Lignin and the cell wall

Trees are the largest organisms on earth

Plants first had to adapt over millions of years Wing and Chapple 2010, New Phytologist 187:273

Surviving pests and fire- lignin Lignin protects wood’s sugars Lignin has good thermal stability* orest_fire_aftermath.jpg&usg

Typical composition of different plant types J. J. Bozell, et al. U. S. DOE, Report PNNL , Washington, DC 2008.

In reaction wood lignin content is changed As highlighted by Rowell et al. Cell Wall Chemistry 2005

Lignin content within the cell wall of plants

How does lignin get into the wall? Lignification occurs after polysaccharide scaffolding is in place. Terashima et a J. Wood Sci 55:409

Monomers of lignin Courtesy W. Glasser

Chiang, 2002 Nature Biotech 20:557

Polymerization pathways: Free Radical Coupling M= molecules that form covalent bonds, while propagating reactive species (*) I* M M M M M M M I-M* I* +M I-M* I-M-M* +M I-M-[M] n -M 1. initiate 2. propagate 3. terminate Initiating reactive species Monomers With lignin the radical is coupled between monomers and then the initiator must create a new radical. Step 1 and then step 3.

Enzyme initiated polymerization Source ?, Courtesy W. Glasser

Monomer ratios are different base on plant type J. J. Bozell, et al. U. S. DOE, Report PNNL , Washington, DC 2008.

HW and SW have different monomers and this results in guaiacyl (G) and syringyl (S) lignins of different G:S ratios G:S ratio impacts properties G:S ratio changes within cell wall based on location Whetten et al Ann Rev Plant Phys Plant Mol Bio 49:585

Linkages in lignin J. J. Bozell, et al. U. S. DOE, Report PNNL , Washington, DC 2008.

Proportional linkages Saake 2007Ullmann’s Encyclopedia of Industrial Chemistry, Wiley-VCH

BIOLOGICAL vs. THERMODYNAMIC CONTROL Precursors are formed in CYTOPLASM (living cell): syr:gua ratio is under biosynthetic/genetic control Precursors are polymerized by “coupling” reactions of free radicals (which are created by enzymes): bonding types are under thermodynamic control Courtesy W. Glasser

Enzyme catalyzed polymerization Reihmann and Ritter 2006 Adv Polym Sci 194:1

Historical literature claims coniferin transported out of cytoplasm Source ?, Courtesy W. Glasser

Movement of monomers to reactive sites within wall

Locating glucosidase and peroxidase Dharmawardhana et al Plant Physiology, 107:331

Lignification with different materials present Compound middle lamellaSecondary wall

Globular modules in cell corners Zhuo Li, 2009

Smaller modules within secondary cell wall

Lignin carbohydrate complexes (LCC) Grabber et al. J. Ag. Food Chem :4984

LCC FORMATION Source ?, Courtesy W. Glasser

Lignin is usually cited as random network Lignin is optically inactive Lignin has a variety of linkages that could facilitate a cross-linked network However, there is experimental evidence and developing theories to the contrary- that lignin is oriented in the cell wall and branched, but not necessarily a highly cross-linked network.

Orientation of lignin rings within cell wall

One picture of lignin organization Modular organization High degree of order

A model of a branched lignin J. Ralph et al Phytochemistry Reviews 3: 29-60, 2004

Delignification has a large impact on structure

How big are lignin chains of isolated lignin? Polymer properties are always influenced by their molecular weight (MW) and molecular weight distribution (MWD) Isolating lignin disrupts native lignin bonds Technical lignins are isolated through kraft pulping and sulfite pulping These technologies impact size of chains Break linkages and form new linkages Newer conversion methods for biofuels are being developed (organosolv pulping and steam-explosion processing) “Gentle” isolation can be used to limit the number of broken bonds in lignin for laboratory isolation Obtain 20 to 50% of the lignin using a “milled-wood” acidolysis process

Lignin MW studies Holtman et al Journal of Wood Chem Tech 27:179 A wide range of sizes for the compounds from 3 to 4 units bonded together to 30 to 40 units bonded together.

Summary Lignin is a polymer made out of phenylpropane units The polymer is built by a process of radical polymerization outside of the cytoplasm More than one monomer type influences lignin structure for different plants There is some spatial patterning of lignin linkages Lignin can bond with carbohydrate creating lignin carbohydrate complexes Attempts to examine lignin’s structure disrupts lignin’s structure Progress has been made to view in situ lignin with newer analytical techniques HSQC 2-D NMR