Lecture 9 Lignin Biosynthesis Wood Chemistry PSE 406 Lecture 9 Lignin Biosynthesis PSE 406 Lecture 9 1

Class Agenda Lignin I Basic Lignin Biosynthesis Nomenclature Methoxyl Groups Phenoxy Radical Formation Radical Coupling PSE 406 Lecture 9

Lignin-introduction Phenolic Polymer - the glue that holds the fibers together Increase the mechanical strength properties of plants Present in higher plants in vascular tissues (liquid transport and mechanical strength) Distribution in the cell wall and parts of tree differs Mechanical barrier against pathogens Polydisperse (Large and Small Polymers) 3 Dimensional Networks - Branched Polymer PSE 406 Lecture 9

Lignin distribution PSE 406 Lecture 9

Lignin Biosynthesis I PSE 406 Lecture 9 The biosynthesis of lignin has been very well studied and a significant amount is known about the various pathways. There is still, however, a significant amount left to discover. In this class, we are only going to address this topic superficially. In the above slide, the starting material for this whole process is CO2. Through a large number of photosynthetic steps, CO2 is converted into glucose. Glucose is converted through other pathways into other monosacharides, cellulose, hemicelluloses, lignin and extractives. In the formation of lignin, glucose is converted through the shikimic acid pathway into shikimic acid which is further converted to either L tyrosine or to L phenylalanine. These are both amino acids. Notice that they are also now both aromatic compounds. PSE 406 Lecture 9 2

Lignin Biosynthesis II L-tyrosine and L- Phenylalanine are converted through a series of reactions known as the cinnamate pathways into three different chemicals which are the precursors of lignin. It is important to note that all of these reactions and the reactions discussed in the previous slide are controlled by enzymes. The tree is manufacturing exactly the chemicals it wants with exactly the correct stereochemistry. Using advances made over the last two decades, scientists have been recently manipulating these pathways in order to change the structure of lignin or to reduce the amount of lignin in the tree. Reduced lignin trees have now been produced through genetic manipulation. (Why would this be important?). These trees are still quite young so it will still be some time before their properties are known. The three structures shown above, p-coumaryl alcohol, coniferyl alcohol, and sinapyl alcohol are the precursors (monomers) for lignin. It is important that you learn the structures of these compounds as we will be working with these quite a bit. PSE 406 Lecture 9 3

Lignin Biosynthesis Nomenclature Side Chain The lignin precursor shown in this figure is referred to by lignin chemists as a phenylpropane unit or a C9 unit. This is obviously because: a. it is a combination of a phenol ring and a propane (C3) side chain and b. because there are 9 carbons in the structure ignoring the methoxyl group. It is important to note the numbering of the ring and the naming of the side chain. This is something you need to know. The carbon on the side chain nearest the ring is the alpha carbon. Next up the line is the beta carbon and finally the gamma carbon. Some lignin chemists have recently taken to numbering the entire structure so that the three side chain carbons are numbered C7, C8, and C9 instead of alpha, beta, and gamma. } Phenylpropane Unit C9 Common Names PSE 406 Lecture 9 5

Lignin Structure Methoxyl Content As you can see in the slide, grasses, softwoods and hardwoods all use different percentages of lignin precursors. There are wide differences in the levels of lignin precursor used between these groups and within different species in each group. Grasses (bamboo, reeds, etc.) use the highest levels of p-coumaryl alcohol. Softwoods and hardwoods use only very limited levels of this precursor. On the average, coniferyl and sinapyl alcohol are used equally by grasses although this varies dramatically between species. Softwoods are made up almost entirely from coniferyl alcohol. If sinapyl alcohol is used, it is in only very trace amounts. Hardwoods use both coniferyl and sinapyl alcohols. The ratio of these two varies significantly between hardwood species. As you can see, the difference between these precursors is the number of methoxyl groups (none, 1, or 2). While this doesn’t seem like a large difference, the number of methoxyl groups affects the size and shape of the resulting lignin molecule PSE 406 Lecture 9

Lignin Biosynthesis Nomenclature Once incorporated into lignin, the ring structures of the precursors are given these names. PSE 406 Lecture 9 4

Lignin Biosynthesis Enzymatic Formation of Phenoxy Radical The initial step in the polymerization of lignin is an enzymatic directed one electron abstraction forming a phenoxy radical. The lignin precursor is transported to the lignification site in a water soluble form. It is attached to a sugar. This dimer is cleaved leaving the precursor. Polymerization is initiated through an enzyme directed abstraction of one electron from the phenolic hydroxyl leaving a phenoxy radical. PSE 406 Lecture 9

Lignin Biosynthesis Resonance Forms of Phenoxy Radical The phenoxy radical is only one of several resonance structures. This figure shows a second resonance structure with the free radical on the 5 carbon of the ring. PSE 406 Lecture 9

Lignin Biosynthesis Resonance Forms of Phenoxy Radical This figure shows the resonance forms of the phenoxy radical of coniferyl alcohol. Notice that the radical is located on the following positions and only the following positions. The phenolic hydroxyl on the 4 (ring) carbon. The 5 (ring) carbon. The 1 (ring) carbon. The 3 (ring) carbon. It is important to note that because of steric reasons (the methoxyl group), even though this position has a free radical structure linkages are typical not formed. The b (beta) position of the side chain PSE 406 Lecture 9

Lignin Coupling -O-4 PSE 406 Lecture 9 Free radical lignin precursors couple forming covalent bonds linking the structures. In the figure above, the coupling is between 2 coniferyl alcohol molecules; one with with a free radical on the b carbon and one with a phenoxy radical. The linkage formed above is known as a b-O-4 linkage. You will see later that this is the most common lignin linkage. In the structure with the b radical, the ring structure is known as a quinone methide structure. This is a very reactive structure. It is very susceptible to nucleophilic addition reactions. In the above example, water (or an alcohol) adds to the a carbon forming an ether linkage. There are obviously a large number of ways the lignin precursors can couple. We will discuss these in the next lecture. An important issue which is currently being debated is whether the way the lignin precursors couple is 1. A random event, 2. An event affected by cell conditions, or 3. An enzymatically directed event. It is pretty much known that this is not a random event as research has shown that many variable will affect the linkage formation. Lewis and Sarkanen have discovered a cell protein which directs the coupling reaction to a specific linkage. To date, only one directing protein has been discovered and the linkage which is formed is also found in wood extractives. The jury is still out on this issue. PSE 406 Lecture 9