Effect of Fusiform Rust 

(Cronartium quercuum 

f.sp. fusiforme) on 

the Composition of 

Loblolly Pine Lignin

Roderquita K. Moore

Allisha N. Blood

Cherrelle I. Esekie

Forest

Products

Laboratory

General

Technical 

Report

FPL–GTR–230

United States Department of Agriculture

Forest Service
March 2014

Moore, Roderquita K.; Blood, Allisha N.; Esekie, Cherrelle I. 2014. Effect 

of Fusiform Rust (Cronartium quercuum f.sp. fusiforme) on the Composition of Loblolly Pine Lignin. General Technical Report FPL-GTR-230. 

Madison, WI: U.S. Department of Agriculture, Forest Service, Forest Products Laboratory. 5 p.

A limited number of free copies of this publication are available to the 

public from the Forest Products Laboratory, One Gifford Pinchot Drive, 

Madison, WI 53726–2398. This publication is also available online at 

www.fpl.fs.fed.us. Laboratory publications are sent to hundreds of libraries 

in the United States and elsewhere.

The Forest Products Laboratory is maintained in cooperation with the 

University of Wisconsin. 

The use of trade or firm names in this publication is for reader information 

and does not imply endorsement by the United States Department of 

Agriculture (USDA) of any product or service.

The USDA prohibits discrimination in all its programs and activities on the 

basis of race, color, national origin, age, disability, and where applicable, 

sex, marital status, familial status, parental status, religion, sexual orientation, genetic information, political beliefs, reprisal, or because all or a part 

of an individual’s income is derived from any public assistance program. 

(Not all prohibited bases apply to all programs.) Persons with disabilities 

who require alternative means for communication of program information (Braille, large print, audiotape, etc.) should contact USDA’s TARGET 

Center at (202) 720–2600 (voice and TDD). To file a complaint of discrimination, write to USDA, Director, Office of Civil Rights, 1400 Independence 

Avenue, S.W., Washington, D.C. 20250–9410, or call (800) 795–3272 

(voice) or (202) 720–6382 (TDD). USDA is an equal opportunity provider 

and employer.

Abstract

Loblolly Pine, Pinus taeda, is a common southern softwood ranging from Georgia and the Carolinas to Texas. The 

fungus Cronartium quercuum f.sp. fusiforme is one the most 

destructive forest diseases in the south. This fungus infects 

both loblolly and slash pine (P. elliottii Engelm.), causing 

canker formation that frequently kills the infected branch. 

April and early May are when the pine infection cycle occurs in Georgia. The elongated swelling of the branches is 

the result of individual attacks on different parts of the tree. 

Many of the infected trees are unsuitable for product, causing millions of dollars to be lost a year because of the death 

of infected trees. In addition, trees with large galls on the 

main stem are unsuitable for many products.

We report the chemical composition of loblolly pine infected with fusiform rust. Branches were collected in Athens, 

Georgia, and frozen. The branches not infected by fusiform 

were collected as the control. Spectroscopy was used to 

analyze lignin. This report presents and discusses the results. 

The aim of this study was to apply Fourier transform infrared spectroscopy (FTIR) and nuclear magnetic resonance 

spectroscopy (NMR) to determine whether there were any 

structural modifications to the chemical composition of 

lignin from the fungus. Published information on fusiform 

rust analyzed by FTIR and NMR is limited in the literature. 

In this paper, we present a FTIR and NMR spectroscopic 

analysis of the chemical changes occurring in a softwood, 

loblolly pine, by fusiform rust. 

Keywords: FTIR, NMR, sugar analyses, lignin, fusiform 

rust, loblolly pine, southern pine, invasive species

Acknowledgments 

Thanks to Pauline Spaine of the USDA Forest Service 

Southern Research Station for the collection of loblolly pine 

fusiform rust branch samples. Thanks to Sally Ralph, Fred 

Matt, and the Analytical Chemistry and Microscropy Lab 

(ACML) of FPL for conducting NMR, FTIR, and Klason 

analyses. 

Cover Photo: Loblolly pine infected by fusiform rust 

Content

Introduction........................................................................1

Materials and Experimental Methods................................1

 Samples..........................................................................1

 Lignin Preparation .........................................................1

 Nuclear Magnetic Resonance ........................................1

 Fourier Transform Infrared Spectroscopy......................1

Results and Discussion ......................................................1

 Nuclear Magnetic Resonance ........................................1

 Carbon NMR..................................................................1

 Fourier Transform Infrared Spectroscopy......................2

 Klason Lignin.................................................................2

Conclusions........................................................................4

References..........................................................................4
Effect of Fusiform Rust (Cronartium 

quercuum f.sp. fusiforme) on the 

Composition of Loblolly Pine Lignin

Roderquita K. Moore, Research Chemist 

Forest Products Laboratory, Madison, Wisconsin

Allisha N. Blood, Summer Intern, Chemistry

Spelman College, Atlanta, Georgia 

Cherrelle I. Esekie, Summer Intern, Chemistry

Clark Atlanta University, Atlanta, Georgia

Introduction

Loblolly pine, Pinus taeda, is a common southern softwood 

ranging from Georgia and the Carolinas to Texas. The fungus Cronartium quercuum f.sp. fusiforme is one of the most 

destructive forest diseases in the South. This fungus infects 

both loblolly and slash pine causing canker formation that 

frequently kills the infected branch (Brown and Coder 

2009). April and early May are when the pine infection cycle occurs in Georgia. The elongated swelling of the branches is the result of individual attacks on different parts of the 

tree. Many of the infected trees are unsuitable for product, 

causing millions of dollars to be lost a year because of the 

death of the infected trees. In addition, trees with large galls 

on the main stem are unsuitable for many products.

Changes in wood chemistry resulting from fungal decay 

of Scots pine (P. sylverstris L.) have been studied directly 

using Fourier transform infrared spectroscopy (FTIR) and 

nuclear magnetic resonance spectroscopy (NMR). Scots 

pine sapwood was exposed to brown rot, selective white rot, 

and nonselective white rot fungi, and the decay process was 

followed using FTIR (Pandey and Pitman 2003). After 

12 weeks, the wood exposed to the brown rot fungus resulted in progressive increase in lignin content relative to cellulose and hemicellulose, whereas the lignin content of the 

wood exposed to the selective white rot decreased as decay 

proceeded. For the wood exposed to the nonselective white 

rot wood, both occurred. 

The aim of this study was to apply FTIR and NMR to determine whether the pathogen caused any structural modifications to the chemical composition of lignin. 

Materials and Experimental 

Methods

Samples

The samples selected for this study were loblolly branches 

affected by fusiform rust. The control was branches that 

grew in the same environment but were not attacked by 

fusiform rust. The branches were collected by Pauline 

Spaine of the U.S. Forest Service Southern Research Station 

from loblolly pines located near Athens, Georgia. 

Lignin Preparation

These branches were debarked, cut, and ground into 

40-mesh grain sizes. The branches (50–100 g) were soxhletextracted with 700 ml of acetone:water (9:1) and ethanol 

(95%) for 8 hours. The isolation of milled wood lignin 

(MWL) was slightly modified from Björkman MWL 

(Björkman 1956). 

A portion (500 mg) of MWL was acetylated with equal portions of pyridine and acetic anhydride for 4 hours. The mixture was then co-rotary evaporated with toluene and then 

acetone to remove the reaction co-products. The acetylated 

MWLs were then dissolved in deuterated acetone for NMR 

analysis. Acetylation was not done for the FTIR analysis.

Nuclear Magnetic Resonance

A Bruker 250 DPX Nuclear Magnetic Resonance Spectrometer (Bruker Corporation, Billerica, MA) was used to 

acquire 1H, 13C, Distortionless Enhancement by Polarization 

Transfer (DEPT) 135 and Heteronuclear Single Quantum 

Coherence (HSQC) spectra using standard Bruker pulse programs. The internal reference was set to the center peak of 

the acetone solvent 2.04/29.83 ppm.

Fourier Transform Infrared Spectroscopy

A Mattson Galaxy 5020 FT-IR system (Mattson Instruments, Inc., Madison, WI) was used to characterize KBr 

pelleted samples. FT-IR spectra were obtained in transmission mode by averaging 64 scans from 4000 to 450 cm−1 at 

8 cm−1 resolution.

Results and Discussion

Nuclear Magnetic Resonance

Both the proton and carbon NMR spectra were acquired, 

along with a DEPT experiment that was run to distinguish 
General Technical Report FPL–GTR–230

2

the degree of protonation on the carbons in the 13C NMR. 

A short range C-H correlation, HSQC, experiment was run 

to determine C-H correlations for signals of interest.

Carbon NMR

A comparison of the 13C spectra from infected and noninfected (control) loblolly by fusiform rust shows areas of 

interest at approximately 170, 122, and 68 ppm (Fig. 1). The 

region between 173 and 169 ppm shows some differences 

(Fig. 2a). The signals shown in Fig. 2a are due to the carbonyl group of the acetate moieties (Shown by arrows). The 

primary, secondary, and phenolic hydroxyl group acetates 

are at 171, 170.5, and 168 ppm, respectively, in this region. 

The signal at 172 ppm is due to residual acetic acid from the 

derivitization procedure and may be disregarded. 

The spectra of the carbonyl group were normalized on the 

gamma carbonyl signal 171 ppm. The carbonyl at the beta 

position 170.5 ppm intensity was the same; however, peak 

broadening was observed for the infected branches, indicating that neighboring groups were changing. The free hydroxyl phenolic acetates position at 169 ppm intensity increased, indicating there are more free phenolic side groups 

in the infected branches available for acetylation. Literature 

indicates that an increase in lignin and more phenolic compounds are observed in the infected sample (Rowan 1970).

Between 134 and 122 ppm were also differences (Fig 2b). 

Two methine carbon signals seen at 129 and 122 ppm are 

missing or diminished for the infected sample. This observation indicates that oxidation is occurring and is forming 

an aldehyde because of the fungal attack. Oxidation of this 

kind is observed in wood-decay-forming benzoates (Eriksson and others 1999).

The proton NMR observed peaks at 5.4 and 5.1 ppm (see 

arrows) in the control proton spectrum (Fig. 3). From 2D 

NMR, the two peaks seen at 68.8 and 68.3 ppm in the 13C 

NMR correlates, respectively, to the two proton peaks. From 

the DEPT experiment, the peaks observed represented CH 

groups. HSQC showed the proton was low field. This observation correlated with the diminished peaks observed in the 

aromatic region at 129 and 122 ppm. 

Fourier Transform Infrared Spectroscopy

FTIR spectra of infected and control loblolly pine are shown 

in Figures 4–7. The areas of interest are 2900 and 1640 cm–1 

(See arrows). A strong hydrogen bonded (O-H) stretching 

absorption is seen at 3400 cm–1 and a prominent 

C-H stretching absorption around 2900 cm–1. In addition, 

there are many peaks in the fingerprint region between 

1800 and 600 cm–1. It has been shown that the guaiacyl 

component (softwood lignin) absorbs near 1268 and 

1230 cm–1 (Pandey 1999; Pandey and Pitman 2003). 

One area of difference occurs at around 1650–1600 cm–1; 

unfortunately, this is also an area where overlapping with 

moisture occurs. Several spectra were run to eliminate other 

possibilities for the difference (Fig. 5). Both samples as well 

as an extra sample of loblolly pine were oven dried (Fig. 6). 

The peak at 1640 cm–1 remained for the infected sample.

Another area of difference occurs at 2900 cm–1. This is the 

area where C-H stretching arises. The small new shoulder 

that forms on the infected loblolly pine is not seen on either 

the control or the loblolly sample. This could be a result of 

a new CH bond that has been formed (Fig. 7). Because of 

the shoulder at 2900, this could result in the modification 

at 1640 where we see broadening of the band of infected 

sample. Quinones are known intermediates in modification 

of lignin by fungi. The shoulder at 2900 cm–1 and the peak 

broadening at 1640 cm–1 could be due to this degradation of 

lignin by the fungi (Eriksson and others 1999; Heitner and 

others 2010).

Klason Lignin

The percentage difference in Klason lignin (Hatfield and 

others 1994) was calculated from the control and the infected loblolly pine (Table 1). The Klason lignin percentage 

increased for the infected sample. This percentage increase 

correlates with modifications in the chemical composition 

changes in the lignin that were observed in the NMR spectra. The percentages for the mannan, glucan, and xylan 

and total carbohydrates decreased significantly for the infected loblolly pine. The infected sample acid soluble 

Figure 1. Full spectra of 13C NMR. The red line (a) is 

loblolly pine infected by fusiform rust. The black line 

(b) is the control sample of loblolly pine. 
Effect of Fusiform Rust (Cronartium quercuum f.sp. fusiforme) on the Composition of Loblolly Pine Lignin

3

Figure 2. (a) The red line shows the 13C NMR between 173 and 167 ppm of loblolly pine infected by 

fusiform rust and the black line is the loblolly pine control. (b) The red line shows 13C NMR between 

135 ppm and 120 ppm of loblolly pine infected by fusiform rust, and the black line shows the loblolly pine control. 

Figure 3. Partial expansion of 1H NMR spectrum. The 

red line (a) shows loblolly pine infected by fusiform 

rust. The black line (b) shows the control sample of 

loblolly pine. 

Figure 4. Full FTIR spectra of (a) infected (A2) and 

(b) control (UA) loblolly pine. 
General Technical Report FPL–GTR–230

4

percentage increased, leading us to conclude that low molecular weight acids were formed during the fusiform rust 

attack. 

Conclusions

FTIR and NMR spectroscopy were used to examine changes 

in the lignin chemical composition after fusiform rust fungal attack on loblolly pine. Changes were observed in the 

spectra for the infected loblolly pine lignin. NMR spectra 

showed changes in the carbonyl groups. These changes 

indicated there are more ß-O-4 linkages forming that could 

result in higher molecular weight because of the attack. 

NMR results supported the observation from the FTIR that 

modification of the lignin is occurring. Klason lignin percentage increased from the fungal attack. The acid soluble 

from the infected lignin increased, which could indicate that 

low molecular weight compounds are formed. Sugar analysis percentage differences were similar to what is found in 

literature (Rowan 1970), even though fusiform rust is not a 

wood-decaying fungi glucan, and xylan decreased because 

of fungal attack. The chemical composition of lignin was investigated, and from the study lignin chemical composition 

is being modified because of the fusiform rust attack in loblolly pine. Future investigation will include chromatography. 

This analytical method will be added to further investigate 

what type of changes and degradation are occurring because 

of this fungal attack. 

References

Brown, E. II; Coder, K. 2009. Fusiform rust on pines. Circular 440. Athens, GA: The University of Georgia, College 

of Agricultural and Environmental Sciences Cooperative 

Extension. 4 p.

Björkman A. 1956. Studies on finely divided wood. Part 1. 

Extraction of lignin with neutral solvents. Svensk Papperstidn, 59:477–85.

Eriksson, K-E.; Blanchette, R.; Ander, P. 1999. Microbial 

and enzymatic degradation of wood and wood components. 

Berlin: Springer-Verlag. 

Figure 5. Full FTIR spectra of (a) dioxane, (b) infected (A2) loblolly pine, (c) control (UA) sample 

of loblolly pine in dioxane: H2O and (d) control (UA) 

loblolly pine.

Figure 6. Partial FTIR spectra of oven-dried (a) loblolly pine, (b) infected (A2) loblolly pine, (c) control 

(UA) loblolly pine.
Effect of Fusiform Rust (Cronartium quercuum f.sp. fusiforme) on the Composition of Loblolly Pine Lignin

5

Hatfield, R; Jung, H.J; Ralph, J; Buxton, D.; Weimer, P. 

1994. A comparison of the insoluble residues produced by 

the Klason lignin and acid detergent lignin procedures. Journal of the Science of Food and Agriculture, 65:1. 51–58. 

Heitner, C.; Dimmel, D.; Schmidt, J. 2010. Lignin and lignans advances in chemistry. 1st Edition. Boca Raton, FL: 

CRC Press. 

Pandey, K.K. 1999. A study of chemical structure of soft 

and hardwood and wood polymers by FTIR spectroscopy. 

Journal of Applied Polymer Science, 71:1969–1975.

Pandey, K.K; Pitman, A.J. 2003. FTIR studies of the 

changes in wood chemistry following decay by brown-rot 

and white-rot fungi, International Biodeterioration & 

Biodegradation, 52(3):151–160.

Rowan, S.J. 1970. Fusiform rust gall formation and cellulose, lignin, and other wood constituents of loblolly pine. 

Phytopathology, 60:1216–1220.

Figure 7. Full FTIR spectra of oven-dried (a) loblolly 

pine, (b) infected (A2) loblolly pine, (c) control (UA) 

loblolly pine.

Table 1. Klason lignin percentage difference of infected and control 

loblolly pine including sugars found in lignin

Sample 

Control (UA)

(%)

Infected (A2)

(%)

Difference

(%)

Higher

percentage 

AI ash 0.00 0.20 0.20 A2 

AS lignin 0.80 1.10 0.30 A2 

K. lignin 33.70 38.40 4.70 A2 

Arabinan 1.24 1.99 0.75 A2 

Galactan 4.83 5.16 0.33 A2 

Rhamnan ND 0.67 0.67 A2 

Glucan 33.00 26.74 6.26 UA 

Xylan 9.52 5.92 3.60 UA 

Mannan 7.62 7.53 0.09 UA 

Total carb 56.21 48.01 8.20 UA 

Total yield 90.70 87.70 3.00 UA