Soil Biology & Biochemistry 33 (2001) 1137±1140

www.elsevier.com/locate/soilbio
Short communication

Control of amino acid mineralization and microbial metabolism

by temperature
L.C. Vinolas a, V.R. Vallejo a, D.L. Jones b,*
a
Departament de Biologia Vegetal, Universidad de Barcelona, Diagonal 645, 08028 Barcelona, Spain
b
School of Agricultural and Forest Sciences, University of Wales, Bangor, Gwynedd LL57 2UW, UK
Received 19 May 2000; received in revised form 24 August 2000; accepted 13 September 2000

Abstract
Amino acids constitute a major reserve of soil organic-N and studies demonstrating direct uptake of amino acids by plants has indicated
that understanding their bioavailability and fate in soil is important to understanding terrestrial N cycling. The aim of this study was to
determine the effects of temperature and sorption on the mineralisation of three amino acids (glycine, lysine glutamate) in soil. Amino acid
sorption followed the series lysine . glycine . glutamate, whereas mineralisation rate followed the series glutamate . glycine . lysine.
These observations support the concept that sorption reduces the bioavailability of amino acids to the soil microbial population. Although the
amino acids were used preferentially for making new biomass rather than respiration, differences were apparent between the individual
amino acids with microbial assimilation ef®ciency (biomass production) following the series, lysine . glycine . glutamate. Our results
suggest divergences in the uptake and metabolism of the individual amino acids with a rapid mineralisation of amino acids which readily
enter general metabolic cycles (e.g. glutamate) compared to the amino acids which typically form the terminus of metabolic pathways (e.g.
lysine). Temperature signi®cantly affected the rate of amino acid mineralisation which increased up to 308C (Q10 ˆ 2.0) followed by a decline
as the temperature approached 408C. Rapid mineralisation occurred even at very low temperatures (18C). Amino acid mineralisation across
three experimental soil treatments followed the trend acidi®ed . control . eroded soil. In summary, the results indicate that mineralisation is
highly amino acid species dependent, has a mesophilic optimum, is retarded by sorption and is most rapid in soils which are not degraded.
q 2001 Elsevier Science Ltd. All rights reserved.
Keywords: Amino acids; Metabolism; Microorganisms; Mineralisation; Sorption

Ef®cient N cycling plays a critical role in natural and
agricultural ecosystem sustainability and a full understand-
ing of the fate of organic compounds within the soil-plant
system is key to improving the economy of crop production
and preventing freshwater pollution (Stevenson, 1982).
Recently, it has been demonstrated that in addition to the
uptake of NH41 and NO32, a wide range of plants directly
absorb and assimilate organic N in the form of amino acids
(Kielland, 1994; Jones and Darrah, 1994). These studies,
however, have focused largely on the plant and have ignored
possible competitive loss of organic-N by rhizosphere
microbial immobilization. In most surface soils amino
acids normally account for 7±50% of the total N with
most present in peptide and proteins (Stevenson, 1982).
Despite the recognition that amino acids may contribute a
key cog in soil N cycling (Chapin, 1995), to date, the in¯u-
* Corresponding author. Tel.: 144-1248-354997; fax: 144-1248-
382579.
E-mail address: d.jones@bangor.ac.uk (D.L. Jones).
0038-0717/01/$ - see front matter q 2001 Elsevier Science Ltd. All rights reserved.
PII: S 0038-071 7(00)00243-1
ence of soil management practices and environmental
factors on the behavior of amino acids in soil remains under-
studied. From studies with other C and N substrates,
temperature and microbial biomass/activity can be expected
to have major impacts on the availability of amino acids to
plants (Nicolardot et al., 1994). The aim of this study was to
determine the rate of mineralisation and sorption reactions
of three contrastingly charged amino acids as a function of
temperature. Further, the in¯uence of simulated erosion and
acidi®cation was assessed.
A loam textured Calcic Palexeralf was taken from experi-
mental ®eld plots located on the Plain of Barcelona, NE
Spain [41822 0 59 00 N, 286 0 44 00 E; mean annual precipitation
614 mm; mean annual temperature 15.58C (range 0±
358C)]. Three contrasting replicated experimental treat-
ments with differing soil organic matter contents were
sampled: (1) untreated plots; (2) simulated erosion plots in
which the top 10 cm of soil were manually removed 4 years
previously, and (3) acidi®ed, non-eroded plots in which
solid S was added 10 years previously to lower soil pH
1138 L.C. Vinolas et al. / Soil Biology & Biochemistry 33 (2001) 1137±1140

(Vinolas et al., 2001; Table 1). Plots were 20 m 2 and Table 1

contained mixed herbaceous scrub vegetation. Samples Selected properties of the three soil treatments. The control soil is non-
eroded and non-acidi®ed. All soils (Calcic Palexeralfs) were sampled from
were collected from the soil surface (0±10 cm) using a 0 to 10 cm
soil corer as described in Vinolas et al. (2001), sieved to
pass 2-mm mesh and stored ®eld-moist at 48C. The soil Control Eroded Acidi®ed
characteristics are provided in Table 1 with measurements 21
Sand, g kg 480 352 480
undertaken as described in Vinolas et al. (2001). Silt, g kg 21 309 440 309
Sorption of amino acids to the soil solid phase was deter- Clay, g kg 21 218 208 218
mined as described in Jones et al. (1994). Brie¯y, 1 g of CaCO3, g kg 21 54 29 10
moist soil was shaken with 10 ml of uniformly 14C-labelled Organic-C, g kg 21 18 3 19
Total-N, g kg 21 1.2 0.2 4.5
glycine, glutamate or lysine (5 mm; 0.15 kBq ml 21; ICN
Bulk density, kg dm 23 1.21 1.51 1.21
Pharmaceuticals Inc., Costa Mesa CA) for 10 min followed Microbial biomass, mg 285 56 115
by centrifugation (15,000 g, 5 min) and the amount of sorp- N kg 21 soil
tion determined by liquid scintillation counting. A measure pH(H2O) 7.88 8.12 4.97
of the amino acid binding strength was obtained by deter- NO32-N, mg kg 21 371 122 140
NH41-N, mg kg 21 174 174 277
mining the amino acid buffer power (Barber, 1995) where:
K, mmolc kg 21 333 322 338
b ˆ Ctot =Csol where Ctot ˆ …Csol £ u† 1 …Cads £ g† Na, mmolc kg 21 0.04 0.04 0.03
Ca, mmolc kg 21 34 30 120
and where Ctot is the total amount of amino acid in the soil Mg, mmolc kg 21 0.15 0.14 0.03
(mmol cm 23), Csol is the equilibrium solution concentration
(mmol cm 23), Cads is the amount of amino acid sorption
(mmol g 21 soil), u is the volumetric water content Table 2
(cm 3 cm 23) and g the soil bulk density (g cm 23). Buffer power values describing the sorption of three contrastingly charged
amino acids in three experimental soil treatments. Values are means ^ SE
Amino acid mineralisation was undertaken as described in (n ˆ 2). For a de®nition of the buffer power see text. Superior letters after
Jones (1999) using 14C-labelled glycine, lysine and glutamate. the SE values indicate signi®cant differences between amino acid/treatment
Brie¯y, the amino acid solution (5 mM) was mixed by light (P , 0.05)
agitation with 0.5 g of soil to give a ®nal concentration of
Amino acid Soil treatment
0.5 mmol g 21 dry wt soil and a matric potential of
20.033 MPa. 14CO2 evolved over a 48 h period was recovered Control Eroded Acidi®ed
with 1 ml 1 M NaOH traps. After 48 h, the amount of 14C label a a
Glycine 0.64 ^ 0.12 0.69 ^ 0.00 1.63 ^ 0.15 b
remaining in the soil was determined by extraction with 5 ml
Glutamate 0.44 ^ 0.10 a 0.47 ^ 0.04 a 0.59 ^ 0.06 a
of 0.5 M K2SO4 (Joergensen and Brookes, 1990). Any 14C- Lysine 9.27 ^ 0.12 c 16.18 ^ 0.10 d 4.69 ^0.07 e
label not recovered in either the NaOH traps or K2SO4 extracts
was assumed to be in the microbial biomass (biomass- 14C;
Jones and Hodge, 1999). Apparent microbial assimilation ef®-
ciency (Ma) was determined using the equation: acidi®ed soil . control soil . eroded soil. Amino acid
mineralization increased with temperature from 1 to 308C
Ma ˆ biomass-14 C=…biomass-14 C114 CO2 †
for all soil treatments and substrates studied but declined
All treatments were performed in duplicate with some towards 408C indicating a mesophilic optimum (Fig. 1). The
treatments independently replicated on different days and mean Q10 values (^SE) across all soil treatments from 1 to
with soil collected at another sampling date. Statistical 308C was 2.0 ^ 0.2 for lysine, 2.0 ^ 0.3 for glycine and
analysis was performed with SPSS (SPSS Inc., Chicago, 1.9 ^ 0.2 for glutamate.
ILL). Microbial assimilation ef®ciencies greater than 40% were
Our ®ndings indicate that for lysine (net charge 11), a obtained for all treatments, with assimilation ef®ciency
strong interaction exists with the soil's cation exchange following the series lysine . glycine . glutamate (Fig. 1).
complex, while glycine (net charge 0) and glutamate (net Assimilation ef®ciency increased with decreasing tempera-
charge 21) were weakly bound and remained mainly in tures in agreement with the hypothesis that maintenance
solution (Table 2). In contrast, the rate of amino acid miner- costs are greater at elevated temperatures (Atlas and Bartha,
alization followed the series glutamate . glycine . lysine 1993; Nicolardot et al., 1994). At low temperatures (1±
(Fig. 1). The negative correlation between mineralization 108C), assimilation ef®ciencies . 90% were apparent for
rate and sorption strength indicated that sorption to the soil's glycine and lysine, but only 70% for glutamate. This indi-
solid phase probably limits bioavailability (Bartlett and cated that the actual amount of amino acid taken up by the
Doner, 1988). The results indicated highly signi®cant microbial biomass at low temperatures (18C) was actually
effects (P , 0.01) of amino acid type, soil experimental much greater than that revealed by the CO2 evolution
regime, and temperature on mineralization rate. The rate measurements. Signi®cant differences (P , 0.01) were
of amino acid mineralization generally followed the series observed in microbial assimilation ef®ciency between
 L.C. Vinolas et al. / Soil Biology & Biochemistry 33 (2001) 1137±1140 1139

Fig. 1. Amino acid mineralisation as a function of temperature. Panel A (left) represents the time dependent mineralisation of 14C-labelled lysine, glycine and
glutamate. Only the control soil data is shown. Panel B (center) represents the effect of temperature on the rate of amino acid mineralisation to CO2. Rates were
calculated from the linear portion (1±3 h) of the mineralisation curves of each experimental treatment (control, eroded and acidi®ed). Panel C (right)
represents the impact of temperature and soil management regime on the incorporation of amino acid C into new cell biomass (microbial assimilation
ef®ciency) for each amino acid. Note the y-axis scales are different for each amino acid. All values represent means ^ SE (n ˆ 2).

each treatment, generally following the series eroded .
control . acidi®ed soil.
Although glycine and glutamate were both weakly
sorbed, they exhibited different rates of CO2 production
and different microbial assimilation ef®ciencies suggesting
a divergence in either their microbial transport or metabo-
lism. This hypothesis is supported by pure bacterial culture
studies which show that amino acids are transported into the
cell by group speci®c transporters (e.g. neutral, basic,
acidic) and differentially metabolized (Anraku, 1980;
Bender, 1985). In many organisms glutamate is the common
precursor for the synthesis of many other amino acids as it
forms a major pathway by which NH41 is assimilated (e.g.
GOGAT/GS and transamination using keto acids; Moat,
1979). In contrast, amino acids such as lysine typically
form end products in amino acid metabolism, the precursor
for which is typically aspartate (Moat, 1979; Bender, 1985).
Therefore in addition to the proposed reduction of microbial
lysine uptake due to sorption, the repression of internal
lysine metabolism due to internal metabolic constraints is
also likely (i.e. lack of suitable enzymatic machinery to
reverse the synthesis pathway; e.g. lysine to aspartate).
Glutamate was mineralized to CO2 to the greatest extent
possibly due to its pivotal role in microbial amino acid
transamination reactions producing keto acids which can
be readily catabolised to CO2 (e.g. succinate; Fig. 1; Bender,
1985).
Our results suggest that soil experimental treatment
signi®cantly in¯uenced amino acid turnover with the eroded
soil generally exhibiting a lower ability to recycle amino
acids due to it having a lower starting biomass. In contrast,
the acidi®ed soil had a greater ability to mineralize amino
acids per unit of microbial biomass suggesting possible
shifts in community structures between these two soil treat-
ments (e.g. greater fungal biomass).
In conclusion, our results suggest that signi®cant differ-
ences exist between the behavior of different amino acids
in soil with basic amino acids strongly sorbed while
neutral and negatively charged amino acids remain
largely in solution. The sorption of the amino acids
appeared to signi®cantly impact their bioavailability. In
addition, it is likely that internal metabolic control of
microbial amino acid transport and synthesis pathways
also signi®cantly in¯uences the rate of amino acid miner-
alization. However, we also speculate based upon
our results that other factors such as temperature and
microbial community structure also strongly in¯uence
mineralization rate.
1140 L.C. Vinolas et al. / Soil Biology & Biochemistry 33 (2001) 1137±1140
References
Anraku, Y., 1980. Transport and utilization of amino acids by bacteria. In:
Payne, J.W. (Ed.). Microorganisms and Nitrogen Sources. John Wiley,
New York, pp. 9±33.
Atlas, R.M., Bartha, R., 1993. Microbial Ecology: Fundamentals and
Applications. 3rd ed. Benjamin Cummings, Wokingham.
Barber, S.A., 1995. Soil Nutrient BioavailabilityÐA Mechanistic
Approach. 2nd ed. John Wiley, New York.
Bartlett, J.R., Doner, H.E., 1988. Decomposition of lysine and leucine in
soil aggregatesÐadsorption and compartmentalization. Soil Biology &
Biochemistry 20, 755±759.
Bender, D.A., 1985. Amino Acid Metabolism. 2nd Ed. John Wiley, New
York.
Chapin, F.S., 1995. New cog in the nitrogen-cycle. Nature 377, 199±
200.
Joergensen, R.G., Brookes, P.C., 1990. Ninhydrin-reactive nitrogen
measurements of microbial biomass in 0.5 M K2SO4 soil extracts.
Soil Biology & Biochemistry 22, 1023±1027.
Jones, D.L., 1999. Amino acid biodegradation and its potential effects on
organic nitrogen capture by plants. Soil Biology & Biochemistry 31,
613±622.
Jones, D.L., Darrah, P.R., 1994. Amino-acid in¯ux at the soil-root interface
of Zea mays L. and its implications in the rhizosphere. Plant and Soil
163, 1±12.
Jones, D.L., Hodge, A., 1999. Biodegradation kinetics and sorption
reactions of three differently charged amino acids in soil and their
effects on plant organic nitrogen availability. Soil Biology & Biochem-
istry 31, 1331±1342.
Jones, D.L., Edwards, A.C., Donachie, K., Darrah, P.R., 1994. Role of
proteinaceous amino acids released in root exudates in nutrient acquisi-
tion from the rhizosphere. Plant and Soil 158, 183±192.
Kielland, K., 1994. Amino acid absorption by arctic plants: implica-
tions for plant nutrition and nitrogen cycling. Ecology 75, 2373±
2383.
Moat, A.G., 1979. Microbial Physiology. John Wiley, New York.
Nicolardot, B., Fauvet, G., Cheneby, D., 1994. Carbon and nitrogen cycling
through soil microbial biomass at various temperatures. Soil Biology &
Biochemistry 26, 253±261.
Stevenson, F.J., 1982. Organic forms of soil nitrogen. In: Stevenson, F.J.
(Ed.). Nitrogen in Agricultural Soils. ASA-CSSA-SSSA, Madison, WI,
pp. 67±122.
Vinolas, L.C., Healey, J.R., Jones, D.L., 2001. Kinetics of soil microbial
uptake of free amino acids. Biology and Fertility of Soils (in press).