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BIOLOGICAL PERFORMANCE OF CUPROUS AND
CUPRIC COPPER AGAINST HOOD
DECAY FUNGI

presented by

Reining Cut

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BIOLOGICAL PERFORMANCE OF CUPROUS AND CUPRIC COPPER
AGAINST WOOD DECAY FUNGI

By

Weining Cui

A THESIS

Submitted to
Michigan State University
in partial fulfillment of the requirement
for the degree of

MASTER OF SCIENCE
Department of Forestry

1999

ABSTRACT

BIOGICAL PERFORMANCE OF CUPROUS AND CUPRIC COPPER
AGAINST WOOD DECAY FUNGI

By
Weining (hi

Copper naphthenate and copper ethanolamine solutions with various copper metal
concentrations (0 to 1%) were formulated and used to treat southern yellow pine and
sugar maple. After treatment, a post-treatment steaming was conducted at various length
of time, from O to 240 minutes. Atomic absorption spectrometer (AAS) was used to
analyze the copper concentration in treating solutions and in treated wood.

A colorimetric method based on the specific reaction of cuprous copper (Cu (1))
and 2,2'-biquinoline in acetic acid matrix was used to monitor and to quantify Cu (1) in
solutions and in treated wood. Data clearly indicate that the post-treatment steaming
promotes the conversion of Cu (H) to Cu (1). This was confirmed by the increase of Cu
(I) with the increase of the post-treatment steaming duration period.

A laboratory soil block test was conducted using both white and brown rot fungi.
The decay index of copper naphthenate treated southern yellow pine and sugar maple,
and copper ethanolarnine treated sugar maple challenged by copper tolerant brown rot,
Poria placenta, were significantly influenced by the content of Cu (I) / Cu (11). The
weight loss was higher in treated wood with higher Cu (1) content, suggesting that the
toxicity of Cu (1) to decay fungi is lower than that of Cu (II). It was hypothesized that
Cu20 is less toxic because it is less soluble in water, which blocks the fiirther interactions

between copper and fungi or wood components.

ACKNOWLEDGEMENT

I would like to express my sincere gratitude to my advisor Dr. D. Pascal
Kamdem, for his guidance, support and patience during the course of my graduate
studies. Thanks to the committee members, Dr. Prof. Emmett Braselton and Dr. Eugene
A. Pasek for their support and their valuable suggestions.

I am much obliged to Dr. Jun Zhang, Ismail Jusoh and Justin Zyskowski for their
help and advise. Thanks to the student workers for their help in the sample preparation.

The USDA-C SREES Eastern Hardwood Utilization Program in the Department
of Forestry at Michigan State University provided funding for this research. The
continued financial support throughout my research program was gratefully
acknowledged.

I would like to thank my parents for their love and understanding.

iii

TABLE OF CONTENTS

LIST OF TABLES ................................................................................................... vii
LIST OF FIGURES .................................................................................................. viii
INTRODUCTION .................................................................................................... 1
1.1 Background and proposed work ......................................................... 1
LITERATURE REVIEW ......................................................................................... S
2.1 White rot decay .................................................................................. 5

2.2 Brown rot decay ................................................................................. 6

2.3 Soft rot decay ..................................................................................... 8

2.4 Classification of wood degradation by wood decay fungi .................... 8

2.5 Chemical properties of copper ............................................................ 9
MATERIALS AND METHODS .............................................................................. 11
3.1 Wood treatment ................................................................................. 11

3.1.1 Wood species and wood sizes ................................................. 11

3.1.2 Treating solutions .................................................................... 11

3.1.3 Wood treatment ...................................................................... 12

3.1.4 Sterilization of wood blocks .................................................... 12

3.2. Copper analysis ................................................................................... 13

3.2.] Analysis of elemental copper content in wood ......................... 13

3.2.2 Analysis ofCu (I) in wood ...................................................... 13

iv

3.3 Soil-block test of Cu-N and Cu-EA treated wood

after post-treatment steaming ............................................................... 14
3.4 Statistical analysis ................................................................................ 16
RESULTS AND DISCUSSION ............................................................................... 17
4.1 Wood appearance ................................................................................ 17
4.2 Copper retention in treated wood ......................................................... 17
4.3 Cu (1) analysis in wood ......................................................................... 17

4.3.1 Characterization of Cu (I)—2,2'-biquinoline
by UV/VI S spectrophotometer ................................................. 18

4.3.2 Quantification of Cu (I)-2,2'-biquinoline

complex in solution .................................................................. 19
4.3.3 Cu (1) and Cu (11) content in treated wood .............................. 19
4.4 Mass balance for Cu (I) and Cu (II) analysis ........................................ 21
4.5 Soil block test results ........................................................................... 21
4. 5.1 Cu -N treated SYP after Poria placenta exposure .................. 22
4. 5.2 Cu —N treated SYP after Gloeophyllum trabeum exposure ..... 22
4. 5.3 Cu -N treated SM after Poria placenta exposure ................... 22
4. 5.4 Cu -N treated SM after Gloeophyllum trabeum exposure ....... 23
4. 5.5 Cu —EA treated SYP after Poria placenta exposure ............... 23
4.5.6 Cu —EA treated SYP after Gloeophyllum trabeum exposure ...23
4. 5.7 Cu —EA treated SMafier Poria placenta exposure .................. 24
4.5.8 Cu -—EA treated SM after Gloeoplnzllum trabeum exposure ....24
4.5 The weight losses and the copper content in wood .............................. 24
CONCLUSIONS ...................................................................................................... 27

APPENDIX: Initial data of weight of wood blocks
for pressure treatment and soil block test ........................................... 63

REFERENCES ......................................................................................................... 8O

Table 1.

Table 2.

Table 3.

Table 4.
Table 5.
Table 6.

Table 7.

Table 8.

Table 9.

Table 10.

Table 11.

LIST OF TABLES

Oxidation states and stereochemistry of copper ........................................ 28
Cu (I) content in Cu-N treated wood

after various periods of post-treatment steaming ...................................... 29
Cu (1) content in Cu-EA treated wood

after various periods of post-treatment steaming ...................................... 30
Mass balance for Cu (I) and Cu (II) analysis .............................................. 31
Weight losses of SYP after laboratory soil block test ................................ 32
Weight losses of SM alter laboratory soil block test .................................. 33

Spearman rank order correlation for copper species
and average weight losses after soil block test .......................................... 34

Overall effect of post-treatment steaming on Cu-N treated SYP:
weight losses fi'om soil block test:
Statistical One Way ANOVA and Tukey ................................................. 35

Overall effect of post-treatment steaming on Cu-EA treated SYP:
weight losses from soil block test:
Statistical One Way ANOVA and Tukey ................................................. 36

Overall effect of post-treatment steaming on Cu-N treated SM:
weight losses from soil block test:
Statistical One Way ANOVA and Tukey ................................................. 37

Overall effect of post-treatment steaming on Cu-EA treated SM:
weight loss from soil block test:
Statistical One Way AN OVA and Tukey ................................................. 38

vii

Figure 1.
Figure 2.
Figure 3.
Figure 4.
Figure 5.

Figure 6.

Figure 7.

Figure 8.

Figure 9.

Figure 10.
Figure 11.
Figure 12.
Figure 13.
Figure 14.
Figure 15.
Figure 16.
Figure 17.

Figure 18.

LIST OF FIGURES

Building blocks of lignin ........................................................................ 39
Oxidation/reduction potential data of Cu (I) and Cu (11) ......................... 40
Copper retention in wood after pressure treatment with Cu-N ................ 41
Copper retention in wood afier pressure treatment with Cu-EA .............. 42
Cu (1) analysis using 2,2'-biquinoline ....................................................... 43

UV/VIS spectrum of (a) Cu20 and Cu-N with 2,2'-biquinoline
(b) Cu-N with 2,2'-biquinoline
(c) Cu20 with 2,2'-biquinoline ................................................................ 44

UVNIS spectrum of (a) Cu20 and Cu-EA with 2,2'-biquinoline
(b) Cu-EA with 2,2'-biquinoline

(c) Cu20 with 2,2'-biquinoline ................................................................ 45
Calibration curve of Cu (I) -2,2'-biquinoline ........................................... 46
Cu (1) content in Cu-N treated SYP after post-treatment steaming .......... 47

Cu (1) content in Cu-EA treated SYP after post-treatment steaming ....... 48
Cu (1) content in Cu-N treated SM after post-treatment steaming ........... 49

Cu (1) content in Cu-EA treated SM after post-treatment steaming ......... 50

Cu (11) in Cu-N treated SYP after post-treatment steaming ..................... 51
Cu (11) in Cu-N treated SM afier post-treatment steaming ...................... 52
Cu (11) in Cu-EA treated SYP after post-treatment steaming .................. 53
Cu (II) in Cu-EA treated SM after post-treatment steaming .................... 54
Average weight losses of Cu-N treated SYP to brown-rot fungi ............. 55
Average weight losses of Cu-N treated SM to brown-rot firngi ............... 56

viii

Figure 19.
Figure 20.

Figure 21.

Figure 22.

Figure 23.

Figure 22.

Average weight losses of Cu-EA treated SYP to brown-rot fungi ........... 57
Average weight losses of Cu-EA treated SM to brown-rot fiingi ............ 58
Average weight losses (%) after a soil block test

versus the Cu (1) content (%) in Cu-N treated SYP

(Cu retention: 1.88 mg/g) ...................................................................... 59
Average weight losses (%) after a soil block test

versus the Cu (1) content (%) in Cu-EA treated SYP

(Cu retention: 5.00 mg/g) ...................................................................... 60
Average weight losses (%) after a soil block test

versus the Cu (1) content (%) in Cu-N treated SM

(Cu retention: 2.50 mg/g) ...................................................................... 61
Average weight losses (%) after a soil block test

versus the Cu (1) content (%) in Cu-EA treated SM

(Cu retention: 4.44 mg/g) ...................................................................... 62

ix

INTRODUCTION

1.1 Background and proposed work

Wood bio-deterioration is generally caused by biological decay including fungi
stains, insect infestation, etc. The properties mostly afl'ected are the density and the
mechanical properties.

The wood-destroying microorganisms are subdivided into four main groups:
white rot, brown rot, soft rot, and mold/stain firngi (Eaton and Hale, 1993).

White and brown rot fiingi are predominant agents of firngal degradation of wood.
Their enzymatic capacities to depolymerize wood provide them with the potential to
colonize wood at all stages of decomposition. Brown-rot firngi are mostly common in
coniferous species, whereas white-rot firngi are frequent in angiosperms (Rayner and
Boddy, 1988). Therefore, brown rot and white rot are the most frequently used firngi in
study of wood protection.

Copper compounds have been used as fungicides over a century. Water-home
copper based preservatives such as ammoniacal copper arsenate (ACA), ammoniacal
copper zinc arsenate (ACZA), acid copper chromate (ACC), chromated copper arsenate
(CCA), copper bis (dimethyldithiocarbamate) (CDDC), ammoniacal copper citrate (CC),
ammonical copper quat (ACQ-B and ACQ-D), copper azole-type A (CBA-A), and oil-
borne copper based wood preservatives such as copper naphthenate, oxine copper
(copper-8-quinolinate) are listed in American wood-preservers’ association standards

(AWPA, 1999).

Copper naphthenate (Cu-N) was introduced as a wood preservative in 1900. Cu-
N is particularly efi'ective against cellulose-destroying fimgi and could afford long-lasting
protection (Eaton and Hale, 1993). Freshly treated Cu-N products have a green / blue
color and a strong odor that negatively impact their use (De Groot et al., 1988). To
reduce the green color, the strong odor for environmental reasons, and to obtain a clean
dried surface, Cu-N treated products are sometimes suggested to post-treatment steaming
at 240°F (115°C) as recommended in the footnote of the AWPA book of standards
(AWPA, 1996).

The reduction of Cu (11) to Cu (I) and the formation of cuprous oxide (Cu20) in
post-treatment steamed Cu-N treated southern pine were reported by Kamdem and Zhang
(1998). It is explained that the formation of Cu20 is promoted by the carboxylic acids
generated during the post-treatment steaming. These wood acids can protonate the
naphthenate ligand of Cu-N into napthenate acid and the formation of copper carboxylate
/ acetate / forrnate / glucuronate / hydroxide. Under some specific conditions, the new
form of copper may be transformed partially or totally into Cu20 (Kamdem and Zhang,
1998). The firngicidal activities of copper-based compounds such as copper naphthenate,
copper amine, copper-8-quinolinate may depend on the valence or the oxidation states of
the copper in the treating solutions and in the treated wood. Little information is
available in the literature about the toxicity of metach copper, cuprous and cupric
copper.

Copper amine system is one of the main ingredients of ACQ-D, CDDC, and

copper azole. Its relatively low mammalian toxicity and environmental risk may help its

promotion as the preservatives in the future. Cu20 can also be obtained fi'om a copper-
amine-treated wood after a post-treatment steaming, which is monitor in the lab by XRD.

This provides an interesting question about the bio-performance of wood
containing Cu20. Could the post-treatment steaming promote the formation of Cu (I)? Is
there any difference in the bio-perfonnance between cupric copper and cuprous copper as
wood preservatives?

Efforts have been invested in studying the mechanisms of copper toxicity to fungi
(Hill, 1977). Mocquot et al. (1996) reported that the activities of certain enzymes are
afi‘ected by the copper content in young maize. Gastaldi et a1. (1993) reported that the
cupric ion could inhibit the alcohol dehydrogenase 1 from Kluyveromyces marxianus. It
is hypothesized that copper acts as a cofactor for electron donor and acceptor for many
enzymatic reactions including oxidation, hydroxylation and dismutation. Excess of
copper is reported to cause damage through oxidizing proteins and lipids and enhancing
the toxic intra- and extra-cellular free radicals (Goyer and Cherian, 1995).

Superoxide radicals (02") and H202 are known to be produced in many biological
systems, but these are relatively harmless as they react with bio-molecules at low rates.
Simpson et al. (1988) suggested that in the presence of Cu (II) ion complexes, 02" and
H202 could lead to the formation of hydroxyl radical (OH), which could damage the
protein molecules in tissue. Similar reactions could take place in the presence of organic
hydroperoxides (ROOH), Cu (II) could be reduced to Cu (1) resulting in the formation of
peroxy (ROO’) radical and the more reactive alkoxy (RO') radical (Hunt et al., 1988).

Little information is available about the biochemistry of copper and proteins

under normal conditions (Hamer, 1986). The absorption / reaction sites of copper and

proteins are hypothesized to the carboxylic groups and the amino groups in the amino
acids of the proteins. Therefore, the copper-oxygen and copper-nitrogen interactions may
be determinant factors to the copper-fungi toxicity.

The objective of this work is to characterize Cu (1) and Cu ([1), quantify Cu (I) and
Cu (II) in treated wood and determine the biological performance of Cu (I) and Cu (II) in
treated wood by challenging with laboratory pure culture of white and brown rot decay

firngi.

LITERATURE REVIEW

It is important to understand the factors that afi‘ect the growth and development of
fungi in wood substrates. Like all living organisms, fungi have certain requirements for
growth and survival. The major growth needs of wood-inhabiting firngi are water,
oxygen from air, a favorable temperature and a good food source. Fungi can digest wood
to obtain energy and some compounds important for their metabolism (label and
Morrell, 1992). Decay fungi are clarified in three main groups: white rot, brown rot and
soft rot decay firngi. The control of oxygen and temperature to reduce the decay process
is a little difficult. Water can be limited by using dried wood. The hygroscopicity nature
of wood can not limit the absorption of water by wood used in eternal application. One
of the most effective ways of controlling decay is to put toxicants in the food source,

which is wood.

2.1 White rot decay

White-rot fungi are known to degrade lignin. The degraded wood is white and
soft. Most of the white-rot fungi prefer hardwoods. This may be due to the amount and
the type of lignin (Schultz and Nicholas, 1997). The content of lignin in hardwood is
about 18 to 25 % in dry weight and about 25 to 35 % in softwood (Haygreen and
Bowyer, 1996). The building blocks for hardwood and softwood are shown in Figure 1.
It is proposed that lignin in wood can act as antioxidant and disrupts various white-rot

free radical degradative mechanisms (Schultz and Nicholas, 1997). An attack by white-

rot fungi causes a decrease of strength properties and an increase of swelling (Fengel and
Wegener, 1983).

The most characteristic property of white-rot firngi is that they can produce
ligninases. Staining and growth test of the fungal mycelium gave evidence of the
presence of these enzymes (Paice and Jurasek, 1979; Schmidt and Liese, 1980). Lignin
peroxidase was found in T rametes versicolor and other white rot fungi. It plays a key
role in lignin degradation. Trametes versicolor requires H202 to oxidize and break the
bond between carbons in phenyl propane units in lignin (label and Morrell, 1992). From
the elemental analysis of isolated lignin after fungal attack, the amount of oxygen is
higher compared to that in the isolated lignin without fungal attack (Kirk, 1971; Kirk and
Chang, 1974). The increase of oxygen in the lignin molecule derives fi'om an oxidation
of lignin.

The hyphae of white rot fungi penetrate the wood tissue through the pit
membranes and through the cell walls by forming bore holes (Rayner and Boddy, 1988).
At the macroscopic level, this leads to a remarkable honeycomb-like pattern of decay
(Kirk, 1973). Species of white-rot fungi include T rametes versicolor, Plumerochaete
chrysoworium, Pleurotus ostreatus and Schizophyllum commune, etc. (Eaton and Hale,

1993).

2.2 Brown rot decay
Brown-rot decay results in a selective degradation of cellulose and hemicellulose.
Brown rot decay firngi can produce cellulase and hemicellulase, to digest or hydrolyze

cellulose and hemicellulose. It is reported that the enzymatic degradation of cellulose is

influenced by the presence of monosaccharides, which act as a starter-substance or aid in
the decomposition of the crystalline structure (Koenigs, 1974; Highley, 1976, 1977,
1980)

Chemical analysis of wood decayed by brown-rot firngi such as Poria monticola,
Lenzites trabea and Lentinus lepideus shows a significant reduction of the polysaccharide
content. The wood cellulose and polyoses undergo an extensive depolymerization before
any significant weight loss is noticed (Kirk and Highley, 1973). The reduction of the
mechanical properties at an earlier stage of decay is a good indication of brown rot decay.
Lignin is also subjected to a limited degradation. UV spectrophotometn’c studies of
brown rot decayed wood show a relative increase of the lignin content. This can be
explained by the relatively larger amount of cellulose and hemicellulose depolymerized
compared to that of lignin. X-ray difi‘raction study of cellulosic material treated with
cellulase shows a reduction of the crystallite width of cellulose, indicating that attack by
brown-rot firngi starts at the amorphous region of cellulose (Rayner and Boddy, 1988).

Brown-rot fungi are more frequent on softwood. The effect of lignin type and
quantity on the rate of brown rot decay is considerably less than that recorded with white
rot fungi (Schultz and Nicholas, 1997). Decayed wood becomes fiiable, ultimately
powdery, and cracks cubically. The mechanical strength of wood is reduced. The
hyphae of brown rot decay fungi grow mainly in the lumen of the wood cell walls
(Wilcox, 1973). Abnormal longitudinal shrinkage and deformation of the cell walls is
also observed (Fengel and Wegener, 1983).

Koenigs (1974) found that brown-rot fungi produce more H202 than the white-rot

fungi, and suggested that the initial attack on crystalline cellulose by brown-rot fungi be

via a non-enzyme H2O2/Fe2+ system, which generates hydroxyl radical. Species of
brown-rot fungi include Coniophora puteana, Setpula lacrymans, Antrodia vaillantii,

Gloeophyllum trabeum and Poria placenta, etc. (Eaton and Hale, 1993).

2.3 Soft rot decay

Soft-rot is found both in softwoods and hardwoods, and results in various rates of
reduction of the strength properties (F engel and Wegener, 1983). Soft-rot fungi degrade
cellulose and hemicellulose but do not appear to break down lignin appreciably (Rayner
and Boddy, 1988). Sofi-rot attack is typified by the formation of unique longitudinal
bore holes in the secondary cell wall (label and Morrell, 1992). The decay is
characterized macroscopically by surface softness. Soft-rot fungi can produce cellulases
and hernicellulases such as endo- and exo-l, 4-B-glucanases, B-glucosidases, which are
capable of degrading cellulose and hemicellulose in wood (Eriksson and Wood, 1985).
Species of soft-rot fungi include Chaetomium globosum, Lecythophora hofiinamii,

Monodictys putredinis and Humicola alopallonella, etc. (Eaton and Hale, 1993).

2.4 Classification of wood degradation by wood decay fungi
Liese (1970) presented a classification of decay types to wood tissue in decreasing
severity of cell-wall damage:
1. Simultaneous white-rot fungi attack all cell-wall constituents, essentially uniformly
during all decay stages;
2. Sequential white-rot fungi attack all cell-wall constituents; the initial attack is

selective for hemicellulose and lignin;

3. Brown-rot fimgi primarily attack the cell-wall carbohydrates, leaving a modified
lignin at the end of the decay process;

4. Soft-rot fungi preferentially attack cell-wall carbohydrates in the 82 layer of the
secondary cell wall, forming longitudinal cavities or eroding the wood cell wall fiorn

lumen surface in hardwoods or the $2 in conifers.

2.5 Chemical properties of copper

Copper is the first element of Group [B in the periodic table. The most abundant
oxidation states for copper are Cu (0), Cu (I) and Cu (11).

The relative stability of the Cu (1) and Cu (11) are indicated by the oxidation /
reduction potential data presented in Figure 2 (Cotton and Wilkinson, 1988). In aqueous
solution only low equilibrium concentrations of Cu (I) can exist compared to the
concentration of Cu (11) unless Cu (I) is complexed. For an example, CU(NH3)2+, etc.
The only copper compounds stable in water are the highly insoluble Cu2O, Cu2S and
CuCl, etc. The water instability of Cu (1) compounds is due partly to the greater lattice
and solvation energies and higher formation constants for Cu (H) compounds. Ionic Cu
(1) derivatives are readily to be oxidized to Cu (II), which is much more stable. Further
oxidation to Cu (III) is possible but more dimcult. There are only a few Cu (III)
complexes known (Cotton and Wilkinson, 1988).

Table 1 lists some copper complexes and their oxidation states (Cotton and

Wilkinson, 1988).

Cu (1) is a d’" ion and it is diarnagnetic. Cu (11) is a d9 ion and it is paramagnetic.
Cu (II) can be detected using electron paramagnetic resonance (EPR). No paramagnetic
signal is obtained from Cu (I).

The identification and quantification of the types and forms of copper is well
established for liquid samples. Speciations of copper can be done on liquid samples
using compleximetric titration, polarography and anodic stripping volametry (Zirinoode
and Kounaves, 1980).

X-ray photoelectron spectroscopy (XPS) is routinely used to determine the atomic
composition and the valence states present on the surface of solid sample. It provides
information such as the binding energy, the chemical shift of photoelectrons and Auger
electron, which is used to identify the correspondent atom and the oxidative states. It was
reported to detect the cupric and cuprous copper in Cu-N treated wood (Kamdem and
Zhang, 1999).

Cu (11) compounds have a specific color. This provides an opportunity to explore
spectrophotometry as a means to identify and quantify the presence of Cu (I) and Cu (II)
in liquid and solid. The color derives fi'om one of the following three rrrain possibilities:

charge-transfer transitions, d —> d transitions and intraligand-orbital transitions.

Most Cu (11) complexes have a blue or green color due to the d —9 d transition.
Cu (11) has a specific absorption band in the 600 to 900 nm regions (Cotton and
Wilkinson, 1988). Cu (O) is tough, soft, and ductile reddish metal. Little information is
obtained describing the color of Cu (1) complexes. This might be due to the instability of

Cu (1) complexes in aqueous solution.

10

MATERIALS AND METHODS

3.1 Wood preparation
3.1.1 Wood species and wood sizes

Defect-flee kiln dried sapwood boards of southern yellow pine (SYP) and sugar
maple (SM) were selected in the study. Cubes measuring 14 mm for SYP and 17 mm for
SM were prepared from these boards and dried in at 40°C for 24 hours. The moisture
content afier dried in oven was 4 :1: 1 percent. The densities of wood blocks were 32
pounds per cubic foot (pct) for SYP and 36 pcf for SM.

3.1.2 Treating solutions

Cu-N and Cu-EA solutions with the copper strength in treating solutions ranging
from 0.0, 0.25, 0.5 and 1.0 percent (w/w) were prepared for treatment.
3.1.2.1 Cu-Napthenate (Cu-N)

A stock solution of Cu-N containing 2 % elemental copper, received fiom ISK
Biosciences Corporation, was diluted with toluene from IT Baker Inc. Naphthenic acid
is distilled from crude oil and may contain C5 to C35 carboxylic acids (Dzidic et al.,
1988)
3.1.2.2 Cu-Ethanolamine (Cu-EA)

Cu-EA was made by mixing copper hydroxide and monoethanolarnine with
ethanolamine to copper molar ratio of 4:1 at room temperature in order for copper
complexed with ethanolamine completely. Copper hydroxide and ethanolamine were

both obtained from Aldrich Chemical Co.

11

3.1.3 Wood treatment

A modified full cell process was used to pressure treat wood blocks. The
treatment consists of two steps. During the first step, a cylinder measured 3 inches in
diameter and 6 inches in length was used for the pressure treatment with Cu-N / Cu-EA
solutions. An initial vacuum of about 84.6 kPa (25 inches of mercury) was pulled for 10
minutes prior to the injection of the treating solutions. A pressure level of 690 kPa (100
psi) was applied for 30 minutes. The extra solution was removed from the cylinder and
then a final vacuum applied for 30 minutes.

The second step consisted of post-treatment steaming. The treated blocks were
placed on a tray and air-dried at room temperature for 24 hours. Three hundred blocks of
Cu-N or Cu-EA treated wood were divided into 6 groups. Every fifty blocks were post-
treatrnent steamed at 240°F (115°C) in autoclave for 0, 20, 40, 60, 120 or 240 minutes,
respectively. After post-treatment steaming, wood samples were dried at 40°C until the
weight reached constant weight.

3.1.4 Sterilization of wood blocks

Wood samples were sterilized by ionizing radiation using Co-60 sources
according to AWPA standard B 10-91 (AWPA, 1999). The wood blocks were sealed in
polyethylene bags. Nitrogen was introduced in the bags to reduce the oxygen content and
limit the oxidization. The envelopes were subjected to radiation at a level of 2.0 to 2.5
Mrad. Half of these treated wood blocks were used in soil block tests and the other half

were used for analysis.

12

3.2 Copper analysis
3.2.1 Analysis of elemental copper content in wood

Analysis of elemental copper content in treated cubes was determined by atomic
absorption spectroscopy (AAS) using Atomic Absorption Spectrophotometer Model:
Perking Elmer 3110 according to AWPA Standard A1 1-93 (AWPA, 1999). The wood
blocks were ground into sawdust. About 0.2 g of wood sawdust was oven dried at 105°C
and weighted. Samples were acid digested with nitric acid and perchloric acid following
the AWPA Standard A7-93 (AWPA, 1999). The solutions after digestion were diluted to
around 10 ppm for AAS analysis. The working standard for AAS was at wavelength of
324.8 nm with hollow cathode lamp as light source and air-acetylene as flame.

3.2.2 Analysis of Cu (1) in wood
3.2.2.1 Characterization of Cu (I) by UV/VIS spectrophotometery

The 2,2'-biquinoline reagent was prepared at room temperature by dissolving
about 0.25 g of 2,2'-biquinoline in 250 ml of glacial acetic acid. The chemicals were both
obtained from Aldrich Chemical Co.

The spectrum for different copper species in 2,2'-biquinoline reagent were
collected by scanning respective solutions from 450 nm to 1000 nm using ultraviolet I
visible spectrophotometer. A Beckrnan DU640 B spectrophotometer with a 10—mm light
path silica cuvette fiom Sigma was used. All UV/VIS scans were performed at a speed
of 600 nm/min. Cu2O mixed with Cu—N or Cu2O mixed with Cu-EA were prepared by
mixing Cu2O with Cu-N or Cu-EA treating solutions in 2,2'-biquinoline reagent.

3.2.2.2 Quantification of Cu (1) -2,2'-biquinolinc complex in solution

13

The Cu (I)-2,2'-biquinoline complex was prepared by dissolving Cu2O in 2,2'-
biquinoline reagent (0.004 moi/l). The concentration of Cu (I) ranged fi'om 0 ppm to 100
ppm (0 to 0.0016 mon) was determined by AAS. The solutions were stored for 20
minutes before detecting the WMS absorption at 540 nm, which is the wavelength of
the maximum absorbance for Cu (I)-2,2'-biquinoline complex in glacial acetic acid.
3.2.2.3 Cu (I) content in treated wood

The wood was ground to pass through 40-mesh sieve and then dried at 40°C in
oven for 24 hours. The moisture content of wood powder was 4 i 1 percent.

About 25 ml of 2,2'-biquinoline reagent was used to extract about 0.1 g wood
powder by ultrasonic extraction for 5 minutes using ultrasonator Model: ULTRA sonik
57X from Cole-Farmer Instrument Co. with nitrogen protection against air oxidization.

After centrifugation, the supematant was used for UV/VIS analysis. The
absorption of the solution was measured at 540 nm (Felsenfeld, 1960).
3.2.2.4 Mass balance of Cu (1) and Cu ([1) analysis

After the ultrasonic extraction, the wood powder was washed with 3 ml of 2,2'-
biquinoline reagent for 4 to 5 times until the filter liquor turned colorless. The solutions
were collected for both Cu (1) analysis and AAS analysis for the elemental copper. The
wood powder was analyzed by AAS to determine the amount of copper that remained in
wood. The copper contents detected from these two steps were compared to the
elemental copper content in treated wood.

3.3 Soil-block test of Cu-N and Cu-EA treated wood after post-treatment

steaming

l4

Wood cubes treated by 0.5 % in elemental copper (w/w) Cu-EA treating solution
and 0.25 °/o in elemental copper (w/w) Cu-N treating solution were used in study the bio-
perforrnance of cupric and cuprous copper challenged by wood decay fimgi.

Two brown rot fungi, Poria placenta (Pp) (F r.) Cooke (Madison 698,ATCC
11538) and Gloeophyllum trabeum (Gt) (Pers ex. Fr.) Murr (Madison 617, ATCC
11539), two white rot fimgi, T rametes versicolor (Tv) (L. ex Fr.) (FP-lOl664-Sp, ATCC
42462) and Pleurotus ostreatus (Po) (Jacq. Ex Fr.) Kummer (ATCC 3223 7) were used to
evaluate the bio-performance of cupric and cuprous copper in wood cubes as described in
AWPA Standard E10-91 (AWPA, 1999) with some slight modification.

Aspen cut in 3 by 28 by 34 m (1/8 x 1‘/. x 13/. in.) blocks were used as feeder
strips. About 3.5 :i: 0.5 g of malt extract agar was poured on the surface of feeder strips
and the blocks were sterilized in an autoclave before introducing the wood cubes.

The malt extract agar was prepared by dissolving 7.5 g Difco Bacto malt extract
and 10.0 g and Difco Bacto-agar fiom Difco Laboratories in 500 ml distilled water by
stirring and heating below 80°C until it became transparent (Wang and label, 1990).

Boxes were inoculated with a monoculture of firngus and incubated until the
feeder strip was covered. Five replica for each sample group were introduced. The soil
boxes were kept in a growing room at the temperature of 76°F and relative humidity of 90
% during the test. After 16 weeks for brown-rot firngi exposure and 20 weeks for white-
rot fungi exposure, the cubes were removed from the culture boxes, scraped clean to
remove superficial mycelium and reconditioned in an oven at 40°C until the weight

stabilized. The percentage of weight loss after exposure to pure monoculture of test

15

fungus was used as the index of decay. Wood blocks were introduced to soil boxes with
no fungi to obtain the operational weight losses due to the process.

The average moisture content of a soil box was taken by comparing the weight of
the culture medium to the dried culture medium. The soil boxes were randomly selected
and the weight of dried culture medium was obtained by oven drying at 105°C until the

weight reached constant.

3.4 Statistical analysis

The correlation between the copper species (Cu (1) and Cu (11)) and the weight
losses obtained after a soil block test were evaluated by Spearman order correlation
analysis. One way AN OVA and Tukey analysis were used to evaluate the overall efi‘ect
of post-treatment steaming on the weight losses of Cu-N and Cu-EA treated wood. All
pairwise multiple comparison were evaluated within different duration periods of post-

treatment steamed wood to certain fungus exposure.

16

RESULTS AND DISCUSSION

4.1 Wood appearance

Cu-N treated wood has a green / blue color with a strong odor. Cu-EA treated
wood has a blue color with no noticeable odor. The color changed to brown after post-
treatment steaming and became darker with the increase of post-treatment steaming

duration period for both Cu-N and Cu-EA treated wood.

4.2 Copper retention in treated wood

The copper retentions were achieved in mg/g: 1.88, 3.44 and 5.31 mg/g for Cu-N
treated SYP, 3.13, 5.00 and 10.31 mg/g for Cu-EA treated SYP, 1.39, 2.50 and 4.17 myg
for Cu-N treated SM and 2.50, 4.44 and 7.50 for Cu-EA treated SM in elemental copper
(Tables 2 and 3). The copper retention after pressure treatment for SYP was higher than
that of SM for the same treatment. The copper retentions in wood increase with the
increase of the concentration of copper in the treating solutions as shown in Figures 3 and

4.

4.3 Cu (1) analysis in wood

Little research has been reported to estimate the Cu (1) content in treated wood
quantitatively. Kamdem and Zhang (1998) used X-ray difi‘raction photoelectron
spectroscopy (XRD) and (1999) X-ray photoelectron spectroscopy (XPS) successively to

detect the Cu (1) in wood. Klotz and Klotz (1955) used a colorimetric reagent namely

17

 

2,2'-biquinoline in glacial acetic acid to determining the concentration of cuprous copper
in Busycon hemocyanin. This method has been applied to determine the concentration of
Cu (1) in protein chemistry, notably for the tyrosinase enzyme (F elsenfeld, 1960). The
reaction scheme between Cu (1) and 2,2'-biquinoline is presented in Figure 5.

4.3.1 Characterization of Cu (1) - 2,2' - biquinoline by UVNIS spectrophotometer

Cu (1) reacts with 2,2'-biquinoline in glacial acetic acid matrix yielding a pink
color solution. Such color is not noticeable with cupric copper. The 2,2'-biquinoline
reagent was colorless with pH value ranged around 2 :1: 0.5. It takes about 20 minutes for
the pink color to develop at room temperature. The pink color results from the
solubilization of cuprous oxide and the formation of a Cu (1) complex with 2,2'-
biquinoline (Kertesz, 1957).

Figure 6 presents the UV/VIS spectrum of Cu2O, Cu-N, Cu20 mixed with Cu-N
in 2,2'-biquinoline reagent. An absorption band with Amax at 540nm was observed fi'om
Cu2O dissolved in 2,2'-biquinoline reagent in spectra (c). The band was attributed to the
new complex formed between Cu2O and 2,2'-biquinoline. This maximum absorbance at
540 nm remained stable in presence of Cu-N as shown in spectra (3). There is no
absorbance from Cu-N dissolved in 2,2'-biquinoline reagent at 540 nm in spectra (b).

Figure 7 presents the UVNIS spectrum of Cu2O, Cu-EA, Cu2O mixed with Cu-
EA in 2,2'-biquinoline reagent. The maximum absorbance for Cu (1)- 2,2'-biquinoline
complex at 540 nm remained stable in presence of Cu-EA as shown in spectra (a). There

is no absorbance from Cu-EA dissolved in 2,2'-biquinoline reagent at 540 nm in spectra

(b).

18

The absorbance band for Cu-N and Cu-EA in 2,2'-biquinoline reagent were
broadened when the concentrations of Cu-N and Cu-EA were decreased. These
attributed to that Cu-N and Cu-EA were destroyed in acetic acid and Cu (11) - acetate was
formed in acetic acid matrix (Zyskowski and Kamdem, 1999).

These results agreed with the conclusion from Klotz and Klotz (1955) that 2,2'-
biquinoline react with cuprous copper to form a stable complex. Therefore, 2,2'-
biquinoline was used to complex Cu (1) and the complex was quantified as the cuprous
copper.

4.3.2 Quantification of Cu (I) -2,2'-biquinoline complex in solution

A calibration curve was constructed for cuprous concentration and the Cu (I)-2,2'-
biquinoline complex in solution. Known amount of cuprous oxide was diluted with 2,2'-
biquinoline reagent. A linear regression of Y = (X-0.681)/6.885 with R2 value of 0.991
was obtained between the copper concentration determined by AAS and the UVNIS
absorbance between 4 and 10 ppm of elemental copper (Figure 8). X is the concentration
of cuprous ion in ppm and Y is the UVNIS absorbance at 540 nm. This regression line
assumed that all Cu (1) in the solution was complexed with 2,2'-biquinoline and the
intensity of absorbance was proportional to the amount of complex formed.

4.3.3 Cu (1) and Cu (II) content in treated Wood

About 7.9 °/o of the 1.88 mg/g elemental copper in Cu-N treated SYP was
determined to be in the form of Cu (1). The content of Cu (1) increased to 37.0 % after 20
minutes of post-treatment steaming. Cu (1) content increased to 53.2 % after 60 minutes.

The Cu (1) content was 55.6 % after 120 minutes and 51.9 % after 240 minutes (Table 2).

l9

The Cu (1) content increased with the increasing of the post-treatment steaming
duration period. The formation of Cu (1) increased sharply during the first 60 minutes of
post-treatment steaming and reached a plateau after 60 nrinutes. This confirms the XRD
and XPS finding reported by Kamdem and Zhang (1998, 1999). The equations
presenting the Cu (1) content related to the post-treatment steaming time are listed in
Figures 9, 10, 11 and 12. The equations were obtained fi'om the every mean value of
three replicas. Y represents the Cu (1) content (%) in form of elemental copper in wood,
and X represents the post-treatment duration periods in minutes. Cuprous copper was
detected from Cu-N and Cu-EA treated wood without post-treatment steaming as well.
Nearly 10 :1: 5 % ofCu (1) in terms ofelemental copper was detected. The Cu (1) contents
in untreated wood were 0.1% for SYP and 0.1% for SM and less than 0.1% in treating
solutions. This suggests that the Cu (1) detected came from the vacuum pressure
treatment or drying in oven at 40°C. The equations reflected the Cu (1) detected with 0
minute of post-treatment steaming.

There was a slight decrease of Cu (1) content observed after 240 minutes of post-
treatrnent steaming compared to the Cu (1) content after 120 minutes of post-treatment
steaming in most specimens. The reason of this decrease was not clear. But it can be
theorized that solid deposit of copper in the surface was lost during the sample
preparation. Similar trends were observed for both Cu-N and Cu-EA treated wood with
difi’erent copper retentions.

Figures 13, 14, 15 and 16 present the initial Cu (11) content and the Cu (11) content

after post-treatment steaming. The Cu (11) contents in both Cu-N and Cu-EA treated SYP

20

and SM after post-treatment steaming increase with the increase of initial Cu (11) contents

in wood, and decrease with the increase of post-treatment steaming time.

4.4 Mass balance for Cu (1) and Cu (II) analysis

The first column of Table 4 contains the value of Cu (1) determined by UV/VIS
for several specimens. The second column represents the data of the AAS analysis of
elemental copper content in ultrasonic extracts. The third column represents the data of
elemental copper remained in solid wood sawdust after the extraction. The forth column
is the copper content in specimens determined by AAS afier vacuum pressure treatment
with no extrication. Data obtained from the AAS analysis of elemental copper content in
ultrasonic extracts and in wood sawdust after ultrasonic extraction were compared to the
elemental copper content in wood with no extraction. About 96 to 106 °/o of copper was
detected. About 64 °/o of copper was extracted after the ultrasonic extraction by
comparing the elemental copper content in extracts and the elemental copper content in
wood with no extraction. About 79 % of copper in extracts was determined as Cu (1) by
comparing the UV/VIS analysis of Cu (1) content in extracts and the AAS analysis of
elemental copper content in extracts. This suggests that some Cu (11) was extracted by

2,2'- biquinoline reagent during the ultrasonic extraction.

4.5 Soil block test

The moisture content in the soil substrate in boxes was 130 i 5 %.

21

The average weight losses in percentage after fungi exposure and the descriptive
statistics analysis for the standard deviation are listed in Tables 5 and 6 and Figures 17 to
20.

One way ANOVA analysis and Tukey test listed in Tables 8 to 11 present the
overall effect of post-treatment steaming on the weight losses of copper treated wood.
4.5.1 Cu-N treated SYP after Poria placenta exposure

The average weight loss was 1.1 °/o for Cu-N treated SYP with 0 minute of post-
treatrnent steaming after Poria placenta exposure. The weight loss increased to 24.2 %
after 120 minutes of post-treatment steaming. The weight loss for untreated control was
56.9 %. There was a significant difference observed between wood samples groups of 0
minutes and 120 minutes of post-treatment steaming; and 0 minutes and 240 minutes; and
20 minutes and 240 minutes (Table 8 and Figure 17). No significant difference was
observed among other sample groups.

4.5.2 Cu-N treated SYP after Gloeophyllunr trabeum exposure

The average weight loss was 2.4 % for Cu—N treated SYP with 0 minute of post-
treatrnent steaming after Gloeophyllum trabeum exposure. The weight loss increased to
9.0 % after 60 minutes of post-treatment steaming. The weight loss for untreated control
was 40.0 %. There was a significant difference observed between wood samples groups
of 40 minutes and 120 minutes of post-treatment steaming (Table 8 and Figure 17). No
significant difference was observed among other sample groups.

4.5.3 Cu-N treated SM after Poria placenta exposure
The average weight loss was 14.2 % for Cu-N treated SM with 0 minute of post-

treatment steaming after Poria placenta exposure. The weight loss increased to 35.4 %

22

after 40 minutes of post-treatment steaming. The weight loss for untreated control was
58.2 %. There was a significant difi‘erence observed between wood samples groups of 0
minutes and 40 minutes of post-treatment steaming; and 0 minutes and 60 minutes; and 0
minutes and 120 minutes; and 0 minutes and 240 minutes; and 20 minutes and 40
minutes; and 20 rrrinutes and 60 minutes; and 20 minutes and 120 minutes; and 20
minutes and 240 minutes (Table 10 and Figure 18). No significant difference was
observed among other sample groups.
4.5.4 Cu—N treated SM after Gloeoplryllunr trabeurn exposure

The average weight loss was 0.4 % for Cu-N treated SM with 0 minute of post—
treatrnent steaming after Gloeophyllum trabeum exposure. The weight loss increased to
4.6 % after 60 minutes of post-treatment steaming. The weight loss for untreated control
was 66.3 %. There was a significant difference observed between wood samples groups
of 0 minutes and 60 minutes of post-treatment steaming; and 0 minutes and 120 minutes;
and 20 minutes and 60 minutes; and 20 minutes and 120 minutes (Table 10 and Figure
18). No significant difference was observed among other sample groups.
4.5.5 Cu-EA treated SYP after Porr’a placenta exposure

The average weight loss was 32.1 % for Cu-N treated SM with 0 nrinute of post-
treatrnent steaming after Poria placenta exposure. The weight loss increased to 56.4 %
after 20 minutes of post-treatment steaming. The weight loss for untreated control was
56.9 %. There was no significant difference observed among wood samples groups
(Table 9 and Figure 19).

4.5.6 Cu-EA treated SYP after Gloeophyllum trabeum exposure

23

The average weight loss was 4.8 % for Cu—N treated SM with 0 minute of post-
treatrnent steaming after Gloeophyllum trabeum exposure. The weight loss increased to
9.0 % after 60 minutes of post-treatment steaming. The weight loss for untreated control
was 40.0 %. There was a significant difference observed between wood samples groups
of 0 minutes and 60 minutes of post-treatment steaming; and 0 minutes and 120 minutes;
and 0 minutes and 240 minutes; and 20 minutes and 60 minutes; and 20 minutes and 120
minutes; and 20 minutes and 240 minutes; and 40 minutes and 60 minutes; and 40
minutes and 120 minutes; and 40 minutes and 240 minutes (Table 9 and Figure 19). No
significant difi‘erence was observed among other sample groups.

4.5.7 Cu—EA treated SM after Porr'a placenta exposure

The average weight loss was 50.2 % for Cu—EA treated SM with 0 minute of post-
treatment steaming afier Poria placenta exposure. There is a slight increase of weight
losses observed with the increase of post-treatment steaming time. The weight loss for
untreated control was 58.2 %. There was no significant difference observed among wood
samples groups (Table 11 and Figure 20).

4.5.8 Cu-EA treated SM after Gloeophyllunr traberun exposure

The average weight loss was 0.7 % for Cu-EA treated SM with 0 minute of post-
treatment steaming after Gloeophyllum trabeum exposure. There is a slight increase of
weight losses observed with the increase of post-treatment steaming time. The weight
loss for untreated control was 66.3 %. There was no significant difference observed

among wood sample groups (Table 11 and Figure 20).

4.6 The weight losses and the copper content in wood

24

Figures 21 to 24 suggest that the average weight losses of Cu-N treated SYP and
SM increased with the percentage of Cu (1) in wood exposed to Porr'a placenta. The
relation between the weight losses and the Cu (1) content is not clear in Cu-EA treated
wood after Poria placenta exposure. There was a slight variation but no significant
difi‘erence in weight losses observed in most wood blocks exposed to Gloeophyllwn
trabeum, T rametes versicolor and Pleurotus ostreatus. The weight loss is relatively
lower for samples exposed to Gleophyllum trabeum, Trarnetes versicolor and Pleurotus
ostreatus compared to the specimens exposed to Porr'a Placenta. The increment of
weight losses fiom 0 minute of post-treatment steaming to 240 minutes was no more than
10 % compared to the increment of 25 i 5 % for the specimens exposed to Poria
Placenta. This is attributed to the good performance of copper-based wood preservatives
to most wood decay fungi (label and Morrell, 1992). However, Poria placenta is a
copper tolerant fungus (Sutter et al., 1983). The copper tolerance of Poria placenta is
reported in both laboratory experiments and in service situation (Schimidt and Liese,
1996; Da Costan and Kerruish, 1964; Williams and Fox, 1994; Collett, 1992; Thornton
and Tighe, 1987). This explained the relatively high weight losses obtained fiom both
Cu-N and Cu-EA treated wood exposed to Poria placenta and the relatively lower weight
loss exposed to Gloeophyllum trabeum, T rametes versicolor and Pleurotus ostreatus in
this test.

Statistic analysis using Spearman rank order correlation presented the correlation
coefiicient for the Cu (1) or Cu (11) contents in wood and the weight losses after soil block
test as shown in Table 7. A high correlation coefficient (more than 0.8) with low P value

(less than 0.05) were observed in Cu-N treated SYP, Cu-N treated SM and Cu—EA treated

25

SM afier Poria placenta exposure. The weight losses increased with the increase of Cu
(1) content and decreased with the increase of Cu (11) content in wood samples.

No significant correlation was observed between the copper species and the
weight losses for most specimens after Gloeophyllum trabeum, T rametes versicolor and
Pleurotus ostreatus exposure.

Marten and Leach (1994) reported that the toxicity of Cu20 toward Pythium
debaryanum depends on its solubility in difi‘erent conditions. Cu2O is less toxic because
it is less soluble in certain solution, which leads to less susceptible to interact with the
wood decay fungi such as the amino acid functional groups. Comparing the data
obtained from soil block test to the data obtained from copper analysis, we suggest that
the toxicity of Cu-N treated SYP and SM, and Cu-EA treated SM towards wood brown
rot decay fungi Poria placenta is attributed to the Cu (H) in wood in this post-steaming

treatment.

26

CONCLUSIONS

Cuprous copper content in presence of Cu (H) in treated wood could be
determined by the 2,2'-biquinoline method for Cu-N and Cu-EA treated SYP and SM
after post-treatment steaming.

Based on the analysis of Cu (1) and Cu (H) content in treated wood and the data of
soil block tests, Cu (H) can be converted to Cu (1) by using post-treatment steaming. The
conversion ratio increases with the increase of post-treatment steaming duration period in
the first 60 minutes for Cu-N and Cu-EA treated SYP cubes measuring 14 mm and SM
cubes measuring 17 mm of copper retention ranged from 1.88 mg/g to 10.31 mg/g.

The weight losses of Cu-N and Cu-EA treated wood increase with the increasing
of post-treatment steaming duration period in Poria placenta. The weight losses increase
with the decrease of Cu (H) content, while the total copper retention remains the same.

In conclusion, Cu2O in Cu-N treated SYP and SM and Cu-EA treated SM to

Poria placenta is less toxic than Cu (H) in post-steaming treated wood.

27

Table l. Oxidation States and Stereochemistry of Copper

 

 

 

   

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Oxidation Coordination Geometry
state number
2 Linear Cu20, CuClz'
3 Planar KlCu(CN)2] I
4' Tetrahedral CuI, (Cu(CN)4l"
4 Distorted planar CuL° I
I 5 sp (CuLCOl‘ I
J I 6 Octahedral eP Re Cu
Cu (H), d9 3 Trigonal planar Cu2(u-Br)2Br
4"” Tetrahedral (N -Isopropylsalicylaldininato)2Cu
(distorted) Cs,(CuCl,)
! 4"” Square CuO, (N11,),(CuCll4
6"” Distorted CuCl2
octahedral I
5 tbp (CuClslz’ I
5 sp lCu(NH;,),l2+ ‘ I
6 Octahedral K,Pb(Cu(NOz)6]
7 Pentagonal lCu(I-IZO)2(dps)l2+ °
bipyramidal
8 Distorted CaICu(co,Me),1' 611,0
dodecahedron
Cu (1H), d' 4 Square KCuO2 I
6 Octahedral KiCuFi I

 

' Most common states.

" These three cases are often not sharply distinguished.

° L= a macrocyclic N4 anionic ligand.

‘ In KCU(NH3)s (PR);

° dps= 2,6—diacetylpyridine bis(semicarbazone)

28

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Table 7. Spearman rank order correlation for copper species and average weight losses

 

    
   
 

 

 

 

 

 

 

 

 

 

 

 

 

after soil block test
WL (Pp) I WL (Gt) J WL (Po WL Tv
Co 8 'es ’
pp“ pea Cu-N treated SYP
Cu (1) content ‘0.829 0.486 0.714 0.829
"0.0583 0.356 0.136 0.0583
°6 6 6 6
Cu (11) content -0.829 -0.486 -0714 -0.829
0.0583 0.356 0.136 0.0583
6 6 6 6
Cu-EA treated SYP
Cu (I) content 0.257 0.714 0.429 0.771
0.658 0.136 0.419 0.103
6 6 6 6
Cu (II) content -0257 -0714 -0429 -0771
0.658 0.136 0.419 0.103

 

    

   

 

 
 

Cu-N treated SM

   

 

 

 

 

 

 

 

 

 

 

 

. --—~ —-———— - --——— —-+ L —

Cu (1) content 1.000 0.600 0.265 0.706
0.00278 0.242 0.564 0.136
6 6 6 6

Cu (II) content -1000 -0.600 -0.265 -0.706
0.00278 0.242 0.564 0.136
6 6 _ %_ 6

I Cu-EA treated SM

Cu (I) content 0.943 0.257 -0.232 -0.232
0.0167 0.658 0.658 0.658
6 6 6 6

Cu (11) content 0.943 0.257 -0.232 -0.232
0.0167 0.658 0.658 0.658
6 6 6 6

 

 

 

 

 

 

 

‘zCorrelation coefficient; I’: P value; ‘: Number of samples

34

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Cu+ + e = Cu E° = 0.52v
Cu2++ c = Cu+ 15" = 0.153v
Cu +Cu2*= 2Cu+ °=-o.37v

K= [Cu’j / [Cu+ )2 = ~1o‘S

Figure 2. Oxidation/reduction potential data of Cu (1) and Cu (II)
(Cotton and Wilkinson, 1988)

40

Copper retention in wood (mg/g)

 

 

 

 

 

5 5
4 .5
3 ...
2 -l
O... --o-- SM
1 T I 1 I
0.2 0.4 0.6 0.8 1.0 1.2

Copper content in treating solution (%)

Figure 3. Copper retention in wood after pressure treatment with Cu-N

41

Copper retention in wood (mg/ g)

12

 

 

 

 

 

10-
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65
4-
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Copper content in treating solution (%)

Figure 4. Copper retention in wood after pressure treatment with Cu-EA

42

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Absorbance

(I)

(b)

(c)

 

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450 nm

IOOOnm

Figure 6. UVNIS spectrum of (a) Cu20 and Cu-N with 2,2'-biquinoline (b) Cu-N

with 2,2'-biquinoline (c) Cu20 with 2, 2'-biquinoline

 

 

 

 

540 nm
(8)
(b)

§ *—
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Q
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(C)

500 nm 1000 nm

Figure 7. UVNIS spectrum of (a) Cu20 and Cu-EA with 2,2'-biquinoline
(b) Cu-EA with 2,2'-biquinoline (c) Cu20 with 2,2'-biquinoline

UV/VISabsat540nm

 

 

 

 

 

Figure 8. Calibration curve of Cu (1) -2,2'-biquinoline

1O

Cu (1) content in form of elemental copper (%)

 

20..

10~

 

 

 

 

 

O 50 1 00 1 50 200 250 300

Duration of post-treatment steaming (min)

+ Cu: 1.88 mg/g Y= 7.8+50.3X/(12.3+X), R’=o.9s
—0— Cu: 3.44 mg/g Y=10.9+40.2X/(26.8+X), 112:0.95
+ Cu: 5.31 mg/g Y=8.2+36.9X/(37.5+X), R’=o.85

Figure 9. Cu (1) content in Cu-N treated SYP after post-treatment steaming

47

Cu (1) content in form of elemental copper (%)

 

 

 

30“ I4

20~

-.

1o~ /

 

 

 

0 T T I T T I

O 50 100 1 50 200 250 300

Duration of post-treatment steaming (min)

+ Cu: 3.13 mg/g Y=9.0+19.4X/(14.3+X), R’=o.99
—0— Cu: 5.00mg/g Y=5.4+S3.9X/(22.2+X), R’=o.88
—v— Cu: 10.31 mg/g Y=4.3+33.5X/(85.1+X), R’=o.97

Figure 10. Cu (I) content in Cu-EA treated SYP afier post-treatment steaming

48

Cu (1) content in form of elemental copper (%)

 

 

 

 

 

o I I I I I T

O 50 1 00 1 50 200 250 300

Duration of post-treatment steaming (min)

+ Cu: 1.39 mg/g Y=7.5+50.2X/(8.2+X), R2=0.97
—0— Cu: 2.50 mg/g Y=14.9+54.3X/(35.5+X),R2=0.99
+ Cu: 4.17 mg/g Y=9.2+35.7X/(34.4+X), R’=0.94

Figure 11. Cu (I) content in Cu-N treated SM afier post-treatment steaming

49

Cu (I) content in form of elemental copper (%)

 

40q

20-

10~

 

 

 

 

 

I I T I I T

0 50 100 1 50 200 250 300

Post-treatment steaming time (min)

+ Cu: 2.50 mg/g Y=14.7+20.3X/(9.6+X), R2=0.78
—0— Cu: 4.44 mg/g Y=S.6+42.9X/(17.2+X), R2=0.97
—v— Cu: 7.50 mg/g Y=6.8+22.2X/(35.3+X), R’=0.81

Figure 12. Cu (1) content in Cu-BA treated SM afier post-treatment steaming

50

Cu (11) after post-treatment steaming (mg/g)

 

 

 

 

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0 min of post-treatment steaming
20 min of post-treatment steaming
40 min of post-treatment steaming
60 min of post-treatment steaming
120 min of post-treatment steaming
240 min of post-treatment steaming

DIG‘O.

Figure 13. Cu (11) in Cu-N treated SYP afier post-treatment steaming

51

Cu (11) after post-treatment steaming (mg/g)

 

 

 

 

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20 min of post-treatment steaming
40 min of post-treatment steaming
60 min of post-treatment steaming
120 min of post-treatment steaming
240 min of post-treatment steaming

DIQ‘O.

Figure 14. Cu (11) in Cu-N treated SM afier post-treatment steaming

52

Cu (II) after post-treament steaming (mg/g)

 

12'

105

 

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Cu (11) initial (mg/g)

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Cu(Il) 0 min of post-treatment steaming
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Cu (11) 40 min of post-treatment steaming
Cu (II) 60 min of post-treaming steaming
Cu (II) 120 min of post-treatment steaming
Cu (11) 240min of post-treatment steaming

DIG‘O.

Figure 15. Cu (H) in Cu-EA treated SYP afier post-treatment steaming

53

Cu (H) after post-treatment steaming (mg/g)

 

 

 

 

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v Cu (H) 40 min of post-treatment steaming
v Cu (11) 60 min of post-treatment steaming

I Cu (II) 120 min of post-treatment steaming
Cu (II) 240 min of post-treatment steaming

E]

Figure 16. Cu (11) in Cu-EA treated SM after post-treatment steaming

54

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35

 

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0 1O 20 30 40 50 6O 70

Cu (1) content in percentage of elemental copper

Figure 21. Average weight losses (%) after a soil block test versus the Cu (1)
content (%) in Cu-N treated SYP (Cu retention: 1.88 mg/g)

59

Average weight losses (%) after a soil block test

60

 

 

 

 

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Cu (1) content in percentage of elemental copper
0 Pp
0 Gt

Figure 22. Average weight losses (%) after a soil block test versus the Cu (I)
content (%) in Cu-EA treated SYP (Cu retention: 5.00 mg/g)

Average weight losses (%) after a soil blok test

40

 

 

 

 

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Figure 23. Average weight losses (%) after a soil block test versus the Cu (1)
content (%) in Cu-N treated SM (Cu retention: 2.50 mg/g)

61

Average weight losses (%) after a soil block test

70

 

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Cu (I) content in percentage of elemental copper

Pp
Gt

0
0

Figure 24. Average weight losses (%) after a soil block test versus the Cu (I)
content (%) in Cu-EA treated SM (Cu retention: 4.44 mg/g)

62

APPENDIX

63

 

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83

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