Scientific African 4 (2019) e00107

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Scientific African
journal homepage: www.elsevier.com/locate/sciaf

Effect of biochar amendment on soil enzymatic activities,

carboxylate secretions and upland rice performance in a
sandy clay loam Alfisol of Southwest Nigeria
Segun O. Oladele a,b
a
Department of Agronomy, Adekunle Ajasin University, Akungba Akoko PMB 01, Ondo State, Nigeria
b
Department of Crop, Soil and Pest Management, Federal University of Technology, Akure PMB 704, Ondo State, Nigeria

a r t i c l e i n f o a b s t r a c t

Article history: Field studies were conducted over two years to determine changes in soil biochemical
Received 17 January 2019 properties and rain-fed upland rice yield as influenced by biochar amendment. Treatments
Revised 23 May 2019
consisted of incorporation of rice husk biochar at different rates (B1 = 0, B2 = 3 t ha−1 ,
Accepted 24 June 2019
B3 = 6 t ha−1 , and B4 = 12 t ha−1 ). Soil enzymes viz invertase, alkaline phosphatase, ure-
ase, and catalase, were significantly (P ˂ 0.05) higher under the highest application rate
Keywords: of biochar (12 t ha−1 ). Higher rates of enzyme activities were recorded within the top soil
Alfisol layer (0–0.1 m). Soil organic carbon, available nitrogen, Bray 1 phosphorus, total nitrogen
Biochar and soil moisture content were significantly (P ˂ 0.05) increased upon the application of
Carboxylates biochar at a rate of 12 t ha−1 in the top 0–0.1 m layer. Similarly, application of biochar at
Oryza sativa 12 t ha−1 significantly (P < 0.05) increased rice rhizospheric carboxylate secretions in the
Soil enzymes
top 0–0.1 m layer. Carboxylate exudates were increased in the order of citrate ˃ malate
˃ acetate ˃ oxalate, while increasing soil layer reduced carboxylate secretions. In general,
biochar application improved soil enzyme activities, rhizospheric secretions involved in C,
N, P cycling and nutrient solubilization which influence soil nutrient retention, availability
and crop performance. Biochar application at 3–6 t ha−1 significantly (P ˂ 0.05) increased
rice grain yield by 46% and straw yield by 47%.
© 2019 The Author. Published by Elsevier B.V. on behalf of African Institute of
Mathematical Sciences / Next Einstein Initiative.
This is an open access article under the CC BY license.
(http://creativecommons.org/licenses/by/4.0/)

Introduction

The excessive use of inorganic fertilizers has been shown to have detrimental effect on soil health and quality [1]. For
instance, excessive use of nitrogen (N) fertilizer to produce high yielding and nutrient demanding cereal crops has been
associated with a decline in soil organic matter [2]. A study by Prieto-Fernández et al. [3], documented that slash and
burn tillage system commonly practiced by small scale farmers in the tropics, and crop residues export cause significant
losses of soil organic matter and crop nutrients. Thus, in situ conversion of crop residues generated from harvest into a
stable carbon dense material such as biochar for incorporation into soil could be an alternative management practice to
mitigate such losses [4–6]. Biochar is produced by the thermal degradation (pyrolysis) of organic biomass under oxygen-
limiting conditions with the sole intent for use as soil conditioner [6,7]. It is a recalcitrant-carbon rich material with a large

E-mail address: segun.oladele@aaua.edu.ng

https://doi.org/10.1016/j.sciaf.2019.e00107
2468-2276/© 2019 The Author. Published by Elsevier B.V. on behalf of African Institute of Mathematical Sciences / Next Einstein Initiative. This is an open
access article under the CC BY license. (http://creativecommons.org/licenses/by/4.0/)
2 S.O. Oladele / Scientific African 4 (2019) e00107

surface area, highly aromatic structure and dominated by effective functional groups [8]. Biochar has considerable potential
to improve soil quality and store carbon in soils due to its unique properties, such as liming (i.e., increase in soil pH), high
soil organic carbon (Corg ) nutrient and water retention and stimulatory effect on microbial activities [9,10].
Soil enzymes are involved in many biochemical processes in soils, such as soil organic matter (SOM) mineralization and
other biogeochemical cycling of nutrients [11]. These enzymes are sensitive indicators to changes in soil functioning and
show a strong linear relationship with SOM forms and quality [12]. Cellobiosidase is responsible for the decomposition of
cellulose [13], dehydrogenase activity influences soil quality and microbial respiratory activity [14] while Polyphenol oxi-
dase is reported to induce biodegradation of lignin and other phenolic compounds [15]. Furthermore, soil enzymes such as
β -glucosidase, urease, and phosphatase are strongly linked to carbon, nitrogen, and phosphorus cycling. The large specific
surface area of biochar helps to adsorb labile substrates with consequential effects on soil enzyme activity [16,17]. The in-
fluence of biochar on soil enzymatic activity mainly depends on the interaction of substrate and enzyme with biochar [16].
Sorption of substrates and extracellular enzymes to the functional groups on biochar surface may promote or limit enzy-
matic reaction [18]. Variable effect of biochar on soil enzymatic activities and associated soil chemical properties have been
reported; Park et al. [19] and Kumar et al. [20] reported positive effect of biochar on soil enzymes [14,21] and Lehmann et al.
[5] reported negative effects while Wu et al. [22] and Lammirato et al. [16] reported a non-biochar effect on soil enzymes.
However, literatures explaining the mechanisms behind these contrasting observations are scarce.
Changes in soil quality are strongly related with soil enzymatic activity which can serve as a useful indicator for sus-
tainable management of soil and environmental stability [23]. Amongst all soil quality indices, enzymes react rapidly to
changes in soil management and therefore remain a good indicator of soil biological change [24]. A study conducted by
Prieto-Fernández et al., [3] documented significant losses of SOM and soil nutrients caused by removal of crop residues and
non-incorporation of stable carbonized materials such as biochar. The return of soil conditioners such as biochar is expected
to affect microbial processes, which are involved in soil carbon and nitrogen transformations and enzymatic activities con-
trolling the availability of most nutrients required for crop growth and microbial metabolic demands [25,26]. Thus, it was
hypothesized that application of rice husk biochar would improve nutrient cycling, soil enzymatic activities and associated
soil chemical properties with consequential effects on rain-fed rice performance in a sub-humid tropical agro-ecological
zone. Therefore, the specific objective of this study was to determine the effects of different rates of biochar application on
soil nutrient levels and rates of enzymatic activities in an Alfisol and rain-fed rice performance in Southwest Nigeria.

Materials and methods

Site description and experimental design

The experimental site (7°20 N; 5°30 E, altitude – 370 m) is situated within the experimental station of the Department of
Crop, Soil and Pest Management, Federal University of Technology, Akure, Southwest Nigeria. The study area has a tropical
climate and is within the equatorial rain forest belt with annual mean temperature of 25.3 °C [27]. The mean annual precip-
itation at Akure is 1450 mm with most rainfall occurring from May to September. The soil is classified as Isohyperthermic
Typic Oxic-Paleustalf Alfisol according to the USDA soil classification system. The soil textural class is sandy clay loam, and
the proportion of sand, clay, and silt fraction in the top 15 cm of soil were 68.8%, 25.1% and 6.1%, respectively. The physical
and chemical properties of the initial soil (0–15 cm) in 2016 were as follows: pH = 4.9, bulk density = 1.63 g cm3 , soil wa-
ter holding capacity = 33%, EC = 1.5 dSm−1 , Corg = (3.7 g kg−1 ), available N = (15.6 mg kg−1 ), available P = (38.06 mg kg−1 ),
exchangeable K = (0.17 cmol kg−1 ) and total N = (0.41 g kg−1 ).
Field studies with four treatments were established in 2016 and repeated in 2017 to quantify soil enzymatic activities,
soil properties, and crop yield response to residual effect of biochar amendment in a rain-fed upland rice cropping system.
The experiment was set up in randomized complete block design (RCBD) with three replications. Biochar amendments were
arranged in the main plots, and soil depth in subplots, for studies on soil enzymatic activities and soil properties only.
The data for each year was analyzed separately. The straw and yield data of rain-fed upland rice were also analyzed using
time series with year as repeated and fixed factor. The rice husk biochar treatments were B1 = 0 t ha−1 , B2 = 3 t ha−1 ,
B3 = 6 t ha−1 , and B4 = 12 t ha−1 applied at two soil depths (0.1 and 0.2 m) in 2016 and 2017. The experimental field was
ploughed and harrowed before sowing. The area of each plot was 4 m2 (2.0 × 2.0 m) and soil samples were collected from
0.1 m to 0.2 m soil depth in each plot. The biochar amendments were incorporated once in 2016 and maintained for the
next cropping season in 2017. Nitrogen fertilization in form of urea was applied equally to all plots at 30 kg ha−1 every year
in two splits. There was no application of phosphorus (P) and potassium (K) fertilizer as initial soil test revealed adequate
levels. The commonly used upland rice cultivar (N-U-1/FARO 65) by local farmers in the study area was planted at a seeding
rate of 60 kg ha−1 on 24 July 2016, 16 July 2017, by dibbing at a row spacing of 0.25 × 0.25 m within and between plants.
Manual weeding was conducted as required during the field experiment each year.

Biochar

Rice husks collected from a local rice mill in Akure, Ondo State, Nigeria was used to produce biochar at 350 °C, using an
electric biochar reactor. Rice husk biochar production and characteristics were previously reported in [6].
 S.O. Oladele / Scientific African 4 (2019) e00107 3

Fig. 1. Effect of biochar application rates on soil organic carbon.

Error bars indicate 95% confidence interval for means.

Soil sampling and measurement

Soil samples were collected by a core sampler from 0 to 0.1 and 0.1 to 0.2 m from each plot immediately after rice
harvest. Five core samples were randomly collected and homogenized to form a composite sample. This sample was divided
into two parts; one was used for soil nutrients analyses and the other stored at 4 °C, and used for soil enzymes analyses.
Soil organic carbon (Corg ) was determined using the Walkley and Black method, total soil nitrogen was analyzed using the
Kjeldahl method. Soil available nitrogen was determined as described by Dorich and Nelson [28]. Soil available phosphorus
was determined using the Bray-1 method [29]. Soil enzymatic activities viz were determined as described by [30]. Soil cata-
lase, invertase, urease and alkaline phosphatase were expressed as mL 0.1 mol L−1 KMnO4 g−1 soil 20 min−1 , mg glucose
g−1 soil 24 h−1 , NH4 + –N g−1 soil 24 h−1 and phenol g−1 soil 2 h−1 , respectively. Carboxylate secretions in the soil were an-
alyzed by preparing a homogenized soil suspension with 50 mL of 0.2M CaCl2 . The carboxylate compounds such as acetate,
citrate, malate and oxalate concentrations were determined following protocols described by [31].

Grain and straw yield

Six central rows of rice in each plot were manually harvested for the determination of grain and straw yield at 12% grain
moisture content.

Statistical analysis

The general linear model (GLM) was used to determine the main effects and interactions of years and treatments on rice
yields with grain and straw yields as the dependent variables, and years and treatments as fixed factors. Analysis of variance
(ANOVA) was used for each soil biochemical variable by calculating the mean values for comparison of different treatments.
The means were compared by Tukey’s test at P < 0.05. The statistical analyses were performed among treatments on each
soil depth using MINITAB® 17th edition. Pearson’s correlation analysis was used to investigate the relations amongst soil
properties and soil enzymes.

Results

Soil organic carbon

The effects of rice husk biochar on Corg in the different soil depths are presented in Fig. 1. Results showed that increasing
application rates of biochar resulted in a linear increase in Corg content. Biochar application rate of 12 t ha−1 had the highest
Corg content that was significantly (P < 0.05) higher than other application rates in the soil depth of 0–0.1 m (Fig. 1).
However, within the 0.1–0.2 m depth, an inconsistent trend was observed wherein Corg was observed on plots treated with
biochar at 3 t ha−1 and 6 t ha−1 , respectively.

Soil available N and P

Fig. 2 shows the available nitrogen content in both soil depths. The effect of biochar amendment on soil available N was
statistically significant (P < 0.05) in both soil depths. Available N pools were generally low for all treatments in both soil
4 S.O. Oladele / Scientific African 4 (2019) e00107

Fig. 2. Effect of biochar application rates on soil available N.

Error bars indicate 95% confidence interval for means.

Fig. 3. Effect of biochar application rates on soil available P.

Error bars indicate 95% confidence interval for means.

depths. However, biochar application of 12 t ha−1 and 6 t ha−1 had the highest pools of available N in the 0–0.1 m and
0.1–0.2 m depths, respectively. Similar trend was observed in soil available P. Available P increased in the order of 0 ˃ 3 ˃
6 ˃ 12 t ha−1 in both 0–0.1 m and 0.1–0.2 m soil depths, respectively. Biochar application at 3 t ha−1 accumulated more
available P than the other treatments in the 0–0.1 m layer relative to the 0.1–0.2 m layer (Fig. 3).

Soil total N

The significant effects of biochar amendment at measured soil depths on soil total N are presented in Fig. 4. Results
showed no clear trend of corresponding increased soil total N with increasing doses of biochar amendment in years of
study at 0.1–0.2 m soil layer. However, biochar dose of 6 t ha−1 and 12 t ha−1 had higher pools of total N at this layer after
two years of study.

Soil moisture content

Analysis of variance revealed that biochar amendment was only significant (P < 0.05) at the highest dose of application
(12 t ha−1 ) at the soil layer of 0–0.1 m in the first year. Furthermore, SMC increased significantly (P < 0.05) across de-
termined soil depths. Increasing rates of biochar amendment increased SMC with increasing soil depths. SMC values were
 S.O. Oladele / Scientific African 4 (2019) e00107 5

Fig. 4. Effect of biochar application rates on soil total N.

Error bars indicate 95% confidence interval for means.

Fig. 5. Effect of biochar application rates on soil moisture content (%).

Error bars indicate 95% confidence interval for means.

increased by 11% and 35% at the 0–0.1 and 0.1–0.2 m soil depth respectively in the highest biochar application rate after
one-time biochar application (Fig. 5).

Soil urease activity

The effect of rice husk biochar amendment at measured soil depths on soil urease activity is presented in Fig. 6. Urease
activity was significantly higher in the treatment with the highest application rate of biochar (12 t ha−1 ) when compared
with other treatments. Furthermore, soil depths indicated significant differences amongst biochar amended treatments. Ure-
ase activity increased by 40% and 50% in the treatment with the highest rate of biochar application at soil depths of 0–0.1
and 0.1–0.2 m after two years of study, respectively. During the two-year study, urease activity was significantly increased
in the second year especially at the lower soil layer (0.1 ̶ 0.2 m).

Soil invertase activity

The effect of rice husk biochar amendment at measured soil depths on soil invertase activity is presented in Fig. 7. Soil
invertase showed similar trend in activity as earlier presented in urease. Invertase activity was significantly higher in the
highest rate of biochar application (12 t ha−1 ) across determined soil depths after two years of study. Invertase activity
6 S.O. Oladele / Scientific African 4 (2019) e00107

Fig. 6. Effect of biochar application rates on urease activity.

Error bars indicate 95% confidence interval for means.

Fig. 7. Effect of biochar application rates on invertase activity.

Error bars indicate 95% confidence interval for means.

increased by 9% at 0–0.1 m and by 14% at the 0.1–0.2 m soil depths. This suggests an increased activity of invertase at the
lower layer of the soil.

Soil phosphatase activity

Analysis of variance showing the effect of biochar amendment on soil alkaline phosphatase activity at measured soil
depths is presented in Fig. 8. An increased rate of phosphatase activity was recorded after two years of study at the 0–0.1 m
soil layer. The application rate of biochar at 12 t ha−1 had the highest level of phosphatase activity. Phosphatase activity in
the second year was 46% higher than the first year. This enzyme showed a declining trend with increasing soil layer after
rice harvest (Fig. 8).

Soil catalase activity

The effect of biochar amendment on catalase activity is presented in Fig. 9. Application of biochar at the rate of 12 t ha−1
increased catalase activity in all soil depth measured. Catalase activity was increased by 11% and 12% after two years of
study at soil depths of 0–0.1 and 0.1–0.2 m, respectively. This shows an increased catalase activity as the study progressed,
however, an increase in soil depth was accompanied by decreasing catalase activity (Fig. 9).
 S.O. Oladele / Scientific African 4 (2019) e00107 7

Fig. 8. Effect of biochar application rate on phosphatase activity.

Error bars indicate 95% confidence interval for means.

Fig. 9. Effect of biochar application rate on catalase activity.

Error bars indicate 95% confidence interval for means.

Rhizosphere carboxylate secretions

After two years of study, analysis of variance showed significant (P < 0.05) effect of biochar amendment on carboxylate
secretions (Fig. 10). At the soil layer of 0–0.1 m, increasing application rate of biochar was observed to increase carboxylate
secretions. Citrate (2570 μmol g−1 root DW), malate (1575 μmol g−1 root DW) and oxalate concentrations (270 μmol g−1
root DW) were increased in the order of 0 ˃ 3 ˃ 6 ˃12 t ha−1 , while acetate (650.25 μmol g−1 root DW) concentrations
was observed to have peaked at the biochar rate of 6 t ha−1 (Fig. 10(a)). Similar trend was recorded at the 0.1–0.2 m soil
depth (Fig. 10(b)), however, carboxylate secretions reduced with increasing soil depth. In general, carboxylate exudations as
increased by biochar amendment in this study were in the order of citrate ˃ malate ˃ acetate ˃ oxalate.

Grain and straw yield

There was no significant (P < 0.05) interaction detected between biochar amendments and years of study. However,
grain and straw yield of rice were significantly (P < 0.05) affected by main effects of biochar amendment and years of
study (Table 1). Results showed an increase in rice grain and straw yield in amended treatments when compared to the
8 S.O. Oladele / Scientific African 4 (2019) e00107

Fig. 10. (a) Effect of biochar application rate on carboxylate secretions. Error bars indicate 95% confidence interval for means.
(b) Effect of biochar application rate on carboxylate secretions. Error bars indicate 95% confidence interval for means.

Table 1
Effect of biochar amendment on grain and straw yield of rain-fed
rice.

Biochar (t ha−1 ) Grain yield (t ha−1 ) Straw yield (t ha−1 )

2016 2017 2016 2017

0 1.6a 1.2a 2.3a 1.7a

3 2.5b 3.9b 4.0b 5.4b
6 3.6c 3.8b 4.2b 5.3b
12 3.1c 3.4b 3.5c 4.7bc
P-value
∗ ∗
Biochar (B)
∗ ∗
Year (Y)
B∗ Y ns ns

Mean values within a column by different lower case letters are sig-
nificantly different (P < 0.05).
∗
Significant at (P < 0.01), ns – not significant.
 S.O. Oladele / Scientific African 4 (2019) e00107 9

Table 2
Correlation coefficients for relationships between select soil chemical properties and enzyme activities.

Parameters Catalase Invertase Urease Phosphatase SMC Available N Available P Corg Total N
∗ ∗∗ ∗∗ ∗∗
Catalase 1 0.81 0.96 0.98 0.98 0.73∗
Invertase 1 0.83∗ 0.95∗∗ 0.94∗∗ 0.98∗∗ 0.74∗
Urease 1 0.85∗ 0.97∗∗ 0.88∗ 0.89∗ 0.89∗
Phosphatase 1 0.78∗ 0.96∗∗ 0.98∗∗ 0.95∗∗ 0.85∗
SMC 1 ns ns ns ns
Available N 1 ns 0.78∗ 0.88∗
Available P 1 0.73∗ ns
Corg 1 0.89∗
Total N 1
∗
Significant at P ˂ 0.05.
∗∗
Significant at P ˂ 0.01.

control after two years of study. In the first year, rice grain and straw yield was increased in the treatment with biochar
application rate of 6 t ha−1 (Table 1). While, in the second year of study, an increase of 46% and 47% in grain and straw yield
respectively was recorded over values reported in the first year particularly in the treatment with biochar rate of 3 t ha−1 .
Though grain and straw yield were higher in biochar dose of 3 t ha−1 , values recorded were not significantly different from
that observed in 6 t ha−1 .

Relationships between soil properties and enzymes activity

Significant correlations between select soil chemical properties and soil enzyme activities are shown in Table 2. The
Pearson correlation coefficients revealed that soil properties such as Corg , nitrogen, phosphorus and soil water content had
positive correlations with soil enzymes. Corg , available nitrogen, and available phosphorus were significantly (P < 0.05) and
strongly correlated with all assayed enzymes in this study when compared with total nitrogen, and soil moisture content
(SMC). Pearson correlations between SMC and available N, available P, Corg and total N were not shown here because corre-
lations between these parameters were not significant (P < 0.05). Furthermore, available N and available P were not shown
as well due to aforementioned reasons.

Discussion

Soil properties

In the present study, Corg was significantly increased over two years in the top soil layer following the application of
the highest rate (12 t ha−1 ) of rice husk biochar compared to the control (Fig. 1). This finding affirms earlier studies by
Lehmann et al. [32] and Chan et al. [33] who observed a remarkable increase in organic carbon content of the soil after
biochar incorporation owing to its recalcitrant nature. This recalcitrant property explains the mechanism behind soil organic
carbon stabilization in tropical soils due to biochar’s chemical inertness and interaction with clay minerals. The Corg is
widely regarded as the engine room of the soil with great influence on physical, chemical and biological functioning [34].
This finding is expected to have a significant influence on soil structure improvement and carbon sequestration. This in
turn will bode well for the drive towards reducing CO2 losses from agricultural fields and provision of beneficial ecosystem
benefits.
In addition to carbon sequestration, biochar amendment will play a positive role in improving soil physical properties,
reduce the loss of top soil to erosion [35], minimize the use of pesticides, increase crop yield while reducing fertilization re-
quirements [36]. With all these benefits in hindsight, it is suggested that amending degraded tropical soils with biochar will
modulate carbon cycling and its sequestration with greater potential for positive effects on the environment. Furthermore,
increased soil water content and soil temperature – not reported in this study due to biochar amendment increased soil
available N (Fig. 2), which had a cascading influence on improved performance of rain-fed rice. An increase in soil inorganic
N at the highest application rate of biochar was observed, which suggest a reduction in leaching of N with positive impli-
cation for increased soil available N. The mechanism behind this could be that biochar amendment stimulated an increase
in net mineralization of soil organic nitrogen, while also sorbing mineralized N onto the pore spaces and negative charged
colloidal surface area of biochar. Findings from this study showed increased soil phosphorus levels especially in the top soil
layer under the highest application rate (12 t ha−1 ) of biochar (Fig. 3). Biochar amendment is widely noted for its nutrient
retention characteristics especially in poor tropical soils [47]. This has positive implications for nutrient use efficiency partic-
ularly in cereal crops with high nitrogen demand and low N use efficiency in the tropics. Phosphorus concentrations in soil
was increased by sorbing of orthophosphate ions in soil solution within biochar pores where it becomes less susceptible to
loss via run-off and leaching but remain accessible for uptake by plant roots [37]. Reduced P leaching and increased avail-
ability in biochar-amended soils could also be attributed to endothermic adsorption of P onto biochar mesopore surfaces
10 S.O. Oladele / Scientific African 4 (2019) e00107

[38]. Furthermore, increased Ca- bounded P, reduced Al and Fe-bounded P in acidic tropical soils [39] and increased soil
aggregate stability [40,41] as mediated by biochar amendment plausibly contributed greatly to increased P concentrations.
Furthermore, biochar amendment increased SMC in this study (Fig. 5). This is because biochar incorporation into the
soil enhanced water retention within the micropores of biochar, hence increasing the soil moisture content. The general
increase in soil moisture content may be ascribed to the enlarged specific surface area and porosity of biochar particles.
Furthermore, it is possible that biochar characteristics and pore morphology influenced biochar-water interactions through
contact and surface roughness. In addition, biochar amendment could have induced changes in soil structure and texture
thereby affecting soil moisture characteristics. This increase suggests that the application of biochar to croplands could con-
tribute to reduced irrigation requirements and frequency with positive implications for water-scarce regions in the tropics
and cropping systems solely dependent on rain-fall.

Grain and straw yield

The results of this study indicated that biochar amendment significantly increased grain, and straw yield of rain-fed rice
when compared with no biochar amendment (Table 1). This finding is consistent with the report of Koyama et al. [42] and
Haefele et al. [43] who both observed a significant increase in rice grain and biomass yield in an andosol under paddy rice
cultivation. Enhanced grain and straw yield of upland rice in this study could be ascribed to the increase in nutrient supply
to the test crop. In retrospect, the soil at the experimental site was acidic and sandy and would plausibly have benefited
from the ameliorating effect of biochar, which has been widely reported to improve bio-hydro-physical properties and crop
performance in similar soils [44]. Furthermore, increase in grain and straw yield could also be ascribed to the increase in
available N, nutrient use efficiency, cation exchange capacity, soil structure and water-holding capacity mediated by biochar
incorporation. The observed strong effect of rice husk biochar on the rice grain and straw yield was dependent on biochar
application rate. At application rates of 3–6 t ha−1 , grain and straw yield were substantially increased while application rates
beyond 6 t ha−1 appeared to decrease grain and straw yield. The negative effect of high rate of biochar on rice yield could
be ascribed to either immobilization of N onto biochar surfaces rendering them unavailable to plants, or release of toxic or
volatile compounds (e.g. PAHs, PCBs) detrimental to rice growth. However, this suggests that recommendation of optimal
biochar application rate in the study area should not exceed 6 t ha−1 particularly if farmers intend to cultivate upland rice.
Similarly, biochar amendment increased rice biomass yield. This increase in biomass yield after biochar amendment has
also been reported for other crops such as red clover, red fescue, plantain and maize [45–47], thus corroborates this study
findings.

Rhizosphere carboxylate secretions

Amendment of soil with biochar showed significant influence and increase in the secretion of carboxylates (Fig. 10).
This could be ascribed to the conditioning effect biochar had on the rhizosphere and modulation of proton efflux by plant-
microbe interaction. Studies have shown that excretion of carboxylates is often associated with proton extrusion, leading to
a substantial lowering of rhizosphere pH [48]. However, in this study, the trend was reversed as biochar amendment due to
its liming qualities did increase soil pH without negative consequence on organic acid secretions. Higher concentrations of
carboxylate in the rhizosphere following the highest application rate of biochar (12 t ha−1 ) in this study could be ascribed
to the addition of labile carbon and ash from biochar which helped increase root carboxylate secretion and stimulated root
functioning. The role of carboxylates has been documented as one of the several mechanisms for improved phosphorus (P)
efficiency of crops [49]. The exudation of carboxylates exudates helps solubilize and mobilize P from mineral and organic P
fractions in soil for plant uptake, especially in acidic and highly weathered tropical soils with low P availability. An inter-
esting finding from this study is the increase in concentration of carboxylates at the top soil despite high available P levels
mediated by biochar amendment. This is in contrast to the findings of Chen et al. [50] who reported decreased carboxylate
concentration in rhizosphere of soils with high P levels. It is important to note that exudation of carboxylates in the rhi-
zosphere could be dependent on root exudation, microbial degradation, plant re-absorption, soil desorption and adsorption
dynamics [51].

Soil enzymatic activity

After two years of study and single biochar application, biochar application rate of 12 t ha−1 significantly increased
the activity levels of soil invertase, urease, alkaline phosphatase, and catalase in the 0–0.1 and 0.1–0.2 m soil layers. The
increased enzymatic activities were pronounced in the 0–0.1 m layer than in the deeper soil layers. Changes in soil en-
zymatic activities as observed in this study have implications on soil processes such as nutrient cycling, decomposition of
litter, basal respiration and N2 O emissions [52]. Furthermore, increase in enzymatic activities could have been induced by
increased Corg , microbial biomass carbon and nitrogen pools which provided organic substrates for soil enzymes [53]. In this
study, catalase activity was greatly increased, especially in the top 0.1 m depth of the soil under the highest application
rate of biochar (12 t ha−1 ). Similar findings corroborating increased catalase activity upon biochar amendment was reported
by Zhang et al. [54]. The increase in catalase activity suggests that biochar amendment helped increase the oxidative ca-
pacity of soil microbes (Fig. 9). This is because catalase activity drives microbial oxidative-reductase metabolism which is
 S.O. Oladele / Scientific African 4 (2019) e00107 11

related to the metabolic activity of aerobic soil microbes [30]. The increase in invertase activity may be explained by labile
carbon availability following biochar incorporation (Fig. 7). Though, invertase activity slightly decreased with soil depths,
this could be ascribed to declining Corg and MBC pools in deeper soil depth [55]. Zhang et al. [56], also reported that soil
invertase activities in the 0–0.2 m soil depth were positively correlated with Corg , available N and P. It is important to note
that soil invertase activity could be used as an indicator in studying pattern of sugar released as food source for soil mi-
crobes through the hydrolysis of glucose and fructose [57]. Increased urease activity upon biochar application in this study
corroborates the report by Jindo et al. [58], who reported 40% increase in urease after soil amendment with wood biochar
and composts (Fig. 6). Urease enzyme modulates the hydrolysis of N fertilizer (urea) applied to the soil into NH3 and CO2
[11]. However as with other enzymes, urease enzymatic activities decreased with soil depths. This could be attributed to
decreased Corg content and microbial population with soil depth. The effect of biochar amendment on urease activity is
important as this enzyme is critically involved in N cycling and availability, and N2 O emissions from agricultural fields with
consequential effects on the environment [52]. In general, the increased urease activities following the highest application
rate of biochar (12 t ha−1 ) indicates that biochar can greatly affect the hydrolysis of urea fertilizer. Furthermore, the re-
sults of this study indicate an increased phosphatase activity under high rate of biochar application (Fig. 8). This was more
likely induced by the increased pools of microbial biomass which increased enzyme levels in response to biochar applica-
tion. Increased phosphatase activity signifies greater P availability for crops and soil microbe since phosphatase is known
to stimulate the hydrolysis of oxides and mineral bound P, ester phosphate bonds thus releasing orthophosphates for plants
and soil microbes [59]. This observation corroborates the findings of Chen et al. [60] who observed significant increase in
activity of alkaline phosphatases in soil amended with wheat straw biochar at pyrolysis temperature of 350 °C–500 °C.
However, phosphatase activity generally declined with increasing soil depths which could possibly be due to low levels of
microbial activities and Corg contents [56]. Overall, the positive effect biochar had on soil enzyme activity as shown in this
study reflects its beneficial effect on soil biota and nutrient cycling.

Relationship between soil enzymes and soil properties

The soil enzymes determined in this study viz; invertase, urease, alkaline phosphatase, and catalase exhibited strong
relationships with soil Corg , available N and P and soil moisture content (Table 2). However, Corg and available N were
significantly correlated with soil enzyme activity than other soil properties. Therefore, availability or non-availability of soil
nutrients and determination of soil health status could depend on activities of soil enzymes [61]. Furthermore, the release
of labile carbon from rice husk biochar and mineralization of organic N from SOM could also regulate soil enzyme activity.

Conclusion

Results from the current study suggests that amending soils with biochar at a rate of 12 t ha−1 had significant effect
on soil available N and P, and water content, especially in the 0–0.1 m soil layer. Similar trend was recorded for soil Corg
and soil enzymes. However, biochar application rate beyond 6 t ha−1 resulted in decreased grain and biomass yield of
upland rice. This implies that the decision to select the most appropriate biochar application rate for the study area and
areas with similar soil and environmental conditions should be related to soil quality improvement and upland rice yield
goal. If increasing rice yield is the major goal, then the recommended application rate of biochar should be between 3 and
6 t ha−1 depending on availability. However, if the major goal is to improve soil quality while also mitigating nutrient fluxes
and losses from agricultural fields, 12 t ha−1 application rate is recommended. Overall, biochar amendment was found to
mediate soil enzymatic activities involved in C, N, and P cycling which control soil nutrient dynamics and fluxes.

Competing interest

No conflict of interest.

Acknowledgments

The author acknowledges and appreciates the assistance of laboratory technicians at the Department of Crop, Soil and
Pest Management, Federal University of Technology, Akure and Mr. Omosuli of the Department of Agronomy, University
of Ibadan who helped with soil analysis. Thanks to the editor and anonymous reviewers for their shrewd comments and
suggestions.

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