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Assignment 2 Article Summary

Riboﬂavin-sensitized photooxidation of isohumulones and derivatives
Kevin Huvaere, Karsten Olsen, Mogens L. Andersen, Leif H. Skibsted, Arne Heyerick and Denis De Keukeleir
 * [Full Marks JCB]**

Link: http://pubs.rsc.org/en/content/articlepdf/2004/pp/b316210a?page=search DOI: [|10.1039/B316210A]

The flavor component in beer often referred to as “skunk,” is produced by the presence of isohumulones in the beer. When the beer is exposed to light between 350 and 450nm, riboflavin absorbs the light and reacts with the isohumulones to create the off-flavor in the beer.

3-Methylbut-2-ene-1 thiol (MBT), a sulfur and carbon chain compound also known as “skunky thiol” is the primary reason for this particular and bitter “off” flavor in beer known as light struck flavor (LSF). It is developed within the beer from iso-ALPHA-acids (isohumulones) that mix with riboflavin excited by light energy and a sulfur source. Concentrations of the MBT are detectable at just a few nanograms per liter.
 * Introduction**

Isohumulones are formed from humulones found in hops and are released during the boiling process when making beer. They come in dihydro-, tetrahydro-, cis- and trans- formations, the latter being the least stable and more likely to become MBT.

Isohumulones undergo decomposition from visible light exposure, as well as UV-B wavelength irradiation. When the light is absorbed, triplet state energy transfers through the molecule and causes Norrish Type I ALPHA cleavage. Rearrangement and loss of a carbonyl group cause a radical of 3-methylbut-2-enyl, which only has to pick up a thiol group to become MBT. This Norrish Type I cleavage renders the resulting molecule sensitive to the light exposure, but not all types of isohumulones undergo this cleavage.

Isohumulones are transparent in wavelengths in the blue part of the visible spectrum, the part of the spectrum where LSF occurs most. This means that something else must absorb the light and react with the isohumulones. Riboflavin absorbs strongly at 375 and 446 nm and is abundant in beer. The riboflavin absorbs the light and its electrons are excited, but the singlet state energy is short-lived and its triplet state energy is not stronger than that of the isohumulones. This means it could not be the molecule that transfers the energy to isohumulones. However, riboflavin in the excited state is a very strong oxidizer and can induce photooxidation in electron donating species.

Laser flash photolysis at 355 and 440 nm has been used to study the electron transfer to riboflavin in the excited state from isohumulones. Excited riboflavin can photooxidize isohumulones and thusly start the molecular chain reaction to get to MBT.


 * Experimental**

Dicyclohexylammonium salts of cis- and trans- isohumulones, potassium salts of dihydro- and tetrahydro- isohumulones, and sodium salts of humulinones were added to complex solutions containing 125 microMolar riboflavin and protected from light and degassed with Nitrogen. Two types of laser light sources were used providing both 355nm and 440 nm light, the former used to excite solutions only containing riboflavin and the latter used to excite solution of riboflavin plus isohumulone salts.


 * Results and Discussion**

Three maxima were produced in the absorption spectrum of the riboflavin at 355nm, 440nm, and 720 nm. The maximum at 440nm is most interesting because isohumulones are transparent at this wavelength and the peak performance is due solely to riboflavin in the excited state. It is also known that the isohumulone compounds were studied as salts based on their pKa of 3. These is suitable because it is a similar condition to the lager beer pH of 4.2 to 4.4.

The samples containing mixtures of riboflavin and isohumulone salts showed an increased rate of decay for riboflavin proportional to the concentration of the salts added. It can then be deduced that riboflavin is likely to be responsible for the one electron oxidation of the isohumulone salts.

Rearrangements of the humulinones and dihydro- and tetrahydro- isohumulones had small effects on the rate of change of the reaction. It was then deduced that the oxidation involved only the BETA-tricarbonyl chromophore that is common to all of the molecules of interest and has been corroborated by spin trapping experiments of isohumulone salts.

__Formation of reduced riboflavin radicals__

A transient riboflavin species (RFH’) formed from the protonation of riboflavin radical anions is seen to absorb at 520nm.

__Electron transfer from N,N,N ,N -tetramethyl-p-phenylenediamine (TMPD) to radicals derived from trans-isohumulones and dihydroisohumulones__

RFH’ does exist based on its absorption, but it is not possible to prove the existence of radicals of isohumulones in the same manner. But, isohumulone radicals appear to exist based on their reactions and stealing of electrons from TMPD and changes in the absorbance of the TMPD radical resulting.

Mixing TMPD, riboflavin and either trans- or dihydro- isohumulones showed absorption typical of radical TMPD cations. Increased rate kinetics are proportional to concentrations of the isohumulone salts, proving that the TPMD radical comes from interaction with isohumulone salts, not the riboflavin directly.

At a 440nm wavelength, TMPD radical cation peaks at 565nm when it reacts with certain isohumulone derivatives and the overall rate of reaction is fastest with trans-isohumulones.

__Thermodynamic feasibility of electron transfer from the salts of trans-isohumulones and derivatives to triplet-excited riboﬂavin__

Energy cannot be directly transferred from excited riboflavin to isohumulone derivatives because of large differences in energy. However, the change in Gibbs free energy is negative so the reaction is spontaneous. The thermodynamic aspects of the reaction show that the electron transfer is favored.

The absorption spectra obtained in this experiment show that electrons transfer from isohumulones and their derivatives to excited triplet states of riboflavin that was excited by a 440nm laser. The radicals formed by isohumulones become 3-methylbut-2-enyl radical which is likely to become MBT, the sulfur containing compound that causes the light struck flavor in beer. All four isohumulone species and humulinones are responsible for this descent to light struck flavor which contradicts the common misconception that only the cis- and trans- isomers create “skunk” thiol.
 * Conclusion**

Works Cited

Kevin Huvaere, Karsten Olsen, Mogens L. Andersen, Leif H. Skibsted, Arne Heyerick and Denis De Keukeleire Photochem. Photobiol. Sci., 2004, 3, 337-340

Assignment #3 -FAQ- completed


 * 5 Online Sources of Brewing Chemistry**

1. [|www.asbcnet.org] The American Society of Brewing Chemists’ website is available to anyone interested in learning methods of brewing, handling raw materials of brewing, problem solving in brewing, as well as professional opportunities in the brewing industry.

2. [|www.mbaa.com] A website of the Master Brewers Association of the Americas that provides interaction with other brewing professionals, resources for better brewing processes, and practical information for companies, as well as home brewers, on a global level.

3. [] A site dedicated to home brewers explaining in layman’s terms the processes, chemistry, etc. for brewing at home. Zymurgy, the journal of the The Home Brewers Association is also available on the website.

4.[|www.ibd.org.uk] A professional brewer’s resource, The Institute of Brewing and Distilling provides opportunity for professionals to contact and converse with industry colleagues about processes in fermentation and brewing, take exams and gain certifications, and keep up to date in the brewing industry.

5. [|www.foodnavigator.com] A website that provides updates and current news in food and beverage science and development.