Directory UMM :Data Elmu:jurnal:S:Soil Biology And Chemistry:Vol32.Issue6.Jun2000:
Soil Biology & Biochemistry 32 (2000) 747±755
www.elsevier.com/locate/soilbio
Biochemical properties of acid soils under climax vegetation
(Atlantic oakwood) in an area of the European temperate±humid
zone (Galicia, NW Spain): speci®c parameters
C. Trasar-Cepeda a, M.C. LeiroÂs b,*, F. Gil-Sotres b
a
Departamento de BioquõÂmica del Suelo, Instituto de Investigaciones AgrobioloÂgicas de Galicia, Consejo Superior de Investigaciones Cientõ®cas,
Apartado 122, E-15080 Santiago de Compostela, Spain
b
Departamento de EdafologõÂa y QuõÂmica AgrõÂcola, Facultad de Farmacia, Universidad de Santiago de Compostela, E-15706 Santiago de
Compostela, Spain
Accepted 6 October 1999
Abstract
The general and speci®c biochemical parameters of soils are highly sensitive to disturbance of the environment, but their use
for diagnosis of soil degradation is limited by lack of comparable published data and lack of accepted methodological standards.
With a view to establishing an appropriate data base for the soils of Galicia (NW Spain), we investigated the biochemical
properties of the O and Ah horizons of 40 native Umbrisols under climax Atlantic oakwood in this region. We report here our
results on speci®c biochemical parameters (i.e. extracellular hydrolytic enzyme activities) characterizing the phosphorus, nitrogen,
carbon and sulphur cycles. The enzymes studied were phosphomonoesterase (23.51 2 10.37 and 6.62 2 3.29 mmol p-nitrophenol
gÿ1 hÿ1, values for O and Ah horizons, respectively), phosphodiesterase (3.60 2 1.95 and 0.96 2 0.51 mmol p-nitrophenol gÿ1
hÿ1), casein-protease (2.97 2 0.83 and 0.94 2 0.32 mmol tyrosine gÿ1 hÿ1), BAA-protease (23.74 2 11.35 and 15.26 2 8.91 mmol
NH3 gÿ1 hÿ1), urease (24.90 2 13.60 and 16.59 2 10.61 mmol NH3 gÿ1 hÿ1), CM-cellulase (0.59 2 0.17 and 0.23 2 0.10 mmol
glucose gÿ1 hÿ1), invertase (12.6622.75 and 6.9322.14 mmol glucose gÿ1 hÿ1), b-glucosidase (8.4325.14 and 1.5520.89 mmol
p-nitrophenol gÿ1 hÿ1), and arylsulfatase (0.672 0.30 and 0.46 2 0.20 mmol p-nitrophenol gÿ1 hÿ1). For the variables for which
comparable data are available, the values observed are generally within previously published ranges. Principal components
analysis of the combined biochemical, physical and chemical data for these soils shows ®ve factors, of which the three most
important concern microbial activity and its logical dependence on nutrient content, the accumulation of soil organic matter and
the mineralization of soil organic matter. 7 2000 Elsevier Science Ltd. All rights reserved.
Keywords: Soil biochemical properties; Soil enzymes; Temperate forest soils
1. Introduction
It is widely accepted that soil quality can be
expressed in terms of the capability of a soil to accept,
store and recycle the water, minerals and energy
required for optimal crop production while at the
* Corresponding author. Tel.: +34-981-563-100, ext. 15042; fax:
+34-981-594-912.
E-mail address: [email protected] (M.C. LeiroÂs).
same time preserving a healthy environment (Arshad
and Cohen, 1992; Parr et al., 1992). Thus good-quality
soils should carry out the following functions: crop
production, ®ltration and degradation. The most important of these three functions for the evaluation of
soil quality is perhaps degradation, as it de®nes the capacity of a soil to mineralize soil organic matter and
degrade exogenous plant material and anthropogenic
inputs such as organic wastes, pesticides, hydrocarbons, etc. (Dick, 1997). The degradation function
0038-0717/00/$ - see front matter 7 2000 Elsevier Science Ltd. All rights reserved.
PII: S 0 0 3 8 - 0 7 1 7 ( 9 9 ) 0 0 1 9 6 - 0
748
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
depends to a large extent on the biochemical properties
of the soil (Visser and Parkinson, 1992), which are
usually divided into two groups (Nannipieri et al.,
1995): general biochemical parameters, which are
directly related to the number and activity of soil
microorganisms, and speci®c parameters, the activities
of extracellular hydrolytic enzymes stabilized by being
bound in complexes with clay minerals and humic colloids.
Hydrolytic extracellular soil enzymes make nutrients
available to plants and microorganisms by converting
them from unassimilable to readily assimilable forms
(Saratchandra et al., 1984). However, although this
implies that they exert a very important part in the
degradation function, this potential for characterising
soil quality has often been ignored. This is partly due
to there having been relatively few studies in which the
activities of a large number of enzymes have been studied simultaneously, and partly to the diculty in
comparing data obtained for dierent soils, by dierent workers using dierent incubation conditions, substrates and substrate concentrations and buer systems
(Saratchandra et al., 1984). What is needed, if the potential of enzymatic activities and other biochemical
properties for soil quality assessment is to be realized,
is an extensive database of the biochemical properties
of soils from dierent parts of the world. Moreover,
since the goal of sustainable development implies that
soil quality should be determined with reference to
undisturbed native soils (Doran et al., 1994), the compilation of such a database should give priority to climax soils in so far as is possible.
The above considerations have led us to compile an
extensive database of the biochemical properties of climax soils in Galicia (NW Spain), a region situated in
the European temperate±humid zone in which extensive areas of climax vegetation (Atlantic oakwood) still
exist. The general biochemical parameters of these climax soils (all Umbrisols; ISSS Working group RB,
1998) were reported by LeiroÂs et al. (1999). Here we
report the activities of a variety of hydrolytic enzymes
involved in the carbon, nitrogen, phosphorus and sulphur cycles, including enzymes with substrates of both
high and low molecular mass, and we discuss their relationships with each other and with the physical,
chemical and general biochemical parameters of the
soils.
2. Material and methods
2.1. Soils
We studied 40 soils developed under climax vegetation dominated by Quercus robur L. or Q. pyrenaica
L. at sites distributed throughout Galicia, NW Spain.
At all sites, the sampling area displayed little if any
disturbance of human origin, and its tree vegetation
was composed mainly of healthy adult specimens. All
the soil are Umbrisols (ISSS Working Group RB,
1998); their locations, the methods used for sampling
and for determination of their physical, chemical and
general biochemical parameters, and the values of
these parameters, have been described by LeiroÂs et al.
(1999).
2.2. Analytical methods
The activities of urease (EC 3.5.1.5) and of proteases
(EC 3.4.4) hydrolysing benzoylargininamide (BAAprotease) and casein (casein-protease) were determined
as described by Gil-Sotres et al. (1992). Brie¯y, urease
activity was determined using urea as substrate, incubating for 1.5 h at 378C and pH 7.1 (phosphate buer
0.2 M) and measuring the NH+
4 released with an ammonium electrode. BAA-protease activity was determined using the same incubation conditions and the
same method to determine NH+
4 but with a-benzoylN-argininamide (BAA) as substrate. In both cases enzymatic activity is expressed in mmol NH3 gÿ1 hÿ1.
Casein-hydrolysing activity was determined with casein
as substrate, incubating for 2 h at 508C and pH 8.1
(Tris±HCl buer 0.05 M) and determining the amino
acids released by the Folin colorimetric method; enzymatic activity is expressed in mmol tyrosine gÿ1 hÿ1.
Acid phosphomonoesterase (EC 3.1.3.2), b-glucosidase (EC 3.2.1.21), phosphodiesterase (EC 3.1.4.1) and
arylsulfatase (EC 3.1.6.1) activities were determined by
incubating the soils with a substrate containing a pnitrophenyl
moiety
and
spectrophotometrically
measuring the amount of p-nitrophenol liberated by
enzymatic hydrolysis. In these cases the enzymatic activity is expressed in mmol p-nitrophenol gÿ1 hÿ1. Acid
phosphomonoesterase activity was determined with pnitrophenyl phosphate as substrate, incubating at pH
5.0 (Modi®ed Universal Buer) and 378C. After 30
min 2 M CaCl2 was added (to stop the reaction and to
avoid the brown coloration caused by organic matter)
and the liberated p-nitrophenol was extracted with 0.2
M NaOH (Tabatabai and Bremner, 1969; Saa et al.,
1993). b-glucosidase activity was determined as
described for phosphomonoesterase activity except
that the substrate was p-nitrophenyl-b-glucopyranoside
and the p-nitrophenol released was extracted with
THAM±NaOH 0.1 M of pH 12 (Eivazi and Tabatabai, 1988). Phosphodiesterase activity was determined
with bis-p-nitrophenyl phosphate as substrate, incubating at pH 5.0 (THAM buer 0.05 M) and 378C for 1
h (Bowman and Tabatabai, 1978). Arylsulphatase activity was determined with p-nitrophenyl sulphate as
substrate, incubating at pH 5.8 (acetate buer 0.5 M)
and 378C for 1 h (Tabatabai and Bremner, 1970).
Table 1
Values of the speci®c biochemical parameters studied, in O horizons n 40 of climax soils under oakwood in Galicia (NW Spain)
Total C (%)
Total N (%)
Phosphomonoesterasea
Phosphodiesterasea
Arylsulfatasea
Casein-proteaseb
BAA-proteasec
Ureasec
CM-cellulased
b-glucosidasea
Invertased
7.1
8.1
10.1
12.1
13.1
14.1
15.1
16.1
17.1
18.1
19.1
20.1
22.1
23.1
24.1
25.1
27.1
28.1
29.1
31.1
33.1
34.1
35.1
36.1
37.1
39.1
40.1
41.1
42.1
43.1
44.1
45.1
46.1
47.1
48.1
49.1
50.1
51.1
52.1
53.1
31.2
28.6
14.6
21.2
21.0
18.3
17.3
27.8
23.1
17.8
32.3
26.9
18.2
50.0
46.2
37.8
19.2
36.2
32.4
40.0
28.5
31.0
39.8
39.2
26.6
35.7
36.5
39.3
31.5
39.6
33.4
38.8
26.6
31.0
36.8
38.6
34.5
19.4
25.3
28.8
1.57
1.76
0.95
1.19
1.16
0.97
1.07
1.51
1.29
1.10
1.57
1.56
1.52
1.92
2.07
1.60
1.02
1.66
1.92
2.17
1.50
1.83
1.76
1.83
1.51
1.71
1.83
1.95
1.59
1.86
1.58
1.81
1.26
1.75
1.88
1.80
1.68
0.95
0.77
1.28
10.49
40.36
9.17
24.59
12.75
36.91
9.47
30.03
10.79
12.38
19.20
15.51
11.93
16.74
24.05
18.17
11.76
15.90
9.35
14.38
33.89
24.29
38.19
20.78
27.47
19.20
34.40
35.49
12.47
30.68
47.45
34.02
32.84
18.52
34.95
31.97
33.03
25.59
21.22
30.00
1.65
4.33
1.21
1.72
3.50
2.73
2.37
2.90
2.58
2.22
7.44
3.88
2.36
3.61
4.81
3.08
3.47
2.53
2.09
3.62
5.56
1.67
2.49
3.02
5.20
4.02
8.01
5.48
2.16
2.91
3.02
11.82
3.15
3.17
3.19
3.06
4.55
3.10
3.40
3.05
0.20
1.03
0.47
0.27
0.67
0.60
0.88
0.70
0.80
0.47
0.69
0.73
0.41
0.53
0.83
0.35
0.53
0.34
0.29
0.22
0.86
0.72
0.55
0.40
1.12
1.58
0.61
1.06
0.52
0.62
0.56
0.67
0.63
1.50
0.65
0.53
0.81
0.81
0.90
0.50
2.76
4.04
2.88
3.66
3.54
2.84
3.15
4.33
3.47
2.43
4.11
3.71
2.69
4.48
4.08
3.04
1.52
1.78
1.15
2.43
3.60
2.13
2.17
3.99
2.41
1.15
3.81
3.30
2.96
2.90
2.58
2.09
1.85
3.50
3.19
3.32
2.83
3.16
2.85
3.02
11.93
18.33
20.54
8.90
29.90
24.98
30.86
13.02
36.21
18.22
39.14
48.97
29.85
23.81
24.99
22.23
21.87
14.12
5.91
10.07
34.48
26.17
10.47
21.99
41.57
19.30
42.61
14.67
28.81
12.52
27.88
16.92
33.05
23.66
40.05
5.84
44.93
16.17
8.13
26.44
12.66
11.37
18.34
13.89
28.27
16.44
28.50
11.43
29.31
19.62
19.80
40.10
28.18
34.57
24.12
30.71
14.47
16.94
12.04
9.35
29.29
38.70
22.68
31.77
65.60
20.15
35.19
20.08
20.81
14.52
66.26
20.01
33.03
21.00
33.57
10.81
47.12
15.12
1.76
28.32
0.62
0.88
0.31
0.54
0.46
0.51
0.30
0.83
0.45
0.58
0.71
0.54
0.70
0.88
0.93
0.75
0.27
0.57
0.31
0.40
0.67
0.80
0.78
0.74
0.50
0.56
0.56
0.41
0.48
0.79
0.68
0.64
0.52
0.44
0.53
0.86
0.61
0.52
0.51
0.53
7.45
15.26
5.31
6.71
4.36
7.31
4.43
11.64
9.05
6.56
2.05
7.13
5.89
13.86
8.20
12.05
4.00
6.42
4.23
7.07
11.41
6.29
5.68
5.08
4.93
6.98
4.21
29.63
3.77
17.09
17.00
4.34
8.76
14.15
13.79
9.04
7.61
7.73
3.83
6.71
12.44
15.56
11.21
12.40
9.39
14.31
11.48
16.92
13.41
13.76
15.77
12.93
14.59
16.20
16.40
14.35
13.39
18.83
10.19
10.41
13.70
15.44
16.93
17.72
11.52
9.03
8.47
10.08
10.19
12.96
11.33
9.62
9.36
12.19
9.95
12.81
11.31
7.92
10.64
11.28
Mean
S.D.
30.5
8.6
1.54
0.35
23.51
10.37
3.60
1.95
0.67
0.30
2.97
0.83
23.74
11.35
24.90
13.60
0.59
0.17
8.43
5.14
12.66
2.75
mmol
mmol
c
mmol
d
mmol
p-nitrophenol gÿ1 hÿ1.
tyrosine gÿ1 hÿ1.
NH3 gÿ1 hÿ1.
glucose gÿ1 hÿ1.
749
a
b
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Sample
750
Table 2
Values of the speci®c biochemical parameters studied, in Ah horizons n 40 of climax soils under oakwood in Galicia (NW Spain)
Total C (%)
Total N (%)
Phosphomonoesterasea
7.2
8.2
10.2
12.2
13.2
14.2
15.2
16.2
17.2
18.2
19.2
20.2
22.2
23.2
24.2
25.2
27.2
28.2
29.2
31.2
33.2
34.2
35.2
36.2
37.2
39.2
40.2
41.2
42.2
43.2
44.2
45.2
46.2
47.2
48.2
49.2
50.2
51.2
52.2
53.2
10.9
10.0
5.7
9.5
6.3
5.2
7.8
12.3
8.3
6.6
12.3
9.5
8.9
14.5
9.5
12.5
11.1
9.4
14.8
11.6
10.3
14.6
12.2
18.3
11.8
11.5
6.6
13.1
9.7
10.2
7.8
15.1
14.7
9.3
13.2
10.1
15.2
6.7
7.9
15.1
0.61
0.70
0.39
0.51
0.37
0.32
0.56
0.76
0.55
0.40
0.65
0.57
0.59
0.81
0.68
0.57
0.54
0.44
0.93
0.79
0.68
0.96
0.77
0.99
0.85
0.74
0.40
0.79
0.63
0.63
0.43
0.85
0.76
0.72
0.92
0.63
0.98
0.41
0.48
0.69
3.25
8.64
2.23
6.72
2.43
7.87
2.73
6.95
4.10
3.99
6.56
2.24
3.20
4.64
5.90
3.12
4.33
3.50
6.04
5.26
9.97
8.99
8.46
3.76
12.57
8.98
6.76
12.04
3.70
6.64
6.93
13.82
11.31
6.23
6.66
8.22
8.64
6.75
5.00
15.76
Mean
S.D.
10.8
3.1
0.65
0.18
6.62
3.29
a
mmol
mmol
c
mmol
d
mmol
b
p-nitrophenol gÿ1 hÿ1.
tyrosine gÿ1 hÿ1.
NH3 gÿ1 hÿ1.
glucose gÿ1 hÿ1.
Phosphodiesterasea
Arylsulfatasea
Casein-proteaseb
BAA-proteasec
Ureasec
CM-cellulased
b-glucosidasea
Invertased
0.45
0.97
0.40
0.56
0.64
1.16
0.81
0.76
1.12
0.72
1.68
0.97
0.92
1.62
1.96
0.44
1.02
0.63
0.84
0.86
0.84
0.72
0.88
0.54
1.24
1.23
0.62
2.28
0.82
0.86
0.38
0.30
1.56
1.02
0.76
0.88
0.83
0.85
0.69
2.69
0.14
0.68
0.21
0.07
0.42
0.25
0.54
0.31
0.63
0.25
0.66
0.38
0.37
0.56
0.46
0.07
0.58
0.34
0.41
0.35
0.56
0.51
0.44
0.23
0.91
0.43
0.54
0.85
0.41
0.56
0.33
0.52
0.70
0.85
0.43
0.27
0.65
0.67
0.57
0.45
0.83
1.19
0.95
1.44
0.63
1.15
1.34
1.67
1.19
0.84
0.62
0.61
0.66
0.91
0.96
0.36
0.71
0.69
0.87
0.51
0.69
0.68
0.64
1.02
1.17
1.13
0.78
1.40
0.99
1.28
0.72
0.70
0.84
1.75
0.66
1.40
0.88
1.12
0.93
0.82
3.63
6.75
16.06
3.73
15.94
5.50
15.86
7.10
25.44
8.33
25.60
28.96
25.86
21.94
13.16
5.03
14.74
8.52
13.80
5.88
23.44
12.31
12.43
8.02
27.64
14.73
11.69
8.80
24.81
12.11
13.21
14.78
30.10
18.69
33.21
2.67
34.33
8.36
4.48
22.79
7.38
5.94
10.21
5.96
14.53
6.59
17.14
10.33
19.94
10.55
26.08
22.50
18.43
30.03
12.83
6.64
6.99
3.92
18.12
10.28
17.74
28.50
20.33
23.24
47.14
9.47
6.46
13.53
15.47
13.67
49.78
11.98
28.56
18.59
26.58
3.17
25.27
14.58
3.60
21.36
0.26
0.33
0.06
0.25
0.10
0.09
0.11
0.26
0.11
0.15
0.23
0.18
0.18
0.26
0.36
0.22
0.06
0.21
0.32
0.26
0.19
0.47
0.33
0.24
0.19
0.30
0.17
0.22
0.06
0.33
0.22
0.33
0.39
0.13
0.28
0.42
0.14
0.16
0.14
0.34
1.13
1.92
1.02
1.09
0.77
1.48
1.16
2.36
1.63
1.20
1.71
3.86
0.93
1.42
2.72
1.03
0.95
1.41
1.46
1.12
0.94
1.62
1.22
0.67
1.10
1.98
1.07
1.97
0.73
1.72
0.77
1.56
4.58
2.02
1.58
0.95
0.89
1.45
0.83
4.06
5.96
8.67
4.75
5.76
7.29
4.42
9.27
5.36
10.95
7.22
7.53
7.49
6.69
8.47
4.57
2.00
9.37
9.12
8.58
9.91
8.35
5.66
2.97
4.43
10.89
6.49
3.75
7.85
9.85
4.46
6.50
6.82
8.76
8.02
6.87
5.58
8.43
5.53
7.64
4.91
0.96
0.51
0.46
0.20
0.94
0.32
15.26
8.91
16.59
10.61
0.23
0.10
1.55
0.89
6.93
2.14
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Sample
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Invertase (EC 3.2.1.26) activity was determined with
saccharose as substrate, incubating for 3 h at 508C and
pH 5.5 (acetate buer 2 M) and determining reducing
sugars as per Schinner and von Mersi (1990). Carboxymethylcellulase (CM-cellulase) activity was determined
similarly, except that the substrate was carboxymethylcellulose and the incubation time was 24 h (Schinner
and von Mersi, 1990). The enzymatic activities are
expressed in mmol glucose gÿ1 hÿ1.
2.3. Expression and analysis of results
All determinations were performed in triplicate, and
all values reported are averages of triplicate determinations expressed on an oven-dried soil basis (1058C).
Statistical analyses were performed using Statistics 4.5
for Windows (StatSoft Inc., 1993)
3. Results and discussion
The values, means and standard deviations of the
enzyme activities in the O and Ah horizons are listed
in Tables 1 and 2.
3.1. Enzymatic activities
3.1.1. Acid phosphomonoesterase
Mean acid phosphomonoesterase activity in the O
horizons was 23.51 mmol p-nitrophenol gÿ1 hÿ1 (S.D.
10.37, range 9.17±47.45, coecient of variation CV
44%). In the Ah horizons it fell to 6.62 mmol p-nitrophenol gÿ1 hÿ1 (S.D. 3.29, range 2.23±15.76, CV
50%). Although phosphomonoesterase has been one
of the most extensively studied soil enzymes (Burns,
1978), data for the organic layers of woodland soils
are relatively scarce. The values observed in this study
lie within the range found in woodland soils in the
Apennines (Nannipieri et al., 1980), in subarctic
regions (Neal, 1982) and in Canada (Pang and
Kolenko, 1986). Similarly, the Ah horizon values
obtained in this study fall within the range reported in
the much more abundant literature on phosphomonoesterase in mineral horizons (Speir, 1977; Frankenberger and Dick, 1983).
3.1.2. Phosphodiesterase
Mean phosphodiesterase activity in the O horizons
was 3.60 mmol p-nitrophenol gÿ1 hÿ1 (S.D. 1.95, range
1.21±11.82, CV 54%), between six and seven times less
than phosphomonoesterase activity; in the Ah horizons
activity ranged from 0.30 to 2.69 mmol p-nitrophenol
gÿ1 hÿ1 (mean 0.96, S.D. 0.50, CV 52%). References
to phosphodiesterase activities in the literature are
scarce and generally unsystematic. Rastin et al. (1988,
1990) mention very low values for both soil layers
751
(between 0 and 0.02 mmol p-nitrophenol gÿ1 hÿ1). For
Ah horizons, Frankenberger and Dick (1983) reported
values similar to those observed in this work, but Ross
et al. (1995a,b) found much higher values of between 7
and 22 mmol p-nitrophenol gÿ1 hÿ1.
3.1.3. Arylsulfatase
Mean arylsulfatase activity was 0.66 mmol p-nitrophenol gÿ1 hÿ1 in the O horizons (S.D. 0.30, CV 45%,
range 0.20±1.58) and 0.45 mmol p-nitrophenol gÿ1 hÿ1
in the Ah horizons (S.D. 0.20, CV 43%, range 0.07±
0.91). Although the values observed in this study in
both the O and Ah horizons are within the range
reported in the literature for natural ecosystems, the
maximum values are considerably lower than those
found in organic layers by authors such as Baligar and
Wright (1991) and Saratchandra et al. (1984), who
reported maxima of 3.89 and 5.69 mmol p-nitrophenol
gÿ1 hÿ1, respectively. Although this discrepancy may
be partly due to methodological dierences, it suggests
that in native Galician soils the sulphur requirement of
soil microorganisms is satis®ed, at least in the top few
centimetres of the soil, with ensuing downregulation of
arylsulphatase. This implies the possibility of these
soils developing an arylsulfatase de®ciency, the consequences of which for the sulphur cycle are dicult to
predict.
3.1.4. Casein-protease
Casein-protease activity ranged from 1.15 to 9.08
mmol tyrosine gÿ1 hÿ1 in the O horizons (mean 3.10,
S.D. 1.26, CV 41%), and from 0.36 to 1.75 mmol tyrosine gÿ1 hÿ1 in the Ah horizons (mean 0.94, S.D. 0.32,
CV 43%). The values observed in Ah horizons are
within the range reported by several authors for native
soils (e.g. Nannipieri et al., 1980; BonmatõÂ et al., 1991;
Perucci, 1992). We have been unable to ®nd references
to casein-protease activity in O horizons; nevertheless,
the values of 5.50±38.67 mmol tyrosine gÿ1 hÿ1 found
by Dilly and Munch (1995) in rotting Alnus litter are
much higher than those found in this study. The
observed depthwise variation suggests that proteolysis
of substrates of high molecular weight diminishes as
the decomposition of plant debris advances, i.e. in the
order litter, O layer, humus, at least when the results
are expressed as activity per unit mass of soil.
3.1.5. BAA-protease
In the O horizons BAA-protease activity ranged
from 5.84 to 48.97 mmol NH3 gÿ1 hÿ1 (mean 23.50,
S.D. 11.54 CV 49%), and in Ah horizons from 2.67 to
34.33 mmol NH3 gÿ1 hÿ1 (mean 15.26, S.D. 8.91, CV
58%). No other values for temperate zone soils appear
to be available for comparison, in spite of the proposal
of Ladd and Buttler (1972) that BAA be used to determine the activity of proteases acting on small peptides.
1.00
1.00
0.48a
1.00
0.64a
0.59a
1.00
0.31b
0.28c
0.28c
1.00
0.71a
0.24d
0.22
0.34b
1.00
0.35b
0.24
0.76a
0.60a
0.44a
P < 0.001.
P < 0.01.
c
P < 0.02.
d
P < 0.05.
a
CM-cellulase
Urease
BAA-protease
Casein-protease
Arylsulfatase
1.00
0.35b
0.42a
0.32b
0.27c
0.42a
0.27c
1.00
0.45a
0.59a
0.43a
0.31b
0.61a
0.48a
0.39a
b
3.1.9. Invertase
Invertase activity ranged from 2.27 to 18.83 mmol
glucose gÿ1 hÿ1 in the O horizons (mean 12.06, S.D.
3.41, CV 28%) and from 2.00 to 10.95 mmol glucose
gÿ1 hÿ1 in the Ah horizons (mean 6.85, S.D. 2.22, CV
28%). Both sets of values are similar to those found
by Batistic et al. (1980), Frankenberger and Dick
(1983), Speir et al. (1984), Schinner and von Mersi
(1990) and Ross et al. (1995a,b).
1.00
0.69a
0.43a
0.61a
0.33b
0.40a
0.76a
0.73a
0.52a
3.1.8. b-Glucosidase
Mean b-glucosidase activity was 8.42 mmol p-nitrophenol gÿ1 hÿ1 in the O horizons (S.D. 5.14, range
2.05 to 29.63, CV 61%) and 1.55 mmol p-nitrophenol
gÿ1 hÿ1 in the Ah horizons (S.D. 0.89, range 0.67 to
4.58, CV 57%). The Ah horizon values can be considered normal, but the O layer values are quite high
in comparison with those found by Batistic et al.
(1980), Kanazawa and Miyashita (1987) and Deng and
Tabatabai (1996b).
Phosphodiesterase
3.1.7. CM-cellulase
In the O horizons CM-cellulase activity ranged from
0.27 to 0.93 mmol glucose gÿ1 hÿ1 (mean 0.59, S.D.
0.17, CV 29%), and in the Ah horizons from 0.06 to
0.47 mmol glucose gÿ1 hÿ1 (mean 0.23, S.D. 0.10, CV
43%). The Ah horizon values are comparable with
those reported in the literature, but those observed in
O horizons are slightly lower than reported values
(Kanazawa and Miyashita, 1987; Ohtonen, 1994).
Table 3
Pearson coecients of pairwise correlation among the enzymatic activities studied (the analysis pools results from both O and Ah horizons)
3.1.6. Urease
Urease activity ranged from 1.76 to 66.26 mmol NH3
gÿ1 hÿ1 in O horizons (mean 24.90, S.D. 13.60, CV
55%) and from 3.17 to 49.78 mmol NH3 gÿ1 hÿ1 in Ah
horizons (mean 16.58, S.D. 10.67, CV 64%). Although
urease, like phosphomonoesterase, has been one of the
most extensively studied soil enzymes (Burns, 1978),
most published work refers to agricultural soils, urea
being one of the most widely used nitrogenated fertilizers in many parts of the world. Research on urease
activity in woodland soils has been relatively scant.
The values observed in this study are similar to those
found by Nannipieri et al. (1980) in soils from the
Apennine Mountains, but between 2 and 20-times
higher than those reported by Speir (1977), Speir et al.
(1980), Saratchandra et al. (1984) and Deng and Tabatabai (1996a).
b-glucosidase
It may be worth mentioning that the values reported
for three soils from the Apennine Mountains by Nannipieri et al. (1980) are within the range found in this
work.
Phosphomonoesterase
Phosphodiesterase
Arylsulfatase
Casein-protease
BAA-protease
Urease
CM-cellulase
b-glucosidase
Invertase
Invertase
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Phosphomonoesterase
752
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
3.2. Correlations between speci®c biochemical
parameters
As in the companion study of the general biochemical parameters of these soils, and for the same reasons
as were discussed by LeiroÂs et al. (1999), correlations
among dierent biochemical properties were sought
considering the data for the O and Ah horizons
jointly. Almost all the activities reported in this paper
are signi®cantly correlated with each other (P < 0.001
in most cases; see Table 3). As in related studies (BonmatõÂ et al., 1991; Tate et al., 1991; Perucci, 1992), the
enzyme correlating best with other enzymes is phosphomonoesterase, which correlates especially closely
with CM-cellulase and b-glucosidase, followed by
phosphodiesterase and casein-protease. Since phosphodiesterase is also closely correlated with casein-protease
and CM-cellulase, there is clearly a strong interrelationship among enzymatic processes involved in
the carbon, nitrogen and phosphorus cycles. By contrast, arylsulfatase is not very highly correlated with
any other enzymatic activity; this apparent lack of
coupling between the sulphur cycle and the other
major nutrient cycles, which contrasts with the results
of Speir et al. (1980, 1984), Frankenberger and Dick
(1983) and Saratchandra et al. (1984), may be due to
the downregulation hypothesized above.
Among the nitrogen cycle enzymes there is close correlation between urease and BAA-protease, but neither
of these two enzymes correlates highly with casein-protease, which contrasts with the ®ndings of Speir et al.
(1980). This pattern suggests that in Galician soils the
degradation of proteins and the degradation of smaller
nitrogenated compounds such as peptides and urea are
subject to dierent regulatory mechanisms.
Among the carbon cycle enzymes, CM-cellulase is
highly correlated with b-glucosidase and invertase, and
these latter are also moderately well correlated with
each other. This re¯ects the synergism among these
enzymes in the degradation of the carbon compounds
received by the soil (Panda and Sharma, 1994; Deng
and Tabatabai, 1996a,b).
Similarly, the phosphorus cycle enzymes phosphomonoesterase and phosphodiesterase are likewise closely correlated, which is in keeping with the ®ndings of
Frankenberger and Dick (1983) and Rastin et al.
(1988) and shows the coupling between the enzymes of
this cycle.
3.3. Relationships within chemical, physical and
biochemical properties
The enzyme activities we measured also correlate
closely with physical and chemical properties related
to the availability of water and nutrients (results
not shown) and with many of the general biochemi-
753
cal parameters discussed by LeiroÂs et al. (1999). To
clarify the structure of these interdependences, we
performed a joint principal components analysis
(PCA) of our data on the physical, chemical and
biochemical properties of the O and Ah layers of
these soils, including both general and speci®c biochemical parameters. The ®ve main factors identi®ed
together account for 76% of the variance; the loadings of the 27 soil characteristics considered on
each factor are listed in Table 4.
Factor I, which accounts for 28% of the total
variance, exhibits close positive correlation with dehydrogenase, microbial biomass C and available
Ca2+ and K+, and somewhat lower positive correlation with ATP, microbial biomass N, catalase, invertase, casein-protease and available P, and may thus be
regarded as related to the size and activity of the microbial community. The presence of available elements
among its de®ning variables may be attributed to the
logical dependence of microbial activity on nutrient
contents. The high loadings of invertase and caseinprotease suggest that, contrary to what has been
suggested by Skujins (1978), Nannipieri et al. (1982)
and Speir and Ross (1990), the activities of both these
enzymes depend more on the microbial community
than on their extracellular accumulation, in these soils
at least.
Factor II accounts for 16% of the total variance.
At its positive pole it is de®ned mainly by phosphomonoesterase, although CM-cellulase, the C-to-N
ratio and total C and N contents also have high
loadings, and at its negative pole by pH. Though it
is not easy to interpret, these associations suggest
that it is related to the accumulation of hydrolytic
enzymes and of organic matter that is poorly humidi®ed because of the acidity of the medium. The
relationship of phosphomonoesterase and CM-cellulase to the accumulation of organic matter has been
mentioned by Harrison (1983) and Sinsabaugh and
Linkins (1988), among others.
Factor III accounts for 15% of the total variance.
Its main de®ning property is nitrogen mineralization
capacity as shown by total inorganic N, although soil
respiration and ammoni®cation also contribute to it.
Together with the relatively high loadings of the total
N and C contents, these properties suggest that Factor
III represents the mineralization of organic matter.
Factor IV, which accounted for only 9% of the total
variance, may be regarded as re¯ecting the nature and
weathering of the parent material, being de®ned by Al
and Fe oxide contents. Factor V is de®ned by BAAprotease and urease; though it accounts for only 8%
of the total variance, its emergence as a separate factor
con®rms the independence of the degradation of low
molecular weight nitrogenated compounds, in keeping
with the correlation results discussed above.
754
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Table 4
Loadings of biochemical, physical and chemical properties on the axes identi®ed by principal components analysis of their values in the O and
Ah horizons of climax soils under oakwood in Galicia (NW Spain)
Factor I
Biochemical properties
Microbial biomass-C
Microbial biomass-N
Evolved CO2-C
ATP
Dehydrogenase
Catalase
N mineralized (N-NH+
4 )
N mineralized (N inorganic total)
Phosphomonoesterase
Phosphodiesterase
CM-cellulase
b-glucosidase
Invertase
Casein-protease
BAA-protease
Urease
Arylsulfatase
Physical and chemical properties
pH KCl
Total C
Total N
C/N
Labile Pi
Available Ca2+
Available K+
Available water
Al2O3
Fe2O3
Explained variance (%)
Accumulated explained variance (%)
Factor II
Factor II
Factor IV
Factor V
0.70
0.61
0.37
0.64
0.84
0.66
0.24
0.34
0.48
0.41
0.50
0.51
0.64
0.66
0.34
0.10
0.48
0.44
0.53
0.38
0.35
0.18
0.12
0.23
0.22
0.71
0.48
0.66
0.53
0.29
0.31
ÿ0.01
0.20
0.29
0.40
0.33
0.59
0.23
0.12
0.20
0.89
0.87
0.17
0.42
0.28
0.26
0.14
0.36
0.07
0.07
ÿ0.10
ÿ0.21
ÿ0.47
ÿ0.17
ÿ0.15
ÿ0.02
0.09
ÿ0.15
ÿ0.13
ÿ0.08
0.13
ÿ0.19
ÿ0.08
ÿ0.02
ÿ0.09
0.29
0.04
0.56
0.02
0.11
0.32
0.28
0.19
0.49
ÿ0.06
0.08
0.20
0.23
0.16
0.02
0.20
0.12
0.82
0.88
0.12
0.40
0.43
0.48
0.18
0.57
0.87
0.73
0.05
ÿ0.15
ÿ0.26
28
28
ÿ0.63
0.63
0.53
0.65
0.38
ÿ0.03
0.32
0.54
ÿ0.18
ÿ0.17
16
44
ÿ0.28
0.56
0.58
0.23
0.48
0.22
0.43
0.09
ÿ0.12
ÿ0.20
15
59
0.47
ÿ0.08
ÿ0.04
ÿ0.12
ÿ0.27
ÿ0.06
ÿ0.22
0.46
0.89
0.79
9
68
ÿ0.12
0.17
0.22
ÿ0.03
0.15
0.05
0.21
0.06
0.02
0.27
8
76
4. Conclusions
The climax soils of Galicia, a region in the European temperate humid zone, are generally acid Umbrisols (ISSS Working Group RB, 1998) with high
organic matter contents. The values of their biochemical properties are in general within the ranges reported
in the soil literature for soils from other geographical
areas, and exhibit close mutual correlation, the chief
exceptions being indicative of a lack of coupling
between enzymes involved in the sulphur cycle and
those involved in the other nutrient element cycles.
The main determinants of the values of biochemical
properties in these soils are the activity of their microbial communities and the processes involved in the
accumulation and transformation of organic matter.
Acknowledgements
This work was ®nanced by the Xunta de Galicia.
The authors thank Ana Isabel Iglesias-Tojo for her
help with the analysis of the samples.
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Biochemical properties of acid soils under climax vegetation
(Atlantic oakwood) in an area of the European temperate±humid
zone (Galicia, NW Spain): speci®c parameters
C. Trasar-Cepeda a, M.C. LeiroÂs b,*, F. Gil-Sotres b
a
Departamento de BioquõÂmica del Suelo, Instituto de Investigaciones AgrobioloÂgicas de Galicia, Consejo Superior de Investigaciones Cientõ®cas,
Apartado 122, E-15080 Santiago de Compostela, Spain
b
Departamento de EdafologõÂa y QuõÂmica AgrõÂcola, Facultad de Farmacia, Universidad de Santiago de Compostela, E-15706 Santiago de
Compostela, Spain
Accepted 6 October 1999
Abstract
The general and speci®c biochemical parameters of soils are highly sensitive to disturbance of the environment, but their use
for diagnosis of soil degradation is limited by lack of comparable published data and lack of accepted methodological standards.
With a view to establishing an appropriate data base for the soils of Galicia (NW Spain), we investigated the biochemical
properties of the O and Ah horizons of 40 native Umbrisols under climax Atlantic oakwood in this region. We report here our
results on speci®c biochemical parameters (i.e. extracellular hydrolytic enzyme activities) characterizing the phosphorus, nitrogen,
carbon and sulphur cycles. The enzymes studied were phosphomonoesterase (23.51 2 10.37 and 6.62 2 3.29 mmol p-nitrophenol
gÿ1 hÿ1, values for O and Ah horizons, respectively), phosphodiesterase (3.60 2 1.95 and 0.96 2 0.51 mmol p-nitrophenol gÿ1
hÿ1), casein-protease (2.97 2 0.83 and 0.94 2 0.32 mmol tyrosine gÿ1 hÿ1), BAA-protease (23.74 2 11.35 and 15.26 2 8.91 mmol
NH3 gÿ1 hÿ1), urease (24.90 2 13.60 and 16.59 2 10.61 mmol NH3 gÿ1 hÿ1), CM-cellulase (0.59 2 0.17 and 0.23 2 0.10 mmol
glucose gÿ1 hÿ1), invertase (12.6622.75 and 6.9322.14 mmol glucose gÿ1 hÿ1), b-glucosidase (8.4325.14 and 1.5520.89 mmol
p-nitrophenol gÿ1 hÿ1), and arylsulfatase (0.672 0.30 and 0.46 2 0.20 mmol p-nitrophenol gÿ1 hÿ1). For the variables for which
comparable data are available, the values observed are generally within previously published ranges. Principal components
analysis of the combined biochemical, physical and chemical data for these soils shows ®ve factors, of which the three most
important concern microbial activity and its logical dependence on nutrient content, the accumulation of soil organic matter and
the mineralization of soil organic matter. 7 2000 Elsevier Science Ltd. All rights reserved.
Keywords: Soil biochemical properties; Soil enzymes; Temperate forest soils
1. Introduction
It is widely accepted that soil quality can be
expressed in terms of the capability of a soil to accept,
store and recycle the water, minerals and energy
required for optimal crop production while at the
* Corresponding author. Tel.: +34-981-563-100, ext. 15042; fax:
+34-981-594-912.
E-mail address: [email protected] (M.C. LeiroÂs).
same time preserving a healthy environment (Arshad
and Cohen, 1992; Parr et al., 1992). Thus good-quality
soils should carry out the following functions: crop
production, ®ltration and degradation. The most important of these three functions for the evaluation of
soil quality is perhaps degradation, as it de®nes the capacity of a soil to mineralize soil organic matter and
degrade exogenous plant material and anthropogenic
inputs such as organic wastes, pesticides, hydrocarbons, etc. (Dick, 1997). The degradation function
0038-0717/00/$ - see front matter 7 2000 Elsevier Science Ltd. All rights reserved.
PII: S 0 0 3 8 - 0 7 1 7 ( 9 9 ) 0 0 1 9 6 - 0
748
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
depends to a large extent on the biochemical properties
of the soil (Visser and Parkinson, 1992), which are
usually divided into two groups (Nannipieri et al.,
1995): general biochemical parameters, which are
directly related to the number and activity of soil
microorganisms, and speci®c parameters, the activities
of extracellular hydrolytic enzymes stabilized by being
bound in complexes with clay minerals and humic colloids.
Hydrolytic extracellular soil enzymes make nutrients
available to plants and microorganisms by converting
them from unassimilable to readily assimilable forms
(Saratchandra et al., 1984). However, although this
implies that they exert a very important part in the
degradation function, this potential for characterising
soil quality has often been ignored. This is partly due
to there having been relatively few studies in which the
activities of a large number of enzymes have been studied simultaneously, and partly to the diculty in
comparing data obtained for dierent soils, by dierent workers using dierent incubation conditions, substrates and substrate concentrations and buer systems
(Saratchandra et al., 1984). What is needed, if the potential of enzymatic activities and other biochemical
properties for soil quality assessment is to be realized,
is an extensive database of the biochemical properties
of soils from dierent parts of the world. Moreover,
since the goal of sustainable development implies that
soil quality should be determined with reference to
undisturbed native soils (Doran et al., 1994), the compilation of such a database should give priority to climax soils in so far as is possible.
The above considerations have led us to compile an
extensive database of the biochemical properties of climax soils in Galicia (NW Spain), a region situated in
the European temperate±humid zone in which extensive areas of climax vegetation (Atlantic oakwood) still
exist. The general biochemical parameters of these climax soils (all Umbrisols; ISSS Working group RB,
1998) were reported by LeiroÂs et al. (1999). Here we
report the activities of a variety of hydrolytic enzymes
involved in the carbon, nitrogen, phosphorus and sulphur cycles, including enzymes with substrates of both
high and low molecular mass, and we discuss their relationships with each other and with the physical,
chemical and general biochemical parameters of the
soils.
2. Material and methods
2.1. Soils
We studied 40 soils developed under climax vegetation dominated by Quercus robur L. or Q. pyrenaica
L. at sites distributed throughout Galicia, NW Spain.
At all sites, the sampling area displayed little if any
disturbance of human origin, and its tree vegetation
was composed mainly of healthy adult specimens. All
the soil are Umbrisols (ISSS Working Group RB,
1998); their locations, the methods used for sampling
and for determination of their physical, chemical and
general biochemical parameters, and the values of
these parameters, have been described by LeiroÂs et al.
(1999).
2.2. Analytical methods
The activities of urease (EC 3.5.1.5) and of proteases
(EC 3.4.4) hydrolysing benzoylargininamide (BAAprotease) and casein (casein-protease) were determined
as described by Gil-Sotres et al. (1992). Brie¯y, urease
activity was determined using urea as substrate, incubating for 1.5 h at 378C and pH 7.1 (phosphate buer
0.2 M) and measuring the NH+
4 released with an ammonium electrode. BAA-protease activity was determined using the same incubation conditions and the
same method to determine NH+
4 but with a-benzoylN-argininamide (BAA) as substrate. In both cases enzymatic activity is expressed in mmol NH3 gÿ1 hÿ1.
Casein-hydrolysing activity was determined with casein
as substrate, incubating for 2 h at 508C and pH 8.1
(Tris±HCl buer 0.05 M) and determining the amino
acids released by the Folin colorimetric method; enzymatic activity is expressed in mmol tyrosine gÿ1 hÿ1.
Acid phosphomonoesterase (EC 3.1.3.2), b-glucosidase (EC 3.2.1.21), phosphodiesterase (EC 3.1.4.1) and
arylsulfatase (EC 3.1.6.1) activities were determined by
incubating the soils with a substrate containing a pnitrophenyl
moiety
and
spectrophotometrically
measuring the amount of p-nitrophenol liberated by
enzymatic hydrolysis. In these cases the enzymatic activity is expressed in mmol p-nitrophenol gÿ1 hÿ1. Acid
phosphomonoesterase activity was determined with pnitrophenyl phosphate as substrate, incubating at pH
5.0 (Modi®ed Universal Buer) and 378C. After 30
min 2 M CaCl2 was added (to stop the reaction and to
avoid the brown coloration caused by organic matter)
and the liberated p-nitrophenol was extracted with 0.2
M NaOH (Tabatabai and Bremner, 1969; Saa et al.,
1993). b-glucosidase activity was determined as
described for phosphomonoesterase activity except
that the substrate was p-nitrophenyl-b-glucopyranoside
and the p-nitrophenol released was extracted with
THAM±NaOH 0.1 M of pH 12 (Eivazi and Tabatabai, 1988). Phosphodiesterase activity was determined
with bis-p-nitrophenyl phosphate as substrate, incubating at pH 5.0 (THAM buer 0.05 M) and 378C for 1
h (Bowman and Tabatabai, 1978). Arylsulphatase activity was determined with p-nitrophenyl sulphate as
substrate, incubating at pH 5.8 (acetate buer 0.5 M)
and 378C for 1 h (Tabatabai and Bremner, 1970).
Table 1
Values of the speci®c biochemical parameters studied, in O horizons n 40 of climax soils under oakwood in Galicia (NW Spain)
Total C (%)
Total N (%)
Phosphomonoesterasea
Phosphodiesterasea
Arylsulfatasea
Casein-proteaseb
BAA-proteasec
Ureasec
CM-cellulased
b-glucosidasea
Invertased
7.1
8.1
10.1
12.1
13.1
14.1
15.1
16.1
17.1
18.1
19.1
20.1
22.1
23.1
24.1
25.1
27.1
28.1
29.1
31.1
33.1
34.1
35.1
36.1
37.1
39.1
40.1
41.1
42.1
43.1
44.1
45.1
46.1
47.1
48.1
49.1
50.1
51.1
52.1
53.1
31.2
28.6
14.6
21.2
21.0
18.3
17.3
27.8
23.1
17.8
32.3
26.9
18.2
50.0
46.2
37.8
19.2
36.2
32.4
40.0
28.5
31.0
39.8
39.2
26.6
35.7
36.5
39.3
31.5
39.6
33.4
38.8
26.6
31.0
36.8
38.6
34.5
19.4
25.3
28.8
1.57
1.76
0.95
1.19
1.16
0.97
1.07
1.51
1.29
1.10
1.57
1.56
1.52
1.92
2.07
1.60
1.02
1.66
1.92
2.17
1.50
1.83
1.76
1.83
1.51
1.71
1.83
1.95
1.59
1.86
1.58
1.81
1.26
1.75
1.88
1.80
1.68
0.95
0.77
1.28
10.49
40.36
9.17
24.59
12.75
36.91
9.47
30.03
10.79
12.38
19.20
15.51
11.93
16.74
24.05
18.17
11.76
15.90
9.35
14.38
33.89
24.29
38.19
20.78
27.47
19.20
34.40
35.49
12.47
30.68
47.45
34.02
32.84
18.52
34.95
31.97
33.03
25.59
21.22
30.00
1.65
4.33
1.21
1.72
3.50
2.73
2.37
2.90
2.58
2.22
7.44
3.88
2.36
3.61
4.81
3.08
3.47
2.53
2.09
3.62
5.56
1.67
2.49
3.02
5.20
4.02
8.01
5.48
2.16
2.91
3.02
11.82
3.15
3.17
3.19
3.06
4.55
3.10
3.40
3.05
0.20
1.03
0.47
0.27
0.67
0.60
0.88
0.70
0.80
0.47
0.69
0.73
0.41
0.53
0.83
0.35
0.53
0.34
0.29
0.22
0.86
0.72
0.55
0.40
1.12
1.58
0.61
1.06
0.52
0.62
0.56
0.67
0.63
1.50
0.65
0.53
0.81
0.81
0.90
0.50
2.76
4.04
2.88
3.66
3.54
2.84
3.15
4.33
3.47
2.43
4.11
3.71
2.69
4.48
4.08
3.04
1.52
1.78
1.15
2.43
3.60
2.13
2.17
3.99
2.41
1.15
3.81
3.30
2.96
2.90
2.58
2.09
1.85
3.50
3.19
3.32
2.83
3.16
2.85
3.02
11.93
18.33
20.54
8.90
29.90
24.98
30.86
13.02
36.21
18.22
39.14
48.97
29.85
23.81
24.99
22.23
21.87
14.12
5.91
10.07
34.48
26.17
10.47
21.99
41.57
19.30
42.61
14.67
28.81
12.52
27.88
16.92
33.05
23.66
40.05
5.84
44.93
16.17
8.13
26.44
12.66
11.37
18.34
13.89
28.27
16.44
28.50
11.43
29.31
19.62
19.80
40.10
28.18
34.57
24.12
30.71
14.47
16.94
12.04
9.35
29.29
38.70
22.68
31.77
65.60
20.15
35.19
20.08
20.81
14.52
66.26
20.01
33.03
21.00
33.57
10.81
47.12
15.12
1.76
28.32
0.62
0.88
0.31
0.54
0.46
0.51
0.30
0.83
0.45
0.58
0.71
0.54
0.70
0.88
0.93
0.75
0.27
0.57
0.31
0.40
0.67
0.80
0.78
0.74
0.50
0.56
0.56
0.41
0.48
0.79
0.68
0.64
0.52
0.44
0.53
0.86
0.61
0.52
0.51
0.53
7.45
15.26
5.31
6.71
4.36
7.31
4.43
11.64
9.05
6.56
2.05
7.13
5.89
13.86
8.20
12.05
4.00
6.42
4.23
7.07
11.41
6.29
5.68
5.08
4.93
6.98
4.21
29.63
3.77
17.09
17.00
4.34
8.76
14.15
13.79
9.04
7.61
7.73
3.83
6.71
12.44
15.56
11.21
12.40
9.39
14.31
11.48
16.92
13.41
13.76
15.77
12.93
14.59
16.20
16.40
14.35
13.39
18.83
10.19
10.41
13.70
15.44
16.93
17.72
11.52
9.03
8.47
10.08
10.19
12.96
11.33
9.62
9.36
12.19
9.95
12.81
11.31
7.92
10.64
11.28
Mean
S.D.
30.5
8.6
1.54
0.35
23.51
10.37
3.60
1.95
0.67
0.30
2.97
0.83
23.74
11.35
24.90
13.60
0.59
0.17
8.43
5.14
12.66
2.75
mmol
mmol
c
mmol
d
mmol
p-nitrophenol gÿ1 hÿ1.
tyrosine gÿ1 hÿ1.
NH3 gÿ1 hÿ1.
glucose gÿ1 hÿ1.
749
a
b
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Sample
750
Table 2
Values of the speci®c biochemical parameters studied, in Ah horizons n 40 of climax soils under oakwood in Galicia (NW Spain)
Total C (%)
Total N (%)
Phosphomonoesterasea
7.2
8.2
10.2
12.2
13.2
14.2
15.2
16.2
17.2
18.2
19.2
20.2
22.2
23.2
24.2
25.2
27.2
28.2
29.2
31.2
33.2
34.2
35.2
36.2
37.2
39.2
40.2
41.2
42.2
43.2
44.2
45.2
46.2
47.2
48.2
49.2
50.2
51.2
52.2
53.2
10.9
10.0
5.7
9.5
6.3
5.2
7.8
12.3
8.3
6.6
12.3
9.5
8.9
14.5
9.5
12.5
11.1
9.4
14.8
11.6
10.3
14.6
12.2
18.3
11.8
11.5
6.6
13.1
9.7
10.2
7.8
15.1
14.7
9.3
13.2
10.1
15.2
6.7
7.9
15.1
0.61
0.70
0.39
0.51
0.37
0.32
0.56
0.76
0.55
0.40
0.65
0.57
0.59
0.81
0.68
0.57
0.54
0.44
0.93
0.79
0.68
0.96
0.77
0.99
0.85
0.74
0.40
0.79
0.63
0.63
0.43
0.85
0.76
0.72
0.92
0.63
0.98
0.41
0.48
0.69
3.25
8.64
2.23
6.72
2.43
7.87
2.73
6.95
4.10
3.99
6.56
2.24
3.20
4.64
5.90
3.12
4.33
3.50
6.04
5.26
9.97
8.99
8.46
3.76
12.57
8.98
6.76
12.04
3.70
6.64
6.93
13.82
11.31
6.23
6.66
8.22
8.64
6.75
5.00
15.76
Mean
S.D.
10.8
3.1
0.65
0.18
6.62
3.29
a
mmol
mmol
c
mmol
d
mmol
b
p-nitrophenol gÿ1 hÿ1.
tyrosine gÿ1 hÿ1.
NH3 gÿ1 hÿ1.
glucose gÿ1 hÿ1.
Phosphodiesterasea
Arylsulfatasea
Casein-proteaseb
BAA-proteasec
Ureasec
CM-cellulased
b-glucosidasea
Invertased
0.45
0.97
0.40
0.56
0.64
1.16
0.81
0.76
1.12
0.72
1.68
0.97
0.92
1.62
1.96
0.44
1.02
0.63
0.84
0.86
0.84
0.72
0.88
0.54
1.24
1.23
0.62
2.28
0.82
0.86
0.38
0.30
1.56
1.02
0.76
0.88
0.83
0.85
0.69
2.69
0.14
0.68
0.21
0.07
0.42
0.25
0.54
0.31
0.63
0.25
0.66
0.38
0.37
0.56
0.46
0.07
0.58
0.34
0.41
0.35
0.56
0.51
0.44
0.23
0.91
0.43
0.54
0.85
0.41
0.56
0.33
0.52
0.70
0.85
0.43
0.27
0.65
0.67
0.57
0.45
0.83
1.19
0.95
1.44
0.63
1.15
1.34
1.67
1.19
0.84
0.62
0.61
0.66
0.91
0.96
0.36
0.71
0.69
0.87
0.51
0.69
0.68
0.64
1.02
1.17
1.13
0.78
1.40
0.99
1.28
0.72
0.70
0.84
1.75
0.66
1.40
0.88
1.12
0.93
0.82
3.63
6.75
16.06
3.73
15.94
5.50
15.86
7.10
25.44
8.33
25.60
28.96
25.86
21.94
13.16
5.03
14.74
8.52
13.80
5.88
23.44
12.31
12.43
8.02
27.64
14.73
11.69
8.80
24.81
12.11
13.21
14.78
30.10
18.69
33.21
2.67
34.33
8.36
4.48
22.79
7.38
5.94
10.21
5.96
14.53
6.59
17.14
10.33
19.94
10.55
26.08
22.50
18.43
30.03
12.83
6.64
6.99
3.92
18.12
10.28
17.74
28.50
20.33
23.24
47.14
9.47
6.46
13.53
15.47
13.67
49.78
11.98
28.56
18.59
26.58
3.17
25.27
14.58
3.60
21.36
0.26
0.33
0.06
0.25
0.10
0.09
0.11
0.26
0.11
0.15
0.23
0.18
0.18
0.26
0.36
0.22
0.06
0.21
0.32
0.26
0.19
0.47
0.33
0.24
0.19
0.30
0.17
0.22
0.06
0.33
0.22
0.33
0.39
0.13
0.28
0.42
0.14
0.16
0.14
0.34
1.13
1.92
1.02
1.09
0.77
1.48
1.16
2.36
1.63
1.20
1.71
3.86
0.93
1.42
2.72
1.03
0.95
1.41
1.46
1.12
0.94
1.62
1.22
0.67
1.10
1.98
1.07
1.97
0.73
1.72
0.77
1.56
4.58
2.02
1.58
0.95
0.89
1.45
0.83
4.06
5.96
8.67
4.75
5.76
7.29
4.42
9.27
5.36
10.95
7.22
7.53
7.49
6.69
8.47
4.57
2.00
9.37
9.12
8.58
9.91
8.35
5.66
2.97
4.43
10.89
6.49
3.75
7.85
9.85
4.46
6.50
6.82
8.76
8.02
6.87
5.58
8.43
5.53
7.64
4.91
0.96
0.51
0.46
0.20
0.94
0.32
15.26
8.91
16.59
10.61
0.23
0.10
1.55
0.89
6.93
2.14
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Sample
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Invertase (EC 3.2.1.26) activity was determined with
saccharose as substrate, incubating for 3 h at 508C and
pH 5.5 (acetate buer 2 M) and determining reducing
sugars as per Schinner and von Mersi (1990). Carboxymethylcellulase (CM-cellulase) activity was determined
similarly, except that the substrate was carboxymethylcellulose and the incubation time was 24 h (Schinner
and von Mersi, 1990). The enzymatic activities are
expressed in mmol glucose gÿ1 hÿ1.
2.3. Expression and analysis of results
All determinations were performed in triplicate, and
all values reported are averages of triplicate determinations expressed on an oven-dried soil basis (1058C).
Statistical analyses were performed using Statistics 4.5
for Windows (StatSoft Inc., 1993)
3. Results and discussion
The values, means and standard deviations of the
enzyme activities in the O and Ah horizons are listed
in Tables 1 and 2.
3.1. Enzymatic activities
3.1.1. Acid phosphomonoesterase
Mean acid phosphomonoesterase activity in the O
horizons was 23.51 mmol p-nitrophenol gÿ1 hÿ1 (S.D.
10.37, range 9.17±47.45, coecient of variation CV
44%). In the Ah horizons it fell to 6.62 mmol p-nitrophenol gÿ1 hÿ1 (S.D. 3.29, range 2.23±15.76, CV
50%). Although phosphomonoesterase has been one
of the most extensively studied soil enzymes (Burns,
1978), data for the organic layers of woodland soils
are relatively scarce. The values observed in this study
lie within the range found in woodland soils in the
Apennines (Nannipieri et al., 1980), in subarctic
regions (Neal, 1982) and in Canada (Pang and
Kolenko, 1986). Similarly, the Ah horizon values
obtained in this study fall within the range reported in
the much more abundant literature on phosphomonoesterase in mineral horizons (Speir, 1977; Frankenberger and Dick, 1983).
3.1.2. Phosphodiesterase
Mean phosphodiesterase activity in the O horizons
was 3.60 mmol p-nitrophenol gÿ1 hÿ1 (S.D. 1.95, range
1.21±11.82, CV 54%), between six and seven times less
than phosphomonoesterase activity; in the Ah horizons
activity ranged from 0.30 to 2.69 mmol p-nitrophenol
gÿ1 hÿ1 (mean 0.96, S.D. 0.50, CV 52%). References
to phosphodiesterase activities in the literature are
scarce and generally unsystematic. Rastin et al. (1988,
1990) mention very low values for both soil layers
751
(between 0 and 0.02 mmol p-nitrophenol gÿ1 hÿ1). For
Ah horizons, Frankenberger and Dick (1983) reported
values similar to those observed in this work, but Ross
et al. (1995a,b) found much higher values of between 7
and 22 mmol p-nitrophenol gÿ1 hÿ1.
3.1.3. Arylsulfatase
Mean arylsulfatase activity was 0.66 mmol p-nitrophenol gÿ1 hÿ1 in the O horizons (S.D. 0.30, CV 45%,
range 0.20±1.58) and 0.45 mmol p-nitrophenol gÿ1 hÿ1
in the Ah horizons (S.D. 0.20, CV 43%, range 0.07±
0.91). Although the values observed in this study in
both the O and Ah horizons are within the range
reported in the literature for natural ecosystems, the
maximum values are considerably lower than those
found in organic layers by authors such as Baligar and
Wright (1991) and Saratchandra et al. (1984), who
reported maxima of 3.89 and 5.69 mmol p-nitrophenol
gÿ1 hÿ1, respectively. Although this discrepancy may
be partly due to methodological dierences, it suggests
that in native Galician soils the sulphur requirement of
soil microorganisms is satis®ed, at least in the top few
centimetres of the soil, with ensuing downregulation of
arylsulphatase. This implies the possibility of these
soils developing an arylsulfatase de®ciency, the consequences of which for the sulphur cycle are dicult to
predict.
3.1.4. Casein-protease
Casein-protease activity ranged from 1.15 to 9.08
mmol tyrosine gÿ1 hÿ1 in the O horizons (mean 3.10,
S.D. 1.26, CV 41%), and from 0.36 to 1.75 mmol tyrosine gÿ1 hÿ1 in the Ah horizons (mean 0.94, S.D. 0.32,
CV 43%). The values observed in Ah horizons are
within the range reported by several authors for native
soils (e.g. Nannipieri et al., 1980; BonmatõÂ et al., 1991;
Perucci, 1992). We have been unable to ®nd references
to casein-protease activity in O horizons; nevertheless,
the values of 5.50±38.67 mmol tyrosine gÿ1 hÿ1 found
by Dilly and Munch (1995) in rotting Alnus litter are
much higher than those found in this study. The
observed depthwise variation suggests that proteolysis
of substrates of high molecular weight diminishes as
the decomposition of plant debris advances, i.e. in the
order litter, O layer, humus, at least when the results
are expressed as activity per unit mass of soil.
3.1.5. BAA-protease
In the O horizons BAA-protease activity ranged
from 5.84 to 48.97 mmol NH3 gÿ1 hÿ1 (mean 23.50,
S.D. 11.54 CV 49%), and in Ah horizons from 2.67 to
34.33 mmol NH3 gÿ1 hÿ1 (mean 15.26, S.D. 8.91, CV
58%). No other values for temperate zone soils appear
to be available for comparison, in spite of the proposal
of Ladd and Buttler (1972) that BAA be used to determine the activity of proteases acting on small peptides.
1.00
1.00
0.48a
1.00
0.64a
0.59a
1.00
0.31b
0.28c
0.28c
1.00
0.71a
0.24d
0.22
0.34b
1.00
0.35b
0.24
0.76a
0.60a
0.44a
P < 0.001.
P < 0.01.
c
P < 0.02.
d
P < 0.05.
a
CM-cellulase
Urease
BAA-protease
Casein-protease
Arylsulfatase
1.00
0.35b
0.42a
0.32b
0.27c
0.42a
0.27c
1.00
0.45a
0.59a
0.43a
0.31b
0.61a
0.48a
0.39a
b
3.1.9. Invertase
Invertase activity ranged from 2.27 to 18.83 mmol
glucose gÿ1 hÿ1 in the O horizons (mean 12.06, S.D.
3.41, CV 28%) and from 2.00 to 10.95 mmol glucose
gÿ1 hÿ1 in the Ah horizons (mean 6.85, S.D. 2.22, CV
28%). Both sets of values are similar to those found
by Batistic et al. (1980), Frankenberger and Dick
(1983), Speir et al. (1984), Schinner and von Mersi
(1990) and Ross et al. (1995a,b).
1.00
0.69a
0.43a
0.61a
0.33b
0.40a
0.76a
0.73a
0.52a
3.1.8. b-Glucosidase
Mean b-glucosidase activity was 8.42 mmol p-nitrophenol gÿ1 hÿ1 in the O horizons (S.D. 5.14, range
2.05 to 29.63, CV 61%) and 1.55 mmol p-nitrophenol
gÿ1 hÿ1 in the Ah horizons (S.D. 0.89, range 0.67 to
4.58, CV 57%). The Ah horizon values can be considered normal, but the O layer values are quite high
in comparison with those found by Batistic et al.
(1980), Kanazawa and Miyashita (1987) and Deng and
Tabatabai (1996b).
Phosphodiesterase
3.1.7. CM-cellulase
In the O horizons CM-cellulase activity ranged from
0.27 to 0.93 mmol glucose gÿ1 hÿ1 (mean 0.59, S.D.
0.17, CV 29%), and in the Ah horizons from 0.06 to
0.47 mmol glucose gÿ1 hÿ1 (mean 0.23, S.D. 0.10, CV
43%). The Ah horizon values are comparable with
those reported in the literature, but those observed in
O horizons are slightly lower than reported values
(Kanazawa and Miyashita, 1987; Ohtonen, 1994).
Table 3
Pearson coecients of pairwise correlation among the enzymatic activities studied (the analysis pools results from both O and Ah horizons)
3.1.6. Urease
Urease activity ranged from 1.76 to 66.26 mmol NH3
gÿ1 hÿ1 in O horizons (mean 24.90, S.D. 13.60, CV
55%) and from 3.17 to 49.78 mmol NH3 gÿ1 hÿ1 in Ah
horizons (mean 16.58, S.D. 10.67, CV 64%). Although
urease, like phosphomonoesterase, has been one of the
most extensively studied soil enzymes (Burns, 1978),
most published work refers to agricultural soils, urea
being one of the most widely used nitrogenated fertilizers in many parts of the world. Research on urease
activity in woodland soils has been relatively scant.
The values observed in this study are similar to those
found by Nannipieri et al. (1980) in soils from the
Apennine Mountains, but between 2 and 20-times
higher than those reported by Speir (1977), Speir et al.
(1980), Saratchandra et al. (1984) and Deng and Tabatabai (1996a).
b-glucosidase
It may be worth mentioning that the values reported
for three soils from the Apennine Mountains by Nannipieri et al. (1980) are within the range found in this
work.
Phosphomonoesterase
Phosphodiesterase
Arylsulfatase
Casein-protease
BAA-protease
Urease
CM-cellulase
b-glucosidase
Invertase
Invertase
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Phosphomonoesterase
752
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
3.2. Correlations between speci®c biochemical
parameters
As in the companion study of the general biochemical parameters of these soils, and for the same reasons
as were discussed by LeiroÂs et al. (1999), correlations
among dierent biochemical properties were sought
considering the data for the O and Ah horizons
jointly. Almost all the activities reported in this paper
are signi®cantly correlated with each other (P < 0.001
in most cases; see Table 3). As in related studies (BonmatõÂ et al., 1991; Tate et al., 1991; Perucci, 1992), the
enzyme correlating best with other enzymes is phosphomonoesterase, which correlates especially closely
with CM-cellulase and b-glucosidase, followed by
phosphodiesterase and casein-protease. Since phosphodiesterase is also closely correlated with casein-protease
and CM-cellulase, there is clearly a strong interrelationship among enzymatic processes involved in
the carbon, nitrogen and phosphorus cycles. By contrast, arylsulfatase is not very highly correlated with
any other enzymatic activity; this apparent lack of
coupling between the sulphur cycle and the other
major nutrient cycles, which contrasts with the results
of Speir et al. (1980, 1984), Frankenberger and Dick
(1983) and Saratchandra et al. (1984), may be due to
the downregulation hypothesized above.
Among the nitrogen cycle enzymes there is close correlation between urease and BAA-protease, but neither
of these two enzymes correlates highly with casein-protease, which contrasts with the ®ndings of Speir et al.
(1980). This pattern suggests that in Galician soils the
degradation of proteins and the degradation of smaller
nitrogenated compounds such as peptides and urea are
subject to dierent regulatory mechanisms.
Among the carbon cycle enzymes, CM-cellulase is
highly correlated with b-glucosidase and invertase, and
these latter are also moderately well correlated with
each other. This re¯ects the synergism among these
enzymes in the degradation of the carbon compounds
received by the soil (Panda and Sharma, 1994; Deng
and Tabatabai, 1996a,b).
Similarly, the phosphorus cycle enzymes phosphomonoesterase and phosphodiesterase are likewise closely correlated, which is in keeping with the ®ndings of
Frankenberger and Dick (1983) and Rastin et al.
(1988) and shows the coupling between the enzymes of
this cycle.
3.3. Relationships within chemical, physical and
biochemical properties
The enzyme activities we measured also correlate
closely with physical and chemical properties related
to the availability of water and nutrients (results
not shown) and with many of the general biochemi-
753
cal parameters discussed by LeiroÂs et al. (1999). To
clarify the structure of these interdependences, we
performed a joint principal components analysis
(PCA) of our data on the physical, chemical and
biochemical properties of the O and Ah layers of
these soils, including both general and speci®c biochemical parameters. The ®ve main factors identi®ed
together account for 76% of the variance; the loadings of the 27 soil characteristics considered on
each factor are listed in Table 4.
Factor I, which accounts for 28% of the total
variance, exhibits close positive correlation with dehydrogenase, microbial biomass C and available
Ca2+ and K+, and somewhat lower positive correlation with ATP, microbial biomass N, catalase, invertase, casein-protease and available P, and may thus be
regarded as related to the size and activity of the microbial community. The presence of available elements
among its de®ning variables may be attributed to the
logical dependence of microbial activity on nutrient
contents. The high loadings of invertase and caseinprotease suggest that, contrary to what has been
suggested by Skujins (1978), Nannipieri et al. (1982)
and Speir and Ross (1990), the activities of both these
enzymes depend more on the microbial community
than on their extracellular accumulation, in these soils
at least.
Factor II accounts for 16% of the total variance.
At its positive pole it is de®ned mainly by phosphomonoesterase, although CM-cellulase, the C-to-N
ratio and total C and N contents also have high
loadings, and at its negative pole by pH. Though it
is not easy to interpret, these associations suggest
that it is related to the accumulation of hydrolytic
enzymes and of organic matter that is poorly humidi®ed because of the acidity of the medium. The
relationship of phosphomonoesterase and CM-cellulase to the accumulation of organic matter has been
mentioned by Harrison (1983) and Sinsabaugh and
Linkins (1988), among others.
Factor III accounts for 15% of the total variance.
Its main de®ning property is nitrogen mineralization
capacity as shown by total inorganic N, although soil
respiration and ammoni®cation also contribute to it.
Together with the relatively high loadings of the total
N and C contents, these properties suggest that Factor
III represents the mineralization of organic matter.
Factor IV, which accounted for only 9% of the total
variance, may be regarded as re¯ecting the nature and
weathering of the parent material, being de®ned by Al
and Fe oxide contents. Factor V is de®ned by BAAprotease and urease; though it accounts for only 8%
of the total variance, its emergence as a separate factor
con®rms the independence of the degradation of low
molecular weight nitrogenated compounds, in keeping
with the correlation results discussed above.
754
C. Trasar-Cepeda et al. / Soil Biology & Biochemistry 32 (2000) 747±755
Table 4
Loadings of biochemical, physical and chemical properties on the axes identi®ed by principal components analysis of their values in the O and
Ah horizons of climax soils under oakwood in Galicia (NW Spain)
Factor I
Biochemical properties
Microbial biomass-C
Microbial biomass-N
Evolved CO2-C
ATP
Dehydrogenase
Catalase
N mineralized (N-NH+
4 )
N mineralized (N inorganic total)
Phosphomonoesterase
Phosphodiesterase
CM-cellulase
b-glucosidase
Invertase
Casein-protease
BAA-protease
Urease
Arylsulfatase
Physical and chemical properties
pH KCl
Total C
Total N
C/N
Labile Pi
Available Ca2+
Available K+
Available water
Al2O3
Fe2O3
Explained variance (%)
Accumulated explained variance (%)
Factor II
Factor II
Factor IV
Factor V
0.70
0.61
0.37
0.64
0.84
0.66
0.24
0.34
0.48
0.41
0.50
0.51
0.64
0.66
0.34
0.10
0.48
0.44
0.53
0.38
0.35
0.18
0.12
0.23
0.22
0.71
0.48
0.66
0.53
0.29
0.31
ÿ0.01
0.20
0.29
0.40
0.33
0.59
0.23
0.12
0.20
0.89
0.87
0.17
0.42
0.28
0.26
0.14
0.36
0.07
0.07
ÿ0.10
ÿ0.21
ÿ0.47
ÿ0.17
ÿ0.15
ÿ0.02
0.09
ÿ0.15
ÿ0.13
ÿ0.08
0.13
ÿ0.19
ÿ0.08
ÿ0.02
ÿ0.09
0.29
0.04
0.56
0.02
0.11
0.32
0.28
0.19
0.49
ÿ0.06
0.08
0.20
0.23
0.16
0.02
0.20
0.12
0.82
0.88
0.12
0.40
0.43
0.48
0.18
0.57
0.87
0.73
0.05
ÿ0.15
ÿ0.26
28
28
ÿ0.63
0.63
0.53
0.65
0.38
ÿ0.03
0.32
0.54
ÿ0.18
ÿ0.17
16
44
ÿ0.28
0.56
0.58
0.23
0.48
0.22
0.43
0.09
ÿ0.12
ÿ0.20
15
59
0.47
ÿ0.08
ÿ0.04
ÿ0.12
ÿ0.27
ÿ0.06
ÿ0.22
0.46
0.89
0.79
9
68
ÿ0.12
0.17
0.22
ÿ0.03
0.15
0.05
0.21
0.06
0.02
0.27
8
76
4. Conclusions
The climax soils of Galicia, a region in the European temperate humid zone, are generally acid Umbrisols (ISSS Working Group RB, 1998) with high
organic matter contents. The values of their biochemical properties are in general within the ranges reported
in the soil literature for soils from other geographical
areas, and exhibit close mutual correlation, the chief
exceptions being indicative of a lack of coupling
between enzymes involved in the sulphur cycle and
those involved in the other nutrient element cycles.
The main determinants of the values of biochemical
properties in these soils are the activity of their microbial communities and the processes involved in the
accumulation and transformation of organic matter.
Acknowledgements
This work was ®nanced by the Xunta de Galicia.
The authors thank Ana Isabel Iglesias-Tojo for her
help with the analysis of the samples.
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