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Phlogiston, bloodletting, and the four humors

Phlogiston – You know, the stuff that’s in stuff. The burny stuff that’s released by fire?Screen Shot 2016-06-09 at 5.20.34 PM.png

Not familiar? Well, that’s because it’s isn’t a thing at all – anymore.

Screen Shot 2016-06-14 at 10.04.59 PMGeorg Ernst Stahl (1659–1734) lived in a complicated time for science. It was just being brought out of the dark ages in many ways and much of what he studied sounds completely foreign and backward to modern ears.

Primarily, Stahl studied the distinction between living and dead material. This vital force was supposedly the anima, or spirit, of a living thing, that gives it ‘agency.’ This was the same force, known as vitalism, that even Louis Pasteur believed was necessary for enzymatic reactions to proceed. Pasteur wasn’t wrong about much, but this one time he fell victim to the prevailing zeitgeist.

Stahl also proposed, in his De motu tonico vitali, that there was a ‘tonic motion’ in things that needed to be permitted for proper circulation of blood. When inflammation or other obstructions occurred, the problem was that this tonic motion was being blocked. One cure for these obstructions was the practice of bloodletting, which addressed the most easily managed of the four humors and was used to treat just about everything.

Although this may sound like a criticism of  Stahl, he was highly regarded as a professor and physician in his time and his work was critical in that it added an experimental element to scientific work. As a testimony to his reputation, he served as physician to both Duke Johann Ernst of Sachsen-Weimar and King Freiderich Wilhelm I of Prussia.

To get to the point here, he proposed the existance of a substance, Phlogiston, that was a component of many things that was released when that thing was burnt. Phlogiston was colorless, odorless, and weightless and it spoke to the question why something, once burnt, could not be burnt again. Ash, for example, was completely deflogistated matter. It contained no more phlogiston and was therefore impervious to further burning.

Additionally, air could fill with phlogiston, becoming saturated. When this happened, the principle of diffusion Screen Shot 2016-06-10 at 4.26.42 PMwould kick in to prevent further diffusion of phlogiston out of a substance. Recall that the basic principle of diffusion is that substances go from regions of high concentration to regions of low concentration (Actually, the random movement of particles will continue unendingly. The apparent result of this movement is that a non-random, concentrated source of particles becomes a random distribution that is effectively uniform. Actually, the particles are still moving, but the random distribution appears stable).

It sets up a simple equation for combustion of any (flamable) thing like this:

Phlogiston(s) + heat + something else –> Phlogiston(g) + ash + energy

Actually, it’s a great hypothesis. It does a servicable job in predicting the behavior of a combustible material in a simple system.  Imagine that phlogiston = carbon. This phlogiston / carbon exists in different forms around us: a waxy hydrocarbon chain in the candle, CO2 in the air, and as the backbone of sugars. However, it fails to recognize a couple of important things too: Mass doesn’t just disappear, the CO2 does have mass, of course, but it’s harder to appreciate. Also, flames don’t necessarily go out because of too much CO2 in the surrounding air, but because of a lack of something else, Oxygen.

However, it does fail to recognize a couple of important aspects. First, mass doesn’t just disappear during combustion. What remains as ash is lighter than the starting material.  CO2 is released and despit that fact that it is harder to appreciate, it does have mass. Second, flames don’t necessarily go out because of too much CO2 in the surrounding air, but because of a lack of something else.

preistly making o2It was by following in Stahl’s footsteps that Joseph Priestley discovered oxygen. Priestley had a knack for studying gasses. He was good at capturing and manipulating them in a controlled way. The figure to the left is an apparatus  of a type common to Priestly’s work, where a substance is heated (e.g., KClO3) to boil off a gas (e.g. O2) in a way that the gas displaces water in an inverted flask so that it may be captured in pure form.

Priestley found that oxygen purified in this way could refresh deflogistated (-perhaps, phlogistated?)26844_lg air allowing it to support combustion once more. It could also rescue an animal from suffocating in a bell jar (something that Preistley did enough that is sounds almost like a hobby of his.) The idea that air was composed of numerous components was a new one, and already Preistley was purifying these substances and demonstrating their requirement for life and for chemical reactions.

So, how does this change the way we needed to think about phlogiston?

It explains that mass doesn’t just disappear when burnt. It goes somewhere, it becomes something else (CO2). It changes the requirement for combustion from one considering the diffusion of matter out of one thing and into the air into a chemical conversion of something into something else.

Instead of the Phlogiston equation, we have the combustion reaction (either proceeding until completion or not):

Screen Shot 2016-06-12 at 8.51.00 PM

Phlogiston might still fit in as carbon if we are insistant, but now we see that something else is required as well: Oxygen.

Flames don’t necessarily go out because of too much Phlogiston (CO2) in the surrounding air, but because of a lack of something else, Oxygen.

The importance of Stahl’s work was not that he was right or wrong, but that Stahl was attempting to bring rigor and experimentation into science. In medicine and chemistry, Stahl believed in taking an empirical approach to his work. Ultimately, this was a stepping stone from the pseudoscience of alchemy to the real science of chemistry.

chemistry_alchemy

:istr makes a nucleophilic attack on chemy, resulting in the leaving group (Al) to leave and precipitate out.

 

 

 

 
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Posted by on June 14, 2016 in Uncategorized

 

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DNA Replication

DNA Replication occurs during the S (Synthesis) phase of cell cycle. The purpose of DNA replication is to create an identical copy of all the DNA in the cell so that, following cell division, both daughter cells will have complete copies of all the information required to build a cell and do all the things the cell does.

Data from several laboratories were elegantly integrated by the work of Watson and Crick to describe the structure of DNA as comprised of two anti-parallel strands bound together by polar (hydrogen) bonds between one purine and one pyrimidine.  Including:

1. Erwin Chargaff ‘s observations that

a) DNA was 50% purine (A and G) and 50% Pyrimidine (C and T) and

b) the proportion of A = the proportion of T; the proportion of C = the proportion of G .

2. Rosalind Franklin’s X ray crystallography data that indicated that DNA had a regular, repeating pattern and the molecule was of a specific width.

3. Oswald Avery’s group along with Hershey and Chase established that DNA was the genetic material (therefore making the structure of this molecule of high importance)

4. Knowledge of the distance between molecules engaged in hydrogen bonds.

5. Knowledge of the chemical properties of nucleotide molecules, comprised of hydrophilic deoxyribose sugars and phosphate groups and hydrophobic bases.

Altogether, this information provided enough background for the pair of researchers to arrive at the structure of DNA by engaging in model building.

How this all leads into the mechanism of DNA replication comes down to the following brief statement at the end of Watson and Crick’s Seminal Paper of the structure of DNA:

“It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.”1

 

What did they mean by this?

“The novel feature of this structure is the manner in which the two chains are held together by the purine and pyrimidine bases…joined together in pairs, a single base from one chain being hydrogen-bonded to a single base from the other chain… [O]nly specific pairs can bond together. These pairs are: adenine…with thymine…, and guanine… with cytosine.”1

So, if the sequence of DNA bases on one strand dictates the sequence of the other, then each of the strands can be used as a template to make another. When this is done with each of the two strands, the result is two identical DNA molecules.

It’s one thing to say that it hasn’t escaped your notice that there is a mechanism for duplicating DNA inherent in its structure, but quite another to say that you know how it works.

This was the question that Matthew Meselson and Franklin Stahl were to solve in 1958.2 They imagined three possibilities:

  1. A Conservative method of replication – the original DNA splits open and new strands are made based on that information, then the original strands comes back together and the new strands zip together. We conserve both strands of the original copy.
  2. A Semi-Conservative method of replication – The original DNA splits open and new strands are synthesized to pair with each of the originals, the new DNA then exists with one original strand and one new one.
  3. A Non-Conservative / Dispersive method of replication – Frankly, I don’t know how this would work, but the result would be two new DNA molecules where bits of each strand of each molecule may be from the original or the new DNA.

How to distinguish between these methods?

Meselson and Stahl devised an experiment that in which they grew the bacteria, E. coli in broth containing DNA made of two different isotopes of Nitrogen. In one broth, let’s call it the ‘light’ broth, they had the light form of DNA with 14N, in the other, ‘heavy’ broth, they had the heavy form of DNA with 15N.

One really is heavier than the other. When they are centrifuged, they will come to rest at different ‘heights’ in the tube.

If the bacteria is grown in broth containing only the heavy DNA, and that DNA is harvested and spun down, you would see a tube like (a) containing a single band of the heavy DNA.

If than bacteria was moved into a new medium containing light DNA, and DNA was allowed to replicate once,

Assuming  semi-conservative or dispersive models of development – you would see (b) a single band of intermediate density – because all new DNA would be partly heavy and partly light.

Assuming the conservative model – you would see (c) two distinct bands – one heavy and one light.

So this immediately tests for or against the conservative model.

The actual result was a single intermediate band was found. This eliminates the conservative model of replication, but a second round of replication in the light broth is required to discriminate between those two models.

If the Semi-Conservative model is correct, then the intermediate band would remain, but a new light band would show up (d).

If the Dispersive model is correct, then the intermediate band would inch upwards (become lighter) as more light elements are mixed in randomly within the strands. (e)

What they found was exactly like that pictured in figure d. Further, if the bacteria were allowed to grow for more generations, the ‘light’ band of DNA would become larger as more light DNA is created, while the intermediate band will remain indefinitely.

 

 

Meselson and Stahl with Chase

 

 

References

  1. Watson. J. D. and Crick F.H.C.  “A Structure for Deoxyribose Nucleic Acid”  Nature 171, 737-738 (1953).
  2. Meselson, M. and Stahl, F.W. (1958). “The Replication of DNA in Escherichia coli”. PNAS 44: 671–82.
 
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Posted by on November 27, 2012 in Education

 

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