the
change from glucose to lactate is coupled with the
formation of ATP from adenosine diphosphate (ADP). If
not immediately required, the energy is stored in the
muscle. ATP is created in greater abundance than the
numbers of carbohydrate, fat or protein molecules
metabolized, creating more of this energy-rich compound
than is required for immediate needs. The excess energy is
stored in muscle tissue as phosphocreatine, a labile
compound that readily absorbs and releases phosphate
groups.
Phosphocreatine 1 ADP!creatine 1 ATP
ATP 2 1 phosphate group!ADP 1 energy
Creatine 1 1 phosphate group!phosphocreatine
3
 History of Biochemistry
The lactate, a waste product, diuses out of the muscle to be
transported by the blood to the liver where most of it is
converted into glycogen. The rate of oxidation of lactic acid
is regulated by the rate of respiration. In prolonged strenuous
exercise lactic acid is formed faster than it can diuse out of
the muscle and this results in muscle fatigue. Glycogen from
the liver is reversibly converted into glucose and nds its way
into the bloodstream, thus completing the cycle.
In the 1930s Warburg investigated the relation between
the chemical and photochemical steps in photosynthesis
using the eects of intermittent illumination on the green
unicellular alga Chlorella. His work led to a study of the
cytochromes. Warburg and Keilin independently exam-
ined the respiratory function of the cytochromes and
showed that the respiratory chain in the cells was located in
the mitochondria. Alternate oxidation and reduction
could be traced through chains of cytochromes, each
reducing the next in line until the last, identied as the
respiratory ferment, reacted with oxygen. At each stage the
small quantities of energy released are stored in high-
energy bonds such as the phosphate link in ATP. The
whole complex system was summarized in the Krebs cycle,
the metabolic pathways of which have been subjected to
minute and intensive research ever since. It also became
clear that other substances besides the cytochromes were
involved and in 1932 Axel Theorell working with Warburg,
isolated the rst so-called yellow enzyme composed of a structures of large molecules was needed. Thus, progress in
protein and the nonprotein yellow coenzyme riboavin
(vitamin B2). Keilin isolated the oxidative enzyme cyto-
chrome c and proposed a new explanation of the action of
the cytochromes involving activated hydrogen as well as
oxygen. These investigations were carried out on heart-
muscle preparations, but similar reactions have been
detected in the cells of plants, microorganisms and fungi.
Thus, they represent a common feature shared by all forms
of life. They were essential early stages in the elaboration of
the citric (or tricarboxylic) acid cycle proposed by Sir Hans
Krebs in 1937. In animal cells the enzymes specic to each
step are located in the mitochondria, in plant cells they are
found in the chloroplasts, and in microorganisms they are
found in the cell walls.
A fuller elaboration of the citric acid cycle came after
1940 with the discovery that the three principal food
constituents, carbohydrates, fats and proteins, all yield a
common product, coenzyme A. This forms citric acid which
is then broken down into carbon dioxide and water with the
liberation of coenzyme A once more. At each intermediate
step in this metabolic chain, hydrogen atoms are transferred
from one carrier to another and small quantities of energy
are released. Thus, the citric acid cycle is the source of
energy refurbishment supporting all the vital processes.
Enzyme reaction rates vary widely and depend on
activators, or coenzymes, ranging from simple metal ions
such as Ca2 1 or Mg2 1 to complex organic molecules such
as vitamins. In 1960 it was found that the coenzyme in
energy-producing processes, ATP, is itself fabricated in the
4
cells by a protein enzyme, ATP synthase. The molecular
structure of ATP synthase was partially determined in 1994
after 12 years of research. In addition to providing fresh
knowledge about how living things produce energy, it is
thought that this research may throw new light on the
processes of ageing and the causes of degenerative diseases.
The large molecules of carbohydrates, fats and proteins are
broken down into smaller molecules such as pyruvates, free
fatty acids and amino acids, all of which take part in the
mechanisms of the citric acid cycle and thus link the
metabolic transformations of carbohydrates, proteins and
fats. Enzymes are now known to control most biochemical
processes, from the breakdown of complex molecules like
carbohydrates and proteins during animal metabolism to
the synthesis of macromolecules within the cells. Most
enzymes have a protein structure and most work with one
or more coenzymes, but certain other complex molecules
not regarded as enzymes, such as messenger RNA, also
show catalytic properties.
Structural Organic Chemistry
Before the details of such cellular mechanisms could be
fully explored, far more information about the chemical
biochemistry had to await the development of structural
organic chemistry.
In 1815 J. B. Biot found that certain natural oils rotated
the plane of polarized light. The same substance could exist
in two forms both having the same empirical formula, yet
capable of rotating the plane of polarized light in opposite
directions. The problem facing organic chemists was to
correlate optical activity with molecular conguration.
The fundamental discovery required to solve this problem
was Kekules recognition in 1858 that the carbon atom is
tetravalent. Using this, J. H. Vant Ho suggested in 1874
that structural formulae must be three-dimensional. A
compound in which a central carbon atom is attached to
four dierent substituents in a tetrahedral structure would
have two non-superposable forms one of which is the
mirror image of the other. Applying this concept Pasteur
investigated molecular asymmetry in the tartrates by
separating potassium tartrate crystals mechanically. Op-
tical activity was later used by Emil Fischer to identify the
many stereoisomers of the sugars.
In 1885 Fischer observed that phenylhydrazine
(C6H5.NH.NH2) forms well-dened crystalline compounds
(osazones) with the sugars and this allowed him to identify
the molecular structures of 16 stereoisomers of glucose.
Before 1891 it had been necessary to assume a conguration
for each sugar, but from that year onwards Fischer applied
himself to the problem of assigning specic congurations to
each isomeric sugar molecule and by 1896 he had done so for
all the monosaccharides. He retained the straight-chain

formulae devised by H. Kiliani, though later studies by W. N.
Haworth and others showed that they contain lactone rings.
Fischer also examined the degradation of sugars by enzymes
and from his study of the saccharases he recognized the
specicity of enzymes and concluded that the enzyme and its
substrate were related as a key to its lock. Later work has
shown this concept to be true to a degree Fischer himself
could hardly have suspected.
Fischers other major contribution to structural organic
chemistry concerned his elucidation of the molecular
structures of the purines, amino acids and proteins. Even
before he began work on the sugars Fischer had been
engaged in a study of three purine derivatives, caeine,
theobromine and xanthine, all structurally related to uric
acid. Ludwig Medicus had given the correct formula for
uric acid in 1875; Fischer conrmed it by synthesis.
Between 1882 and 1900 he isolated about 130 purine
derivatives, thus extending Liebig and Friedrich Wohlers
1838 studies of the oxidation products of uric acid. Fischer
attempted to correlate the molecular structures of these
compounds with their physiological properties. Later, in
1914, he returned to this work and succeeded in preparing
the rst synthetic nucleotide, theophylline d-glucose
phosphoric acid, thus linking his work on sugars with his
studies of the purines. This work was a rst step leading to
the study of the nucleo-proteins. Fischer also investigated
polypeptides and simple proteins, showing how amino
acids are combined in protein molecules. By 1907 he had
synthesized a polypeptide containing 18 amino acids. Its
molecular weight was 1213 and he calculated that there
would be 816 possible optical isomers.
In 1877 Traube suggested that enzymes are related to
proteins, but the isolation of pure enzymes only began in the
1920s. Jackbean urease was the rst enzyme to be crystallized
in a pure state, by J. B. Sumner in 1926. J. H. Northrop, who
isolated several proteolytic enzymes, crystallized pepsin in
1930. Both were found to be complex proteins and little
could be done to determine their structures until special
methods were developed. The sequence of amino acids in
proteins was later found using a method devised by
Frederick Sanger for determining the order of the 51 amino
acids in the insulin molecule in about 1950. Another group of
biological compounds, the hormones, has also caused
considerable diculty for structural organic chemists in
the twentieth century. Those hormones, which like glucagon
are polypeptides, have posed problems in the elucidation of
their amino acid sequences, but others have been found to be
based on the characteristic ring structure of the sterols
containing 17 carbon atoms and 28 hydrogen atoms
arranged in four rings.
Hormones and Endocrinology
In the nineteenth century it was recognized that glands
such as the spleen, thymus, thyroid and adrenals secrete
History of Biochemistry
specic substances which are carried by the blood to other
organs in the body. Towards the end of the nineteenth
century a number of discoveries revealed the essential
nature of these secretions for animal health and it became
common to connect the lack or excess of specic endocrine
secretions with certain diseases. Adrenaline (C9H13O3N)
was discovered in 1894 in experiments on the blood
pressure of a dog. A little later vasopressin, a peptide
hormone released from the posterior lobe of the pituitary
gland, was discovered. This is also capable of increasing the
blood pressure. Adrenaline was isolated independently in
1901 by T. B. Aldrich and J. J. Abel in America and shortly
before by J. Takamine in Japan. In 1902 William Bayliss
and Ernest Starling identied the substance secretin,
produced by epithelial cells in the stomach during
digestion. This is released into the bloodstream and in
the pancreas it causes the secretion of pancreatic juice.
Thus, secretin was recognized as a chemical messenger,
produced in one organ and carried by the bloodstream to
the organ it is intended to stimulate. Starling introduced
the term hormone to describe such chemical messengers
in 1905. However, as secretin is a polypeptide of moderate
molecular weight, Bayliss and Starling were unable to
isolate it, much less determine its molecular structure. The
existence of a number of other mammalian hormones was
surmised, but they all proved dicult to isolate and
synthesize and only a few had been characterized. It is now
known that hormones fall into three main classes: steroids,
peptides and proteins, and amino acid derivatives.
Hormone secretion is controlled by the pituitary gland
situated at the base of the brain. There are three main
divisions of this gland, although the whole complex organ
weighs only about one gram in the human adult. During
the 1920s work on the pituitary gland led to the isolation of
six dierent protein hormones, four of which were found to
stimulate other endocrine glands. Since then other
hormones have been isolated, several of which control
the sexual functions; others govern the rate of growth and
the chemical or physiological balance in the animal body.
The chemical structures of all have been determined and
their chemical syntheses achieved.
Adrenaline, a derivative of catechol, was the rst
hormone to be synthesized in 1904; thyroxin, the active
principle of the thyroid gland, rst isolated by E. C.
Kendall in1914, was synthesized by C. R. Harington and
G. Barger in 19261927. Thyroxine, stored in the thyroid
gland as a protein, is hydrolysed by thyrotrophin, another
hormone produced by the pituitary body. Thus one
hormone calls another into action. Thyroxine is mainly
concerned with the consumption of oxygen and thus the
metabolism of all the cells and tissues in the body; it
appears to increase the production of a number of
enzymes. In 1922 Frederick Banting and Charles Best
identied and prepared the pancreatic hormone, insulin,
for therapeutic use in diabetes. J. J. Abel obtained crystals
of insulin in 1926, but Dorothy Hodgkin using X-ray
5
 History of Biochemistry
diraction elucidated the molecular structure of insulin
only in the 1960s. This work complemented Sangers
earlier determination of the complete amino acid sequence
for bovine insulin. Sangers methods opened the way to the
determination of the structures of many other complex
proteins. He later applied radioisotope labelling using 32P
and other techniques to determine the sequencing in
ribonuclease synthesized in the early 1960s. The pancreas
also produces another hormone, glucagon, a polypeptide
containing 29 amino acids in a known sequence.
The chemical structures of the steroid hormones
produced by the adrenal glands and the sex organs are
related to cholesterol. They are all very similar and even
small structural changes in their molecules produce
profound physiological eects. The steroid hormones
secreted by the adrenal cortex control carbohydrate and
mineral metabolism; some are concerned with the forma-
tion of glucose from proteins and enable the body to
withstand stresses such as intense heat or cold, injury and
infection. The hormones of the adrenal cortex control
carbohydrate and mineral metabolism in the body. Similar
compounds, though with more complex structures, are
produced by the thyroid gland. Work on the sex hormones
was carried out from about 1926. In 19291930 Edward
Doisy in America, Guy Marrian in Britain and Adolf
Butenandt in Germany isolated from the urine of pregnant
women two ovarian hormones related to sterol. Butenandt
also isolated androsterone from male urine and in 1934
Leopold Ruzicka in Switzerland obtained this hormone
from cholesterol. These biologically active substances are
metabolites of the ovaries and testes, respectively. The
dierences in their molecular structures and physiological
actions are relatively slight; both are fat soluble.
The anterior pituitary gland inuences the rate of
secretion of the hormones of the adrenal cortex, the
gonads and thyroid gland. Its action is inuenced by nerve
centres in the hypothalamus, the part of the brain
immediately above the pituitary gland. The secretion of
hormones and the functions they control are therefore
coordinated by the nervous system. In some cases there is a
feedback mechanism whereby the secretions of one
endocrine gland stimulates or damps down secretions
from another. Thus a circle of action and reaction exists
between some of the endocrine organs as the balance of
hormone secretion is maintained.
The eects of injecting adrenaline are similar to those
induced by stimulation of the sympathetic nervous system
and this led to the suggestion that the liberation of
adrenaline at sympathetic nerve endings might transmit
the excitatory or inhibitory impulse to the eector cells of
muscles or glands. Thus it seemed that certain hormones
might control the functions of the central nervous system.
It is now thought that a large number of chemicals can act
in this way, but only a few have been so far identied. The
rst of these, noradrenaline, was isolated by Hans van
Euler in 1946. Others include acetylcholine, dopamine and
6
secretonin. Acetylcholine was rst isolated in 1914; its
function in slowing the heartbeat was identied in 1921 by
Otto Loewi, a German physiologist.
In the 1930s it was observed that one hormone can act in
opposition to another. Frederic Houssay in Argentina
found that extracts of the anterior pituitary gland combat
the action of insulin. Frank Young found that adminis-
tration of anterior pituitary extract causes a persistent
diabetic condition. Cyril Long and Francis Lukens
discovered an antagonism between insulin and the secre-
tions of the adrenal cortex. These and other observations
that hormones can act against each other have proved
useful in elucidating the endocrine control of the vital
functions and in medical and surgical treatments of certain
conditions. However, it is now known that the substance
present in an endocrine gland does not necessarily take the
same form as that of the hormone in the blood.
Furthermore, hormones may themselves undergo meta-
bolic changes, either in the blood or in the tissues, before
they are able to cause reactions in the cells or in the enzymes
they inuence. Thus, increased knowledge in endocrinol-
ogy has created a situation in which the precise chemical
denition of a hormone has become a matter of some
diculty.
Vitamins
Although dietary deciencies have long been recognized as
the cause of certain diseases, the search for vitamins (a term
introduced by Casimir Funk) began about 1912. Gowland
Hopkins observed that animals fed on a sucient quantity
of puried foods ceased to grow unless a small amount of
milk was added. This drew attention to the vitamin
question, though for several years the real existence of
these elusive dietary factors was disputed.
In 1915 Elmer McCollum and M. Davis identied fat-
soluble A and water-soluble B as essential accessory
dietary factors in rats. These were later named vitamins
A and B respectively. The latter was found to prevent beri-
beri, while lack of vitamin A retarded growth and caused
increased liability to infection of the respiratory system.
The antiscorbutic vitamin C was later identied and in
1922 lack of fat-soluble vitamin D was recognized as the
cause of rickets. About the same time vitamin E was
identied. In 1926 it was discovered that pellagra is a
vitamin deciency disease. The vitamin in this case seemed
to accompany the anti beri-beri factor, but was dierent
from it, in that it was much more stable to heat. In 1927 the
anti beri-beri factor was labelled vitamin B1 and the heat-
stable factor became vitamin B2. This was at rst thought
to be a single compound, but was later found to be a
complex including riboavin, a yellow pigment with
growth-promoting properties found in milk. In 1934
pyridoxine, another component of the B2 complex was

identied. Nicotinic acid and nicotinamide (niacin and
niacin amide), the anti-pellagra factor, were identied in
1937. For several decades additional vitamins were
discovered. Three are of particular importance for human
blood: vitamin K promotes the formation of prothrombin
in the liver, folic acid prevents anaemia and vitamin B12
(cyanocobalamin) is the anti-pernicious anaemia factor.
Each of the four fat-soluble vitamin groups, A, D, E and
K, includes several related compounds with biological
activity. All contain one or more units of an isoprene
structure in their molecules (C 5 CHC.(CH3) 5 CH).
These vitamins are transported by lymph from the
intestines to the blood. Bile salts are required for their
ecient absorption. They may be taken up as esters of
palmitic acid or combined with a protein. Vitamins A, D
and K are stored chiey in the liver; vitamin E is found in
body fat. The action of the fat-soluble vitamins is
connected with certain enzymes. Some carotenes also
show vitamin A activity. Only a and b carotenes and
tryptoxanthin are important in human metabolism and b
carotene is the most active. Water-soluble vitamins include
vitamin B1 (thiamin), B2 (riboavin), B3, B6 (pyridoxine),
niacin, vitamin B12, folic acid, pantothenic acid and biotin.
In metabolic processes they act as coenzymes. Thus,
vitamins B1, B2 and B6 become phosphates, biotin under-
goes a change in structure and nicotinic, pantothenic and
folic acids form esters. Water-soluble vitamins are not
stored in the body to the same extent as fat-soluble ones
and any excess is excreted in the urine. In addition to the
true vitamins there are other substances with vitamin
activity such as choline and p-amino benzoic acid, but these
are fabricated in the body, as well as occurring in foods,
and are not considered true vitamins.
Blood Chemistry
The dierence in colour between venous and arterial blood
was known in ancient times, but that the same blood
changed colour as it circulated from veins to arteries
through the lungs was discovered by William Harvey in the
1620s. The cellular structure of the blood was also rst
observed in the seventeenth century. The red blood cells
attracted most attention and in the nineteenth century it
was realized that they were the oxygen carriers. In 1827
Hans Fischer and his co-workers synthesized a large
number of porphyrins, the chemical components of the red
cells, and showed how they transport oxygen in the blood.
As the only uid circulating through all the organs, the
blood was thought to transport nutrients and remove
waste products, maintaining homeostasis in the body,
fundamental to Bernards theory of the internal environ-
ment. In 1862 Hoppe-Seyler observed the characteristic
absorption spectrum of oxyhaemoglobin and in the same
year William Stokes demonstrated the oxidationreduc-
History of Biochemistry
tion of the pigment present in the red cells. The nature of
the active part of haemoglobin and the structure of the
porphyrin ring was investigated by Ernst Kuster in 1913.
Max Perutz and John Kendrew determined the complete
structure of the haemoglobin molecule using X-ray
spectrographic techniques between 1937 and 1959.
The white blood cells, or leucocytes, include a propor-
tion of phagocytes, cells that engulf and digest bacteria,
protecting the body from disease. In addition, 2025% of
the white cells, the lymphocytes, combine with antigens
and remove them from the body, so controlling infections.
There are two types of lymphocytes, called B cells and T
cells. The B cells produce chemical antibodies on activation
by antigens and release them into the bloodstream. In the
1970s the Japanese immunologist Susumu Tonegawa
showed that about 1000 pieces of genetic material in the
antibody-producing part of B lymphocytes can be shued
and recombined in dierent sequences, enabling up to 1
billion dierent types of antibodies to be formed each
specic to a dierent antigen.
Serum albumin accounts for 55% of the total protein in
blood plasma. Its main function is to help maintain the
osmotic pressure between blood vessels and tissues.
Circulating blood tends to force blood out of the blood
vessels and into the tissues, but the colloidal nature of
albumin and to a lesser extent of other blood proteins, the
globulins, keeps the blood within the blood vessels.
Albumin also contains two materials necessary for the
control of clotting: antithrombin keeps the clotting enzyme
thrombin from working unless needed and heparin
cofactor is necessary for the anticlotting action of heparin.
The fate of cholesterol, another chemical substance found
in the bloodstream, was investigated by Henri V. Goldstein
in 1972. He found that low-density lipoproteins, the
primary cholesterol-carrying particles, are withdrawn
from the bloodstream into the bodys cells by receptors
on the cells surface. Although dealing with complex
chemical substances and processes, studies on the blood
tend to be physiological and immunological, rather than
biochemical. They form part of medical research and it is in
such areas that the overlap between biochemistry, medical
chemistry and physiology highlights the diculties of
demarcation and the precise denition of biochemistry as
an independent discipline.
Molecular Biology and the Nucleic Acids
Since the 1950s progress in biochemistry has been led by
molecular biology, a subject concerned with the ultimate
physiological organization of living matter at the mole-
cular level. It is virtually impossible to distinguish between
biochemistry and molecular biology, since both are
concerned with intermolecular transformations within
living cells. Since its inception in the late nineteenth
7
 History of Biochemistry
century biochemistry has sought to develop a molecular
biology in stark contrast to the macrobiology of organs
and tissues. Yet modern molecular biology seeks recogni-
tion as a separate discipline concerned with the molecular
basis of inheritance (genetics) and with protein synthesis in
the cells. It has developed from and is related to
biochemistry, yet distinct from it. In 1950 W. T. Astbury
identied molecular biology with a study of the forms of
biological molecules, their development through higher
levels of organization and their relationship with genesis
and function. The rst part of this denition is conforma-
tional, the second is informational and for some years these
two aspects of molecular biology were studied indepen-
dently.
By the end of the nineteenth century biochemists had
come to recognize that complex biological molecules held
the key to understanding vital processes in living cells. But,
as the methods of classical chemistry were inadequate to
determine the structures of large molecules such as
proteins, polysaccharides or nucleic acids, biochemists
concentrated on the transformations of smaller molecules
forming the components of these complex molecules.
Molecular biologists, on the other hand, adopted physico-
chemical techniques such as X-ray diraction, developed
in the 1920s at the Royal Institution in London by W. H.
Bragg who built up a school of crystallography including
Astbury, J. D. Bernal and Kathleen Lonsdale. Using this
technique it was discovered that the structures of complex
molecules could be determined, but even this was simpler
than the problems of explaining their functions in terms of
their structures. Astbury applied X-ray crystallographic
techniques to bres and discovered that when a natural
bre such as hair is stretched, its diraction pattern
changes due to molecular rearrangements in its structure.
Bernal turned to other living materials and with Dorothy
Hodgkin and Perutz obtained diraction patterns for large
crystalline protein molecules. Perutz analysed the structure
of the haemoglobin molecule and Kendrew made the rst
three-dimensional analysis of the molecular structure of
myoglobin. ATP synthase, an enzyme responsible for
synthesizing ATP, the universal energy carrier in living
cells, was also examined. In the 1930s, however, further
progress was impossible due to the inadequacy of
contemporary methods of analysis.
In America it was recognized that the helix was a
common structural form for large molecules. As X-ray
diraction analysis was unable to cope with such a
structure Linus Pauling suggested that model building
would oer the only hope of solving these dicult
problems. For success, however, precise inter-atomic
distances and angles were required. Pauling therefore
applied X-ray diraction techniques to the analysis of
amino acids and small peptides in order to determine their
dimensions with minute accuracy. By the late 1940s he was
able to use his results to construct models of large protein
and polypeptide molecules by combining these smaller
8
units. From this work a general theory of the diraction of
X-rays by helical structures was evolved. This would be
very important for determining the structures of the nucleic
acids. During the same period another group of workers
settled on the study of bacteria, believing that the
bacteriophage was an ideal subject for the study of heredity
because the transfer of hereditary material was not
confused by other biological functions such as metabolism.
The so-called phage group was indierent, even hostile, to
chemistry and perhaps due to this attitude it was only in
1944 that it was realized that the materials under study
were actually nucleic acids.
The British and American researchers were proceeding
independently of each other until James D. Watson, a post-
doctoral student from the phage group, came to work in
Cambridge. Watson was working on DNA (deoxyribonu-
cleic acid), a compound rst isolated from pus cells in 1869.
Its signicance as genetic material was recognized in 1944
when it was observed that bacterial DNA changed the
genetic material of other cells. In 1953 Francis Crick and
Watson proposed the double helix structure for DNA,
providing the conceptual framework for understanding
DNA replication and protein synthesis. It appeared that
the DNA molecule was composed of two helical phos-
phate-sugar chains running in opposite directions and
crosslinked at regular intervals by four organic bases
always appearing in pairs (thymineadenine and cytosine
guanine). The links between the chains are formed by
relatively weak hydrogen bonds and the separation of the
chains leaves each one as a template for duplication using
the small molecules brought to the cells by the blood.
In 1955 S. Ochoa in Spain discovered an enzyme,
polynucleotide phosphorylase, capable of synthesizing
ribonucleic acid (RNA). It was later found that this
enzyme degrades RNA in the cells, but under test-tube
conditions it runs its natural reaction in reverse. The
enzyme has enabled understanding of processes whereby
hereditary information in genes is translated through RNA
intermediaries into enzymes that determine the functions
and character of each cell. With discoveries such as these
the informational molecular biologists could proceed
without further help from the chemical conformationists
who were free to move on to other structural problems.
Although the way in which molecular biology appar-
ently developed seems to t this simple scenario, it is
incomplete. The development of molecular biology in-
cludes many more aspects of biochemical research like
plant viruses such as Tobacco mosaic virus. These can be
readily extracted from plants and crystallized; they all
contain RNA. The mechanisms of nuclear division have
also been important, as has the role of RNA in protein
synthesis. The contributions of these and other lines of
biochemical research cannot be ignored in considering the
origins of molecular biology. In 1978 the New York
Academy of Arts and Science held a meeting to promote a
broader view focusing attention on the history of protein

research. Since then developments in molecular biology
have involved such important advances as the rst
complete synthesis of a protein, the detailed mapping of
the arrangement of atoms in certain enzymes, the elucida-
tion of intricate mechanisms of metabolic regulation and
the molecular action of hormones. These lines of investiga-
tion are closely related to medical research and prominent
recent researchers in molecular biology are largely drawn
from this eld. They include immunologists Caesar Mill-
stein and Georges Kohler, geneticists Christiane Nusslein-
Volhard and Eric Wieshaus, and Hugh Esmor Huxley,
who proposed the sliding lament theory of muscle
contraction.
Further Reading
Coley NG (1973) From Animal Chemistry to Biochemistry. Amersham,
UK: Hulton.
History of Biochemistry
Florkin M and Stotz EH (19721979) Comprehensive Biochemistry, sect.
VI, vols 3033. Amsterdam: Elsevier.
Fruton JS (1990) Contrasts in Scientic Style: Research Groups in the
Chemical and Biochemical Sciences. Philadelphia: American Philoso-
phical Society.
Holmes FL (1974) Claude Bernard and Animal Chemistry. Cambridge
MA: Harvard University Press.
Needham DM (1971) Machina Carnis; The Biochemistry of Muscular
Contraction in its Historical Development. Cambridge: Cambridge
University Press.
Needham J (ed.) (1970) The Chemistry of Life. Cambridge: Cambridge
University Press.
Neuberger A and Deenen LM van (eds) (19831997) Comprehensive
Biochemistry, vols 3538, 40. Amsterdam: Elsevier.
Olby R (1994) The Path to the Double Helix: the Discovery of DNA
(Reprint of 1974 edn with additions). New York: Dover.
Teich M and Needham DM (1992) A Documentary History of
Biochemistry 17701940. Leicester, UK: Leicester University Press.
Weatherall M and Kamminga H (1992) Dynamic Science: Biochemistry
in Cambridge, 18981949. Cambridge: Wellcome Unit for the History
of Medicine.