I have been having a discussion on another forum, on how it is possible for water to pickup chlorophyll in an extraction, when chlorophyll is basically a hydrocarbon, which is mostly insoluble in water.
I did enough research to know that I was in over my head with chemistry that I took on the late fifties and early sixties, so I asked Joe, our budding biochemist, to take a run at it.
In quick summary, before Joe's response, "define soluble?" The word soluble means different things in biochemistry, than it does in organic chemistry, because of the behavior of the molecules of life.
The chlorophyll molecule has a magnesium (Mg) at its rings center, which makes it ionic and water-loving (hydrophilic) and a ring that is water fearing (hydrophobic) with carbonyl groups near a tail that make it polar (also hydrophilic).
It is held in place in the plant within a water-soluble material known as water-soluble chlorophyll-binding protein (WSCP). WSCP is soluble in water, and mostly insoluble in polar alkane alcohols and non polar alkanes.
Chlorophyll is readily soluble in alcohol, mostly insoluble in non polar alkanes like butane and hexane, and has some special relationships with polar water, because of its polar and ionic groups.
Mostly is a key word in all cases, because of chlorophylls charged polar end and non polar hydrocarbon ring with the ionic Mg.
Before I finish summarizing, here is Joe's response to the question of how water can remove and transport basically insoluble chlorophyll molecules, as well as how saturating water with NaCl table salt aids in the process of washing unwanted chlorophyll out of extractions that have gone awry:
Chlorophyll Info by Joe
Chlorophyll is an intra-membrane chemical within a thylakoid. A thylakoid is a membrane-bound compartment inside chloroplasts. The thylakoid membranes of higher plants are composed primarily of phospholipids and galactolipids that are asymmetrically arranged along and across the membranes.
Chlorophyll is shown as photosystem I and II in this illustration.
Both phospholipids and galatolipids have hydrophilic (water-loving) heads and hydrophobic (water fearing) tails.
In biology this theme is used in almost all life forms to compartmentalized for energy storage, isolate invaders or encase their genome to protect it and many others reasons.
Solubility is a term that has more than one definition. In inorganic chemistry it refers to waters (or another compound) ability to break covalent and ionic bonds of most compounds, dependent on time temperature and pH. The “solublized” atoms are then bonded to their polar opposite ion H3 (+) or OH (-) and are in solution.
In biology however, it is used in the first tense but, it is also used to describe the ability of an organic molecule or complex to form an association with water and be in solution but not be “solubilized” by it. Proteins and other organic molecules use charged ions such as phosphate (PO4) and Sodium (Na) to form micelles. Micelles are little balls of hydrophobic molecules surrounded by a charged ion.
Just like a cell's membrane bilayer. Sometimes micelles are formed by complexes of proteins surrounding a small molecule for transport through water.
Chlorophyll specifically, is only able to form complexes with other molecules to stay in solution at biological pH (7.4). Its natural environment is at a pH of around 4 not 7.4. At this pH it has a net charge of -2 so that it can form a chemo-gradient for electron transport during photosynthesis.
So since pH =-log [OH-/ H+] when at pH 4 the [H+] concentration is higher than the [OH-] thus creating an environment that is more likely to associate with the (-) charged area of the chlorophyll molecule. Hence the low solubility of unbound chlorophyll in water, the large hydrophobic areas compress together and present their hydrophilic areas to exclude water from the center, becoming a mass that will sediment in water.
The point of this is to illustrate that while purified chlorophyll is not likely to stay in solution in pure water; we don’t extract pure chlorophyll and we don’t use pure water.
We use brine to keep the charge on the phospholipid bilayer (Na+ with PO4-3) and no detergents. The alcohol (ROH+) wants nothing to do with Na+ while in its protonated (H+) state.
Non polar solvents for obvious reasons won’t either and also won’t form much of an emulsion with alcohols in their bent state because; the alcohol is denser and forms a micelles like layer to protect itself from the charged Na+. If there is an excess of alcohol it will start forming an emulsion layer at the upper interface.
The phospholipid bilayer of the chloroplasts and of the thylakoid being intact or mostly so, prevent the chlorophyll from being disassociated with the Na+ water and are able to be excluded from alcohol or non-polar solvents .
Alcohol is able to associate with chlorophyll and proteins in its native conformation but not when bent by Na+ because the electron pool concentrated at the (O-) repels the (-) region of the chlorophyll molecule and the PO4-3 of the membrane bilayer.
If the membranes have been broken up by a detergent or broken down by enzymes then the only way to exclude chlorophyll from a non-polar solvent or alcohol is with lots of Na+ and water. Because of the large non polar area of the chlorophyll molecule it can more easily form a hydrophobic interface and shield the charge in the center.
The salt exposes the charge (because ionic is a stronger bond than Van Der Wals forces) and precipitates the chlorophyll into water.
So in conclusion, chlorophyll is not wholly soluble in water, but in its biological complex is able to associate with it.
By manipulating charge/charge interactions chlorophyll can be forced into solution with water and away from polar organic and non-polar solvents. While it doesn’t meet the inorganic chemistry definition of solubility, it will form micelles complexes with an ionic solution and it can be precipitated from that solution under the right conditions.
From a biological perspective chlorophyll can also be solubilized by any solvent under the right conditions.
Sooooo, back to my summary, it appears that some solvents can wash away the cement binding the chlorophyll in the plant cells and free the chlorophyll to be washed away as a micelles, but it also exposes the chlorophyll to the solvent, which in the case of alcohol, will readily dissolve it and hold it in solution.
Freezing the material prior to an extraction, may be holding both the chlorophyll and the WSCP locked up in ice, so that neither the water present or the solvent can reach the chlorophyll.
Merck Index lists chlorophyll as practically insoluble in non polar solvents, so the difference between practically and totally insoluble may offer a clue, as well as butanes practically insolubility in water.
While most sources list n-Butane as insoluble in water, its actual solubility is 0.0325 vol/vol, at 1 atmosphere pressure and 20C/68F. That is 32 ml/liter, which may be enough to account for the light electric green hue that occurs, by both washing away the WSCP and holding some of the chlorophyll in suspension.
When we saturate the water with salt, before washing a polar extract suspended in a non polar solvent like hexane, it forces the chlorophyll into solution and washes it away.
Explanation by Joe, summarized by Graywolf