Mikhail Tsvet: A Life in Five Chapters

Chapter 1 · Qualifying Twice

An Italian birth, a Swiss degree, and a Russian refusal

1872 – 1901 · Asti · Lausanne · Geneva · St Petersburg · Warsaw

Mikhail Semyonovich Tsvet was born at Asti in Italy in May 1872, to a Russian father — an official — and an Italian mother, Maria de Dorozza, who died when he was an infant.

He was brought up and educated in Switzerland, at Lausanne and then Geneva, and took his doctorate at Geneva in 1896 on plant cell physiology, specifically on the structure of chloroplasts.

In 1896 he moved to Russia, and found that his Geneva doctorate was not recognised. To hold any academic position he had to qualify again from the beginning, and he did — taking a Russian master's degree at Kazan in 1901 with a dissertation on plant pigments, and a Russian doctorate later.

So he spent his late twenties redoing work he had already done, in a country whose language and academic culture he had not grown up in.

He worked at Warsaw — then part of the Russian Empire — at the university and the veterinary institute, in modest positions.

His problem was the green colouring matter of leaves. Chlorophyll had been extracted since the 1810s, and there was a persistent question: is it *one* substance or *several*?

The difficulty was that the available methods destroyed what they were meant to examine. Separating pigments meant treating them chemically — with acids, alkalis or reagents — and chlorophyll is fragile. So if you got two substances out, you could never be sure whether they had been in the leaf or had been *made* by your treatment.

That is the problem the method had to solve.

“The substances separate in the column as the layers of a rock are ranged one above another.”

— Mikhail Tsvet, on the appearance of a chromatographic column (1906)

Why this matters

The existing methods of separating pigments destroyed them chemically, so nobody could tell whether the products had been in the leaf or made by the procedure.

You have the second doctorate and the fragile green. What would you ask?

Ask Tsvet

• “What was it like having to qualify all over again?”
• “Why could nobody say whether chlorophyll was one substance?”
• “What is wrong with separating pigments chemically?”
• “Did being an outsider in Russian science cost you?”
• “Why work on leaves at all?”

Chapter 2 · A Tube Packed With Chalk

1903, and the bands that appeared

1903 – 1906 · Warsaw

Tsvet's answer was to separate the pigments *physically* and *gently*, with no chemical treatment at all.

The apparatus is almost absurdly simple. Take a narrow vertical glass tube. Pack it with a powder — he tried many, and powdered *calcium carbonate*, ordinary chalk, worked best. Pour a leaf extract, dissolved in petroleum spirit, onto the top. Then wash it through with more solvent.

And watch.

Because the powder holds some substances more tightly than others, the components travel down the column at *different speeds*. They draw apart. What went in as a single dark green solution emerges as a set of *distinct coloured zones*, ranged one above another down the length of the tube.

The column can then be pushed out of the tube as a cylinder, and the bands cut apart with a knife and each extracted separately.

He described it to the Warsaw Society of Natural Scientists in 1903 and published it fully in 1906.

He named it *chromatography* — writing in colour.

What it showed was that the green of a leaf is *not one substance*. There are two chlorophylls, which he distinguished as chlorophyll a and b, and yellow pigments — the xanthophylls and carotenes — present alongside them and masked by the green.

And — this is the claim that mattered — because nothing chemical had been done to them, they had been in the leaf all along.

The underlying principle is a *partition*: each substance divides itself between the stationary powder and the moving solvent, and how far it travels depends on how it apportions itself. Every form of chromatography since works this way.

Why this matters

Chromatography separates by how each substance apportions itself between a moving solvent and a stationary phase, and it does so without chemically altering anything.

You have the chalk, the solvent and the bands. What is your question?

Ask Tsvet

• “Why do different pigments travel at different speeds?”
• “Why chalk rather than some other powder?”
• “What did you actually see in the tube?”
• “Why does gentleness matter so much here?”
• “Why call it writing in colour?”

Chapter 3 · Willstätter Says the Apparatus Made Them

An objection that could not be answered

1906 – 1919 · Warsaw · Zurich · Berlin

*Richard Willstätter* was the leading authority on chlorophyll in the world. He determined its structure, showed it contains magnesium, and received the Nobel Prize in Chemistry in 1915 for the work.

He did not accept Tsvet's method, and his objection was the sharpest one available: that *adsorption on the powder altered the pigments*. Chlorophyll is known to be unstable and sensitive; calcium carbonate is a reactive surface; on this view the bands were not pre-existing components separated but products *created* by contact with the adsorbent.

In other words: the apparatus makes what it appears to find.

That is not a foolish objection. It is exactly the right question to ask of any new separation, and it is still the first question asked of one now.

Tsvet's answers were reasonable and insufficient. He pointed out that the same bands appeared with different adsorbents, that the recovered pigments retained their properties, and that running the column again gave consistent results. He invited objectors to watch.

What he could not do was *prove* it — because proving that a separated substance is unchanged requires an independent characterisation of it, and he had no such method. He had a coloured band and its position, which is not by itself proof that the band contains a single pure substance.

Willstätter's authority settled the matter practically. The method was set aside for about thirty years.

Tsvet was displaced by the First World War — the Warsaw institutions were evacuated eastward — moved to Moscow and then Voronezh, and lost equipment and position. He fell ill and died at Voronezh in June 1919, aged forty-six.

He never saw the method accepted.

Why this matters

Willstätter's objection — that the apparatus creates what it appears to find — is the right question to ask of any separation, and Tsvet had no way to answer it.

You have the objection and no means of answering it. What would you ask?

Ask Tsvet

• “What exactly was Willstätter's objection?”
• “Why could you not prove the pigments were unaltered?”
• “Is a coloured band evidence of a pure substance?”
• “How much of this was authority rather than argument?”
• “What did the war do to your work?”

Chapter 4 · Revived Thirty Years Later

Kuhn, Karrer, and a number that answered the objection

1931 – 1952 · Heidelberg · Zurich · Leeds

In the early 1930s *Richard Kuhn* in Heidelberg and *Paul Karrer* in Zurich, both working on carotenoids and vitamins, rediscovered Tsvet's papers and applied the method — and it worked exactly as described.

By then the objection had lost most of its force, because chemists had independent means of characterising what came off a column: melting points, absorption spectra, and crystallisation. You could show that the substance in a band was the same substance whether you obtained it by chromatography or by another route.

Both Kuhn and Karrer received Nobel Prizes, in 1938 and 1937, for work that depended on Tsvet's method.

The decisive extension came in 1944, when *Archer Martin*, *Richard Synge*, *Raphael Consden* and *A. H. Gordon* ran mixtures on filter *paper* rather than in a column. Paper chromatography is cheap, needs almost no apparatus, and can be done with tiny quantities.

And it produced the thing Tsvet had lacked: a *number*.

Measure how far a component has travelled from the origin, and how far the solvent front has travelled. The ratio is the *Rf value*, and for a given substance under given conditions it is *characteristic and reproducible*.

That converts a coloured band into an identification. Two laboratories can compare Rf values. A component can be matched against a known standard.

Martin and Synge received the Nobel Prize in 1952. Their citation refers to partition chromatography.

Tsvet had been dead for twenty-five years when the paper method appeared and thirty-three when the prize was given.

Why this matters

The Rf value turned a coloured band into a reproducible number, which is what finally made chromatography an identification method rather than a display.

You have the revival and the number you never had. What is your question?

Ask Tsvet

• “What is an Rf value and why does it matter?”
• “Why did the objection lose its force by the 1930s?”
• “What does paper give you that a column does not?”
• “Should Kuhn and Karrer have credited you more?”
• “Would a number have persuaded Willstätter?”

Chapter 5 · The Commonest Separation in Chemistry

What a tube of chalk turned into

1952 – present · Everywhere

Chromatography in one form or another is now the commonest separation technique in chemistry, and it is difficult to overstate how much depends on it.

*Gas chromatography*, in which the moving phase is a gas and the stationary phase coats the inside of a long fine column, separates volatile mixtures in minutes and is used in forensic laboratories, in doping control, in environmental monitoring and in flavour chemistry.

*High-performance liquid chromatography* — HPLC — forces solvent through a packed column under high pressure and is the workhorse of pharmaceutical analysis; every batch of every drug manufactured is checked by it.

*Thin-layer chromatography* is a plate coated with adsorbent, and does in ten minutes what a column does in hours.

Sanger used chromatography to sequence insulin. The Human Genome Project depended on chromatographic separations. Every drug purity test, every blood test for a metabolite, every check on a food additive uses a descendant of the method.

And the school version — a spot of ink on filter paper, a beaker of solvent, watching the colours separate and climb — is a direct descendant of a tube of chalk in Warsaw in 1903.

The charge Tsvet could not answer in his lifetime has not gone away, and that is worth teaching. *Does the apparatus make what it appears to find?* is the first question asked of any new separation or detection method, and it is a good question. Modern methods answer it with independent verification: run a known standard, use two different techniques, check the result by an unrelated route.

Tsvet had the right method and no way to prove it, and died with the objection standing.

Why this matters

The question Tsvet could not answer — does the apparatus create what it finds? — is still the first one asked of any new separation technique.

You have the tube of chalk and everything it became. What would you ask him?

Ask Tsvet

• “What would you make of a modern HPLC machine?”
• “How does the school ink experiment relate to your column?”
• “How should a new method answer the artefact charge?”
• “What would another twenty years have let you do?”
• “Was Willstätter ever going to be persuaded?”

What Tsvet changed

Chromatography in one form or another is now the commonest separation in chemistry, and the paper chromatogram of a school laboratory is the direct descendant of Tsvet's column. Gas chromatography, HPLC and thin-layer chromatography all rest on the same partition principle. The charge he could not answer in his lifetime — that the apparatus makes what it appears to find — is still the first question asked of any new separation.

A debate that continues

Richard Willstätter, the leading authority on chlorophyll, held that adsorption altered the pigments rather than separating them, and Tsvet had no independent means of proving otherwise; the method was set aside for about thirty years and revived only after his death.

Keep exploring — ask Tsvet

• “How many pigments are actually in a leaf?”
• “What would you have needed to convince the field?”
• “Which separation problem would you take on now?”

Related lives

• Carl Remigius Fresenius — Who Put the Tests in Order
• Emil Fischer — Who Said A Lock Must Fit Its Key
• Rosalind Franklin — The Dark Lady of DNA
• Jan Ingenhousz — The One Who Found the Light

Related themes

Chromatography and separation · Pigments and photosynthesis · Purity and identification