U.S. patent application number 10/476382 was filed with the patent office on 2004-10-07 for method for isolating phenolic substances or javabiones from wood comprising knotwood.
Invention is credited to Eckerman, Christer, Hemming, Jarl, Holmbom, Bjarne, Reunanen, Markku, Sundberg, Kenneth, Willfor, Stefan.
Application Number | 20040199032 10/476382 |
Document ID | / |
Family ID | 8561351 |
Filed Date | 2004-10-07 |
United States Patent
Application |
20040199032 |
Kind Code |
A1 |
Holmbom, Bjarne ; et
al. |
October 7, 2004 |
Method for isolating phenolic substances or javabiones from wood
comprising knotwood
Abstract
The present invention relates to a method for isolating of
phenolic substances or juvabiones from wood comprising knotwood,
said method comprising the steps of a) extracting i) the over-sized
chip fraction obtained by screening chipped wood, or ii) a
knot-rich sub-fraction obtained from said over-sized chip fraction,
or iii) knotwood obtained as residue in finishing of mechanical
wood products with a polar solvent, and b) recovering the
extract.
Inventors: |
Holmbom, Bjarne; (Abo,
FI) ; Eckerman, Christer; (Abo, FI) ; Hemming,
Jarl; (Abo, FI) ; Reunanen, Markku; (Abo,
FI) ; Sundberg, Kenneth; (Abo, FI) ; Willfor,
Stefan; (Abo, FI) |
Correspondence
Address: |
James C Lydon
Suite 100
100 Daingerfield Road
Alexandria
VA
22314
US
|
Family ID: |
8561351 |
Appl. No.: |
10/476382 |
Filed: |
October 30, 2003 |
PCT Filed: |
May 16, 2002 |
PCT NO: |
PCT/FI02/00418 |
Current U.S.
Class: |
585/242 |
Current CPC
Class: |
C07G 17/00 20130101 |
Class at
Publication: |
585/242 |
International
Class: |
C10G 001/00 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 6, 2001 |
FI |
20011194 |
Claims
1-21. (canceled)
22. A method for isolating phenolic substances or juvabiones from
wood comprising knotwood, said method comprising the steps of a)
extracting i) the over-sized chip fraction obtained by screening
chipped wood, or ii) a knot-rich sub-fraction obtained from said
over-sized chip fraction, or iii) knotwood obtained as residue in
finishing of mechanical wood products with a polar solvent, and b)
recovering the extract.
23. The method of claim 22, wherein the over-sized chip fraction is
separated in a knot-rich fraction and a knot-poor fraction, and the
knot-rich fraction is extracted.
24. The method of claim 23, wherein the over-sized chip fraction is
refined before the separation.
25. The method of claim 22, wherein the polar solvent is either a
single polar agent, or a mixture of two or more polar agents, said
polar agent or agents having a dielectric constant that is greater
than 3, determined at 25 Celsius degrees.
26. The method of claim 22, wherein the extract is further purified
for releasing the desired phenolic substance or juvabione.
27. The method of claim 22, wherein a) the phenolic substance is i)
a lignan, which is hydroxymatairesinol, allohydroxy-matairesinol,
matairesinol, conidendrin, pinoresinol, oxomatairesinol,
lariciresinol, liovil, isolariciresinol, secoisolariciresinol,
picearesinol, conidendric acid or nortrachelogenin, or ii) a
flavonoid, which is pinocembrin, dihydrokaempferol, pinobanksin,
naringenin, catechin, 2,4,6-trihydroxychalcone, aromadendrin or
taxifolin, or iii) a stilbene, which is pinosylvin, pinosylvin
monomethyl ether, dihydropinosylvin, methylpinosylvin,
methyldihydropinosylvin or reservatrol, or b) the juvabione is
epijuvabione, dehydrojuvabione, dihydroepijuvabione or epijuvabione
acid.
28. The method of claim 22, wherein the wood comprises coniferous
wood.
29. The method of claim 28, wherein the wood is spruce wood of the
genus Picea.
30. The method of claim 29, wherein the isolated substance is a
lignan which is hydroxymatairesinol, secoisolariciresinol or
liovil.
31. The method of claim 28, wherein the wood is fir wood of the
genus Abies.
32. The method of claim 31, wherein the isolated substance is a
lignan which is secoisolariciresinol, lariciresinol or
isolariciresinol.
33. The method of claim 32, wherein the isolated substance is
epijuvabione or dehydrojuvabione.
34. The method of claim 28, wherein the wood is a Larix genus.
35. The method of claim 34, wherein the isolated substance is a
lignan which is secoisolariciresinol, lariciresinol or
isolariciresinol; or a flavonoid which is catechin, taxifolin or
dihydrokaempferol.
36. The method of claim 28, wherein the wood is pine wood of the
genus Pinus.
37. The method of claim 36, wherein the isolated substance is a
flavoinoid, a lignan, a stilbene or a juvabione.
38. The method of claim 37, wherein the flavoinoid is pinocembrin,
dihydrokaempferol, pinobanksin, naringenin, catechin,
2,4,6-trihydroxychalcone, aromadendrin or taxifolin, the lignan is
lariciresinol, isolariciresinol, secoisolariciresinol,
matairesinol, liovil or nortrachelogenin (NTG), the stilbene is
pinosylvin, dihydropinosylvin, pinosylvin monomethyl ether,
methylpinosylvin, methyldihydropinosylvin, or reservatrol, or the
juvabione is epijuvabione acid.
39. The method of claim 28, wherein the wood is a Pseudotsuga
genus.
40. The method of claim 39, wherein the isolated substance is a
lignan, which is lariciresinol, isolariciresinol or
secoisolariciresinol, or ii) a juvabione such as
dihydroepijuvabione.
41. The method of claim 22, wherein the wood is hardwood and the
isolated substance is a flavonoid or a stilbene.
42. The method of claim 41, wherein the hardwood is aspen and the
isolated and the isolated flavonoid is naringenin or
dihydrokaemferol.
Description
FIELD OF THE INVENTION
[0001] This invention relates to a method for isolating chemical
substances, i.e. phenolic substances or juvabiones from wood
comprising knotwood.
BACKGROUND OF THE INVENTION
[0002] The publications and other materials used herein to
illuminate the background of the invention, and in particular,
cases to provide additional details respecting the practice, are
incorporated by reference.
[0003] It is known that vegetabilic sources, including wood,
contain phenolic substances which are more or less hydrophilic. The
phenolic substances derived from wood are flavonoids, lignans and
stilbenes. In spruce wood the dominant group is lignans while
flavonoids and stilbenes are the dominant phenolic substances in
pine wood and hardwoods. Freudenberg and Knof (1) identified
already in 1957 many different lignans in Norway spruce, including
e.g. hydroxymatairesinol, matairesinol, conidendrin, pinoresinol,
oxomatairesinol, lariciresinol, allohydroxymatairesinol and liovil.
In addition to these lignans, isolariciresinol,
secoisolariciresinol, picearesinol and conidendric acid have been
isolated from spruce (2).
[0004] Pinosylvin (a stilbene) and flavonoids have been identified
in pine wood. The flavonoids aromadendrin and taxifolin have been
isolated from larch wood. Both stilbenes and flavonoids have been
isolated from hardwoods. Many of these phenolic compounds have been
reported to possess valuable therapeutical properties, particularly
as antitumour agents and antioxidants. (S Nishibe, 1997, (3)), J D
Ford et al 1999 (4) and N M Saarinen et al 2000 (5)).
[0005] Juvabiones are a group of cyclohexane derivatives and are
useful e.g. as insecticides.
[0006] Before this invention, it has not been known that the knots
and branches of the trees are particularly rich in flavonoids and
stilbenes, compared to other parts of the tree.
[0007] Although it is been mentioned in the literature (2) that
certain lignans, particularly hydroxymatairesinol, occur in wood
and branches, so far no practically useful method has been
presented for isolating such compounds from wood.
SUMMARY OF THE INVENTION
[0008] This invention in based on the idea of combining the
isolation of phenolic substances or juvabiones from wood with the
utilization of wood in manufacturing of pulp or various mechanical
wood products. The aim of this invention is thus to i) provide a
practically useful source for these useful chemical substances and
ii) improve the economy for the manufacturing processes of pulp or
mechanical wood products in that by-products, hitherto used only
for energy production, are offered a new use as source for phenolic
substances and juvabiones.
[0009] Thus, this invention relates to a method for isolating of
phenolic substances or juvabiones from wood comprising knotwood,
said method comprising the steps of
[0010] a) extracting
[0011] i) the over-sized chip fraction obtained by screening
chipped wood, or
[0012] ii) a knot-rich sub-fraction obtained from said over-sized
chip fraction, or
[0013] iii) knotwood obtained as residue in finishing of mechanical
wood products
[0014] with a polar solvent, and
[0015] b) recovering the extract.
BRIEF DESCRIPTION OF THE DRAWINGS
[0016] FIG. 1 shows a vertical cross section of a tree stem with a
branch, and a vertical cross section of the branch. The figure
shows further sampling of the knots and branches for studying the
distribution of lignans. The dashed vertical line along the
outermost layer of the bark-free stem wood is defined as zero line
(0 cm).
[0017] FIGS. 2A to 2E show the distribution of total lignan in
opposite wood (diamonds), side wood (filled squares) and
compression wood (filled triangles) of five knot and samples and
three branch samples, taken at various positions from the zero line
shown in FIG. 1.
DETAILED DESCRIPTION OF THE INVENTION
[0018] The term "phenolic substances" shall be understood to cover
lignans, oligolignans, flavonoids, isoflavonoids, stilbenes,
tannins and phenolic acids. All these groups are mainly hydrophilic
substances that can be extracted with polar, i.e. hydrophilic
solvents.
[0019] The term "knotwood" shall be understood to include the
"knots", i.e. the part of the branches that is embedded in the
stem, and the branches extending outwards from the stem.
[0020] The "over-sized chip fraction" means the rejected fraction
obtained in the screening stage of the chips aimed for the pulping
process. This over-sized chip fraction, which can constitute about
1 to 5% of the total amount of wood chipped, cannot be forwarded as
such to the pulping process. Up to now, this fraction has been
recirculated to the chipping stage or withdrawn to be burnt. This
over-sized fraction comprises in addition to knotwood also
considerable amounts of "normal wood", i.e wood usable in the
pulping process. The amount of knotwood in the over-sized chip
fraction depends on the wood species and wood quality used, and is
estimated to about 10-30%. The content of knotwood in the
over-sized chip fraction is hereinafter also called "knot-rich
fraction".
[0021] Although it is possible to use the over-sized chip fraction
as such for extraction of phenolic substances, it may be preferable
to first separate the material into a knot-rich fraction and a
knot-poor fraction and to use the knot-rich fraction for
extraction. The "knot-poor fraction" means the "normal wood" that
can be led to the pulping process. This separation can be made
directly from the over-sized chip fraction, or the material can
first be refined before the screening stage.
[0022] "Knotwood obtained as residue in finishing of mechanical
wood products" includes, for example, the plywood sheet pieces
which include knots and are cut out and replaced by corresponding
pieces of normal plywood in the manufacturing stage before the
individual plywood sheets are pasted together to form the finished
product. Other examples are pieces of planks and boards rich in
knots and therefore rejected for various reasons in building and
construction, in furniture industry and the like. Also sawdust is
an example of such residues. However, the useful residues are not
restricted to those listed herein.
[0023] The "polar solvent" is either a single polar agent, or a
mixture of two or more polar agents, where said polar agent or
agents have a dielectric constant that is greater than 3,
determined at 25 Celsius degrees. As examples of polar solvents can
be mentioned pure water only, and mixtures of water and acetone and
water and alcohol, such as water and ethanol.
[0024] The extraction can be carried out on dried wood or on raw
wood material.
[0025] Although the extraction can be physically integrated with
the utilization of wood in the manufacturing of pulp or mechanical
wood products, the extraction can alternatively be carried out as a
separate process, because the knotwood, especially the knot-rich
fraction of the over-sized chips, can easily be transported and
stored for later processing.
[0026] The amount of phenolic substances in knotwood varies greatly
and depends on the phenolic substance in question and the wood
species used. Therefore, the extract derived from the extraction
stage may contain considerable concentrations of a desired phenolic
compound, and may therefore, depending on the purpose, be used as
such without further purification.
[0027] In case further purification is needed, the methods to be
used depend i.a. on the substance to be isolated and the desired
degree of purity. As examples of useful purification methods can be
mentioned chromatography or crystallization.
[0028] As important lignans to be isolated by the method of this
invention can be mentioned hydroxymatairesinol,
allohydroxymatairesinol, matairesinol, conidendrin, pinoresinol,
oxomatairesinol, lariciresinol, liovil, isolariciresinol,
secoisolariciresinol, picearesinol, conidendric acid, and
nortrachelogenin. However, the term "lignans" is not restricted to
these compounds.
[0029] "Oligolignans" are compounds having 3 to 6 phenylpropane
units that are beta-beta linked instead of normal lignans which
have two beta-beta linked phenylpropane units.
[0030] As examples of flavonoids which can be isolated according to
the method of this invention can be mentioned pinocembrin,
dihydrokaempferol, pinobanksin, naringenin, catechin,
2,4,6-trihydroxychalcone, aromadendrin and taxifolin.
[0031] As examples of stilbenes can be mentioned pinosylvin,
pinosylvin monomethyl ether, dihydropinosylvin, methylpinosylvin,
methyldihydropinosylvin and reservatrol.
[0032] As examples of juvabiones can be mentioned epijuvabione,
dehydrojuvabione, dihydroepijuvabione and epijuvabione acid.
[0033] The isolation of phenolic substances or juvabiones from
knotwood is very advantageous compared to the utilization of other
sources. In the knot-rich fraction of the over-sized chips, the
concentration of these substances is 10 to 1000 times higher than
in normal wood. Many of these compounds cannot be located at all in
normal wood. As a result, about 10-50% of the extract obtained
according to this method may be the phenolic agent or agents.
Another interesting feature is that a certain compound may be the
dominating compound of the derived phenolic group of substances.
For example, hydroxymatairesinol may be about 65-85% of the lignans
derived from spruce knotwood.
[0034] This invention thus offers a unique method for deriving the
desired phenolic compound or juvabione in high concentrations in
the extract. Along with this advantage, the wood material utilized
for the extraction is material that hitherto has been regarded as a
wood fraction useful as energy source only.
[0035] The invention will be illuminated by the following
non-restrictive Experimental Section
Experimental Section
[0036] Three different studies were carried out. In a first study,
three trees of Norway spruce (Picea abies) were investigated to
reveal information on the total lignan, hydroxymatairesionol and
oligolignan distribution in the knots and branches (Study 1). In a
second study, seven trees of Norway spruce (Picea abies) were
studied to reveal information on the total lignan,
hydroxymatairesinol and certain other lignans and oligolignan
distribution in the knots and branches (Study 2). In a third study
(Study 3), the content of various lignans in three Abies species,
three Picea species, two Larix species and three Pinus species were
investigated. In this study, also the contents of various stilbenes
and flavonoids were investigated. The methods used in Study 3 were
similar to those used in Study 1 and 2. The results from Study 3 is
shown in Table 5. Furthermore, samples from birch (Betula) and
aspen (Populus) have also been investigated according to the method
of this invention. In aspen, the flavonoids dihydrokaempferol and
naringenin were found, while certain stilbenes were found in
birch.
Study 1
[0037] Material
[0038] Three healthy Norway spruce trees (Table 1), grown in
southern Finland, were cut in May and discs were stored at
-24.degree. C. The heartwood from several 10 knots with a dead
branch (DK), still attached or fallen off, and several knots with a
living branch (LK) were sampled (Table 1). Stem heartwood (HW) and
sapwood (SW), with no visible compression wood, was sampled at 1.5
m height from each tree.
1TABLE 1 Trees, stemwood, and knot heartwood samples analysed.
Growth rings at 1.5 m Heartwood portion at 1.5 m Tree 1 66 Tree 2
71 {close oversize brace} 40-45% Tree 3 64 Stem Knot with dead
branch Knot with living branch Tree 1 HW 1.5* DK 3.5 LK 4.5 LK 13.5
SW 1.5 DK 5.5 LK 9-a LK 15-a LK 12 LK 15-b Tree 2 HW 1.5 DK 4.5 DK
13.5 LK 8 SW 1.5 DK 6.5 LK 9 DK 7.5 LK 14.5 Tree 3 HW 1.5 DK 6 DK
11-b LK 7 LK 17.5 SW 1.5 DK 8 DK 13.5 LK 12 DK 11-a LK 15 *m above
ground
[0039] The heartwood of five additional knots and three branches
were split into 1 cm thick sections and further divided into
opposite, side, and compression wood, for examining the
distribution of lignans (FIG. 1).
[0040] Methods
[0041] The wood samples were splintered, freeze-dried and ground in
a Wiley mill, producing particles passing a 10-mesh screen. A
second freeze-drying step after the milling ensured almost complete
removal of volatile compounds. The wood samples for the lignan
distribution study were freeze-dried, splintered using a scalpel,
and freeze-dried again.
[0042] Sequential extraction was carried out in an ASE-apparatus
(Accelerated Solvent Extractor). The lipophilic extractives were
first extracted with hexane (solvent temperature 90.degree. C.,
pressure 13.8 MPa, 2.times.5 min static cycles) and then the
hydrophilic extractives were extracted with an acetone:water (95:5
v/v) mixture (solvent temperature 100.degree. C., pressure 13.8
MPa, 2.times.5 min static cycles). The samples for the lignan
distribution study were extracted only with an acetone:water (95:5
v/v) mixture (as above, 3.times.5 min static cycles).
[0043] Lignans, free fatty acids, resin acids and free sterols
were, after evaporation of the extract solutions and silylation of
the extractives, analysed on a 25 m.times.0.20 mm i.d. crosslinked
methyl polysiloxane column (HP-1). Heneicosanic acid and betulinol
were used as internal standards. A correction factor of 1.2 for
betulinol was used for the quantification of the lignans. All
results given as mg/g or % (w/w) are calculated on a freeze-dried
wood basis.
[0044] Oligolignans, in the same silylated samples as above, were
analysed according to (6) using cholesteryl heptadecanoate as
internal standard.
[0045] Identification of individual components was done by GC-MS
analyses of the silylated components with an HP 6890-5973 GC-MSD
instrument, using a similar GC column as above.
[0046] Results
[0047] The lignans and oligolignans were the main components of the
hydrophilic extractives in all samples (Table 2). Small amounts
(<1.1 mg/g) of monomeric sugars were also found in most samples.
Trace amounts (<0.2 mg/g) of simple phenols, dominated by
coniferyl alcohol and vanillic acid, were also present in all
samples.
[0048] The amount of lignans was extremely high in the knots,
between 6 and 16% (w/w), compared to the maximum of 0.2% in the
stemwood (Table 2). The variation in the total amount of lignans
was large among the knots of a single tree and also between the
different trees. There seemed to be no difference between knots
with a living and a dead branch. The identified lignans were the
same as found earlier in Norway spruce (7). Ten unidentified
lignans were also detected and quantified. The sum of the two
isomers of hydroxymatairesinol (HMR) dominated all samples.
[0049] Two groups of a complex mixture of oligomeric phenolic
substances were detected and quantified by GC (Table 2).
Preliminary size-exclusion fractionation and MS studies showed
these compounds to be composed of phenylpropane units similar to
those of lignans. The SEC and GC retention times suggested that
these compounds were oligomeric (trimeric and tetrameric) lignans,
here called oligolignans. The total amount of oligolignans ranged
between 2 and 4% (w/w) in the knots. The amount of oligolignans was
about 20-30% of the amount of lignans in all knots.
2TABLE 2 The amount of lignans and oligolignans in the stemwood and
knot heartwood samples. HMR was the most abundant lignan in all
samples. Lignans Oligolignans Total HMR Trimers Tetramers mg/g % of
total mg/g mg/g Tree 1 HW 1.5 1.1 44 0.26 0.18 SW 1.5 0.7 53 0.17
0.08 DK 3.5 118 66 17 13 DK 5.5 123 68 17 10 LK 4.5 119 68 22 10 LK
9-a 124 69 21 9 LK 12 159 73 26 12 LK 13.5 120 73 20 10 LK 15-a 156
77 24 15 LK 15-b 149 80 25 14 Tree 2 HW 1.5 0.5 17 0.11 0.05 SW 1.5
0.2 25 0.07 0.03 DK 4.5 90 76 11 11 DK 6.5 134 75 16 14 DK 7.5 144
73 18 14 DK 13.5 117 76 15 11 LK 8 142 74 18 10 LK 9 129 75 17 10
LK 14.5 154 77 21 9 Tree 3 HW 1.5 2.2 37 0.32 0.13 SW 1.5 0.2 25
0.06 0.02 DK 6 63 72 8 10 DK 8 75 74 10 13 DK 11-a 124 76 17 14 DK
11-b 77 76 11 10 DK 13.5 114 81 14 13 LK 7 95 77 12 13 LK 12 145 82
16 9 LK 15 104 77 17 8 LK 17.5 134 76 25 14
[0050] FIGS. 2A to 2E show the distribution of lignans from near
the base of the knot to 10-20 cm out in the branch. The lignans
were almost absent 20 cm out in the branches from Tree 2 and 3,
while they had decreased strongly in the branch from Tree 1. The
amount of lignans in a radial direction, from the pith into the
outer branch, was quite even inside the stem. The concentration of
lignans decreased in the order opposite wood-side wood-compression
wood. However, since knots usually contain smaller amounts of
opposite wood than side wood and compression wood, the highest
amount of lignans is situated in side wood and compression
wood.
Study 2
[0051] In this study, results are reported from analysis of the
hydrophilic and lipophilic extractives of knots from seven Norway
spruce trees. The objective was to determine the amount and
composition, as well as the variability within a tree and between
trees, of the extractives in knotwood and stemwood. We were also
interested in differences due to geographical location, why two of
the seven trees were sampled from northern Finland.
[0052] Material
[0053] Seven healthy Norway spruce trees (Table 3) were felled, and
samples of stemwood and knots were sawn and put into storage at
-24.degree. C. within 6 h. Trees 1-3 were felled in May, trees 4-5
in January, and trees 6-7 in October. Trees 1-3 and 6-7 were
densely grown in sandy soil, while trees 4-5 were planted on former
arable land. Trees 1-5 had an average diameter of about 30 cm at
1.5 m height, while the diameters of trees 6-7 were about 25 cm.
Trees 4-5 had grown fast. The heartwood from knots at a dead branch
(DK), still attached or fallen off, and knots at a living branch
(LK) were sampled. For short, the expression knots means knot
heartwood. Stem heartwood (HW) from trees 1-3 and 6-7 and stem
sapwood (SW) from trees 1-7, with no visible reaction or decayed
wood, was sampled at 1.5 m height. The number in each sample code
(e.g. DK 3.5) expresses the height in meters above ground while the
suffix a or b expresses different knots at the same height. The
heartwood of seven additional knots and five branches were split
into 1 cm thick discs and further divided into opposite, side, and
compression wood, for examining the distribution of lignans (FIG.
1).
3 TABLE 3 Growth place Growth rings Heartwood proportion, area-%
(DK; LK)* at 1.5 m of cross section at 1.5 m Tree 1 Southern
Finland 66 (2; 6) Tree 2 Southern Finland 71 (4; 3) {close oversize
brace} 40-45 Tree 3 Southern Finland 64 (5; 4) Tree 4 Southern
Finland 17 (--; 2) Tree 5 Southern Finland 17 {close oversize
brace} 0 (--; 2) Tree 6 Northern Finland 150 (2; 1) Tree 7 Northern
Finland 134 {close oversize brace} 60-65 (2; 2) *Number of knot
heartwood samples sequentially extracted and analysed (knot with
dead branch; knot with living branch)
[0054] The methods for isolation and analysis of the isolated
substances were 15 essentially those described in Study 1
above.
[0055] Results
[0056] The lignans and oligolignans were the main components of the
hydrophilic extractives (Table 4). In addition, mainly small
amounts of monomeric sugars, simple phenols (or monolignols), and
dimeric non-lignan aromatic compounds of dilignoltype, such as
1,3-(bis-guaiacyl)-1,2-propan- diol, were detected in most samples.
The amount of lignans was exceptionally large in the knots compared
to the stemwood (Table 4). The knots of the trees from northern
Finland contained 14-24% (w w.sup.-1), while the knots from
southern Finland contained 6-16% (w w.sup.-1) of lignans. Even the
knots from the young trees contained 4-8% (w w.sup.-1) lignans.
Lignans were found in small amounts also in the sapwood.
[0057] The variation in the content of lignans was large between
the knots of a single tree and also between knots from different
trees. The largest amounts of lignans were found in living knots in
all trees. It is not possible to draw any conclusions whether the
larger amounts of lignans in the knots from the northern trees are
due to higher age or place of growth, since the trees from
different location were of different age. The tree genotype was
however different depending on the geographical location. The
branches of the northern trees have a smaller angle between the
stem and the downside of the branch. This is typical for Norway
spruce trees grown north of the pole circle. This, and the fact
that the climate is harder, would certainly cause another type of
stress on the knots than for trees from southern Finland. It has
been suggested earlier that external stress, causing eccentric
growth of Norway spruce stems, is associated with higher lignan
concentrations (2).
[0058] The main identified lignans were mainly the same as found
earlier in Norway spruce (7). The identified lignans were two
epimers of hydroxymatairesinol (HMR), .alpha.-conidendrin, liovil
(two isomers), secoisolariciresinol, lariciresinol, pinoresinol,
matairesinol, isolariciresinol, .alpha.-conidendric acid and a
lignan called lignan A. Seven unknown minor lignans were also
detected and quantified. The two epimers of HMR dominated in all
knots (Table 4). The epimers are here called HMR 1 and HMR 2 based
on their elution order on GC. The structure of the major lignan,
HMR 2, and its minor epimer, HMR 1 or allo-HMR, has been discussed
in the literature (8, 9). The ratio HMR 2/HMR 1 was between 2 and 4
in trees 1-2 and 6-7, while the range was 2-7 in tree 3, 4-5 in
tree 4, and 1-3 in tree 5. HMR was the dominating lignan also in
the stem sapwood and heartwood samples, even though the
contribution of the other lignans was higher. The ratio HMR 2/HMR 1
was between 2 and 5 in heartwood and even up to 11 in sapwood. In
the knots from tree 6 and 7 we found a lignan that has not been
identified earlier in spruce trees, in amounts ranging from 2 to 7
mg g.sup.-1. The mass spectrum and the GC retention time were the
same as for the lignan (-)-nortrachelogenin (NTG) which was
recently identified in knot heartwood of Pinus sylvestris (10).
However, the NTG-enantiomer (+)-wikstromol will have the same mass
spectrum and retention time. Since it was not possible to obtain
the lignan in enough pure form for determining the optical
rotation, it remains unclear, which of the enantiomers occurs in
Norway spruce. Also an earlier name, pinopalustrin, has been
suggested for NTG (11, 12).
[0059] The volume of the pith was quite large compared to the total
knot volume in most knots. Even though the dry mass of the pith is
small compared to the total knot mass, it was of interest to
analyse the extractives in the pith. The pith material contained
mainly lignans, about 120 mg g.sup.-1, with HMR as the most
abundant lignan. Small amounts of an NTG-isomer, with the same mass
spectrum but different GC retention time, was also identified in
the pith material.
4 TABLE 4 Lignans Oligolignans Total amount HMR* Coni* Liovil Seco*
Other Total amount** Trimers mg/g % of total lignans mg/g % Tree 1,
southern Finland, 66 years HW 1.5 0.9 52 6 9 5 28 0.4 59 SW 1.5 0.6
57 3 16 5 20 0.3 67 DK 3.5 115 68 6 5 10 11 30 58 DK 5.5 120 70 4 6
9 12 27 61 LK 4.5 116 70 9 4 4 13 32 69 LK 9-a 120 71 5 5 7 12 29
71 LK 12 159 73 3 5 6 12 38 68 LK 13.5 117 75 4 5 4 12 30 68 LK
15-a 152 80 3 3 3 12 38 62 LK 15-b 149 79 4 3 3 11 39 64 Tree 2,
southern Finland, 71 years HW 1.5 0.3 28 4 12 3 54 0.1 70 SW 1.5
0.1 37 3 10 7 44 0.2 69 DK 4.5 88 78 7 4 2 10 22 51 DK 6.5 131 76 5
4 4 11 30 54 DK 7.5 141 75 5 4 6 11 32 55 DK 13.5 114 78 4 3 4 11
26 57 LK 8 139 76 5 3 5 11 29 64 LK 9 126 76 4 3 5 11 26 63 LK 14.5
151 79 3 5 2 11 30 69 Tree 3, southern Finland, 64 years HW 1.5 1.9
43 2 20 9 25 0.1 72 SW 1.5 0.1 33 3 19 8 37 0.4 71 DK 6 61 75 8 6 3
9 18 45 DK 8 71 78 6 7 1 8 23 43 DK 11-a 119 79 4 7 1 8 31 56 DK
11-b 74 79 5 6 1 8 20 52 DK 13.5 111 84 4 3 1 8 27 51 LK 7 92 80 7
4 2 8 25 48 LK 12 141 85 4 3 1 8 25 65 LK 15 101 79 2 9 1 9 25 67
LK 17.5 131 78 1 10 1 10 39 64 Tree 4, southern Finland, 17 years
SW 1.5 0.3 38 <1 25 8 28 0.4 56 LK 3.5 47 72 1 8 11 9 19 63 LK
5.5 35 73 1 8 10 8 16 62 Tree 5, southern Finland, 17 years SW 1.5
0.6 53 6 19 6 16 0.4 59 LK 4 77 66 <1 6 20 9 23 62 LK 6 39 80
<1 6 8 6 23 61 Tree 6, northern Finland, 150 years HW 1.5 2.5 51
11 8 9 21 0.2 69 SW 1.5 0.2 28 <1 17 25 30 0.9 66 DK 2.5 139 71
9 3 6 10 31 42 DK 3 138 71 10 3 5 11 34 42 LK 9 171 77 8 3 2 10 33
45 Tree 7, northern Finland, 134 years HW 1.5 1.2 44 7 13 9 26 0.4
60 SW 1.5 0.6 28 1 26 19 25 0.9 59 DK 4 171 73 6 5 4 13 43 46 DK 5
172 73 6 4 5 12 48 43 LK 6 209 74 6 5 4 12 56 49 LK 8 244 72 6 5 3
14 44 49 *HMR = hydroxymatairesinol; Coni = .alpha.-conidendrin,
Seco = secoisolariciresinol **trimers and tetramers determined by
GC
[0060] Table 5 shows the results from Study 3.
5TABLE 5 Hydrophilic extractives, % of dry knot wood Abies Larix
alba balsamea sibirica sibirica lariciana Lignans: Isolariciresinol
0.6 0.3 Secoisolariciresinol 3.9 4.1 3.0 1.4 1.3 Lariciresinol 0.8
2.1 0.8 0.3 0.1 Flavonoids: 3,5,7,4',x- 0.3 0.5 pentahydroxy
flavanone Picea Lignans: abies glauca sitchensis
Secoisolariciresinol 0.6 0.3 Liovil 0.2 HMR(1) 2.1 0.8 HMR(2) 6.0
4.3 Pinus contorta sibirica sylvestris Lignans: Isolariciresinol
0.4 Secoisolariciresinol 0.3 0.02 Liovil 0.04 lariciresinol 2.6 NTG
0.1 1.7 Stilbenes: Pinosylvin-Me 0.1 2.4 1.0 Pinosylvin 0.2 0.9 1.1
Dihydropinosylvin-Me 1.4 Dihydropinosylvin 0.2 Flavonoids:
Pinocembrin 0.2 0.5 Pinobanksin 0.1 0.2 Dihydrokaempferol 0.05
0.02
[0061] Additional wood species were investigated according to the
methods described above and the following results were
obtained:
[0062] Abies concolor (secoisolariciresinol 2.5%, HMR 2 1.1%)
[0063] Abies lasiocarpa (epijuvabione 1.1%, dehydrojuvabione
0.8%)
[0064] Picea mariana (HMR 2 3.5%)
[0065] Pseudotsuga menziesii (dihydroepijuvabione 0.7%,
isolariciresinol 6.3%, secoisolariciresinol 2.0%, lariciresinol
1.5%)
[0066] Pinus banksiana (epijuvabione acid 1.2%, pinosylvin
monomethyl ether 1.1%, nortrachelogenin 1.3%)
[0067] Pinus resinosa (pinosylvin 1.4%, pinosylvin monomethyl ether
2.8%, matairesinol 1.2%)
[0068] Larix decidua (dihydrokaempferol 1.1%, taxifolin 2.8%,
isolariciresinol 1.1%, secoisolariciresinol 4.8%, lariciresinol
1.3%)
[0069] Betula pendula, Betula verrucosa, Alnus incana (several
stilbene glycosides 1-2%)
[0070] It will be appreciated that the methods of the present
invention can be incorporated in the form of a variety of
embodiments, only a few of which are disclosed herein. It will be
apparent for the expert skilled in the field that other embodiments
exist and do not depart from the spirit of the invention. Thus, the
described embodiments are illustrative and should not be construed
as restrictive.
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