Comparative Biochemistry and Physiology, Part B 145 (2006) 114 – 117

www.elsevier.com/locate/cbpb

Effects of a 30-km race upon salivary lactate

correlation with blood lactate
R.V.T. Santos ⁎, A.L.R. Almeida, E.C. Caperuto, E. Martins Jr., L.F.B.P. Costa Rosa 1
Laboratory of Metabolism – University of São Paulo, Institute of Biomedical Sciences, São Paulo, Brazil
Received 23 September 2005; received in revised form 26 June 2006; accepted 4 July 2006
Available online 11 July 2006

Abstract

Blood lactate has been used to determine the aerobic capacity and long distance performance. Recently, a new methodology has been suggested to
supplant the invasive blood lactate techniques. Salivary lactate has received attention because it shows high correlation to blood lactate in progressive
overload test. We evaluated the correlation between salivary and blood lactate during a long distance run and assessed possible changes in salivary lactate
concentration. Fifteen expert marathon racers ran 30 km as fast as possible. Saliva and 25 μL of blood were collected at rest and at each 6 km for lactate
determination. Blood lactate concentration increased in the 6th km and then remained constant until the end of the race. Salivary lactate increased after
18 km in relation to basal. We found high correlations between blood and saliva absolute lactate (r = 0.772, p b 0.05) and the blood lactate relative
concentration corrected by protein (r = 0.718, p b 0.05). The highest correlation found between absolute and relative salivary lactate was r = 0.994
(p b 0.001). Our results show that it is possible to use salivary lactate with absolute values or relative protein concentration. In addition, salivary lactate
showed a high correlation with blood lactate in endurance events.
© 2006 Elsevier Inc. All rights reserved.

Keywords: Anaerobic threshold; Blood; Endurance; Lactate; Saliva; Endurance exercise; Protein; Marathon

1. Introduction
Physical exercise promote changes in several hormonal and
biochemistry parameters. Lactate plasma concentration increases
during a short and long exercise bout in several species including
Human Beings (Stainsby and Brooks, 1990), horses (Evans et al.,
2002), dogs (Wakshlag et al., 2004), fish (Clarkson et al., 2005;
Wells and Baldwin, 2006) and chicken (Brackenbury et al., 1987).
Blood lactate is largely used to measure the aerobic capacity
mainly of elite athletes, since its submaximal production and
secretion are more sensitive to training and show higher corre-
lation to endurance performance than maximal parameters, such
as VO2 max (Sjödin et al., 1982; Coyle et al., 1988; Tegtbur et al.,
1993). Blood lactate concentration reflects not just its production,
but also the balance between production and removal (Stainsby
⁎ Corresponding author. Psychobiology and Exercise Research Center (CEPE) –
UNIFESP., Rua Marselhesa, 535 São Paulo/SP – BRAZIL CEP: 04.020-060.
Tel./fax: +55 11 5572 0177.
E-mail address: rvca@usp.br (R.V.T. Santos).
1
In memoriam.
1096-4959/$ - see front matter © 2006 Elsevier Inc. All rights reserved.
doi:10.1016/j.cbpb.2006.07.001
and Brooks, 1990). Therefore low intensity exercise shows cons-
tant lactate concentration, because lactate production is similar to
its removal, as opposed to what happens in exercise performed
above the anaerobic threshold, when blood lactate starts to accu-
mulate in Human Being (McLellan et al., 1991; Urhausen et al.,
1993; Billat, 1996) and rats (Voltarelli et al., 2002; Gobatto et al.,
2001).
As an alternative to blood parameters, substances measured in
saliva have been recently suggested to determine the intensity of
exercise. In this way, the use of the anaerobic threshold deter-
mination by chloride, sodium, amylase or salivary lactate concen-
tration may be an interesting alternative strategy to invasive
techniques (Chicharro et al., 1995, 1999). A high correlation
exists between salivary and blood lactate concentration in pro-
gressive intensity exercise and short intensive exercise, all short
term, acute exercise protocols, both in endurance and sprint/ana-
erobic runners (Ohkuwa et al., 1995; Segura et al., 1996). How-
ever, studies analyzing the correlation between the lactate in saliva
and blood during a long distance race are not known. The effects
of acute exercise on several biochemical parameters (cortisol,
IgA, IL-6, chloride, sodium, potassium) in the saliva have been
 R.V.T. Santos et al. / Comparative Biochemistry and Physiology, Part B 145 (2006) 114–117
studied showing discrepant results (Zhang and Tokura, 1999;
Minetto et al., 2005) explained by differences in the exercise
protocol, diurnal variation and the methods used to express results
(absolute concentration, secretion rate or ratio to total saliva
protein) (Gleeson et al., 2001; Laing et al., 2005). In addition, a
decrease in salivary flow may influence the salivary substance
concentration and consequently the metabolites concentrations
measured in saliva (Mackinnon et al., 1993; Mackinnon and
Jenkins, 1993; Mackinnon and Hooper, 1994).
In this study, we assessed the correlation between salivary and
blood lactate concentration during a long distance run and pos-
sible changes in salivary lactate promoted by saliva concentration
during endurance exercise.
2. Material and methods
The Ethics Committee of the Institute of Biomedical Science at
the University of São Paulo approved the experimental protocol.
All subjects were informed of the purpose and the risks of the
study and their written informed consent was obtained. Fifteen
elite marathoners (12 males and 3 females) ran 30 km at their best
personal time for the distance. The self-reported best time in the
marathon was in the range of 2.5–3 h and all participants had run
at least 3 marathons in the previous year.
2.1. 30-km race
All runners were instructed to run the 30 km as fast as possible.
The 30-km race started at 8:30 a.m. The temperature was 22 °C
and the air humidity 63%.
2.2. Blood and saliva sampling
We withdrew 25 μL of blood from the ear lobe of the runners at
rest and every 6 km, and stored it in 1.5-mL microcentrifuge tubes
containing 50 μL 1% NaF. Samples were stored at −80 °C and
analyzed within 1 week. Simultaneously, we gave each runner a
cup containing distilled water for them to wash their buccal cavity
prior to salivary collect. After washing, a 2-mL sample of saliva
was collected, simultaneously to blood sample collection. The
interval for blood and salivary sampling was less than 45 s.
2.3. Sample analysis and expression of results
About 25 μL of blood and salivary lactate were analyzed by a
Yellow Springs 1500 stat model lactimeter. The protein con-
centration in saliva was measured colorimetrically (Lowry et al.,
1951). The lactate concentration in blood and saliva was
expressed in mmol/L and salivary lactate concentration relative
to protein was determined by the salivary lactate (mmol/Lmg
protein) concentration divided by total protein concentration ratio
(salivary lactate concentration/protein concentration).
2.4. Statistical analysis
We used repeated measures ANOVA to compare blood lactate,
absolute and relative salivary lactate concentrations. When re-
115
Table 1
Blood and salivary lactate concentration through a 30-km race
Blood Salivary absolute Salivary relative
(mmol/L) (mmol/L) (mmol lactate/mg protein)
Rest 1.52 ± 0.10 0.39 ± 0.09 0.16 ± 0.03
6 km 2.39 ± 0.29⁎ 0.39 ± 0.05 0.14 ± 0.0
12 km 2.77 ± 0.39⁎ 0.51 ± 0.10 0.15 ± 0.03
18 km 3.00 ± 0.39⁎ 1.22 ± 0.27⁎ 0.33 ± 0.07⁎
24 km 2.94 ± 0.33⁎ 1.20 ± 0.27⁎ 0.34 ± 0.07⁎
30 km 3.15 ± 0.42⁎ 1.63 ± 0.39⁎ 0.42 ± 0.07⁎
Values are expressed in mmol/L for blood and salivary lactate and mmol lactate/
mg protein for relative salivary lactate concentration ± S.E.M.
⁎ Repeated measures ANOVA and Tukey's post-hoc test significant
difference in relation to rest, P b 0.05.
peated measures ANOVA showed some difference, we used
Tukey's post-test to identify the difference. The level of signi-
ficance adopted was p b 0.05. In addition, we carried out
Pearson's correlation to determine the correlation between sali-
vary and blood lactate concentration.
3. Results
In our study, we measured the salivary and blood lactate
concentration through a 30-km race. Blood lactate concentra-
tion increased during the race in comparison to basal results.
Salivary lactate, on the other hand, increased after 18th km
when expressed as absolute values or in relation to the protein
concentration. Statistical analysis showed that absolute and
relative salivary concentrations during rest and exercise are
significantly lower than those of the blood lactate (Table 1). In
addition, Table 1 shows the difference between absolute and
relative lactate concentration in saliva. The relative salivary
lactate was lower in all results in comparison to absolute
salivary lactate concentration.
Blood lactate concentration is higher than salivary concen-
tration. Two parameters had a similar profile along 30 km. Thus,
we found a significant correlation (r = 0.718, p b 0.05) between
blood and absolute salivary concentration (Table 1).
The blood lactate concentration is also high in comparison to
relative salivary lactate concentration, but the correlation between
both was r = 0.718 (p b 0.01), similarly to blood and salivary
absolute lactate (r = 0.772, p b 0.05) (Table 1).
Table 1 shows a highly significant correlation between
absolute and relative lactate concentration in saliva (r = 0.994,
p b 0.001). The correlation shown in Table 1 is the highest
found in our study.
4. Discussion
Human saliva contains much information regarding hor-
mones, amino acids, immunoglobulins and several metabolites,
including lactate (Mandel, 1990; Chicharro et al., 1998), which
are important for exercise physiologists and biochemists. The
interest in its use has been growing in the last years since saliva
sampling is not invasive and does not need much specialization
to be performed. Furthermore, it subjects the athletes to little
stress (Segura et al., 1996).
116 R.V.T. Santos et al. / Comparative Biochemistry and Physiology, Part B 145 (2006) 114–117
Recent studies have shown high correlation between blood
and salivary lactate concentration during a progressive test and
after anaerobic activities (Ohkuwa et al., 1995; Segura et al.,
1996; Pérez et al., 1999) while the effect of endurance exercise
on salivary lactate remains unknown. In this paper, we assayed
the effect of a long distance race (30 km) upon salivary lactate
concentration and the correlation between the absolute and
relative salivary lactate and blood lactate concentration, since
this kind of exercise may influence the level of salivary secre-
tion and its correlation (Shannon, 1967; Dawes, 1981; Chi-
charro et al., 1998; Walsh et al., 2002; Gleeson et al., 2001;
Laing et al., 2005).
We found that the absolute and relative salivary lactate
concentrations were constant up to the 18th km, when the lactate
concentration increased in relation to basal results. With regard to
blood lactate, we observe an increase in the first sample collected
at 6th km.
The correlations found in our study are in accordance with the
literature, and confirm early studies of Ohkuwa et al. (1995) and
Segura et al. (1996). However, the experimental and climatic
conditions are different in the three studies. Segura et al. (1996)
studied blood and salivary lactate during tests with progressive
overload on ergometric bicycle and controlled laboratory
conditions (r = 0.81). Ohkuwa et al. (1995) observed significant
correlations for long distance and sprint runners after a 400- and
3000-m race with high reproducibility in the two situations,
suggesting that the salivary lactate may be used as an indicator of
lactate accumulation level during short exercise. Our results are
similar to those in other studies and support early studies sug-
gesting that the salivary lactate has a high correlation with the
concentration of blood lactate during exercise of various inten-
sities, duration and muscle groups involved, and it does not
depend on temperature and relative air humidity.
Salivary lactate is possibly formed by passive diffusion from
blood and salivary gland (Ohkuwa et al., 1995). Shannon
(1967) suggested the salivary lactate increases because of an
increase in the concentration of blood lactate, which leads to an
increase in the permeability of the blood–saliva barrier. The
increase in salivary lactate only after 18 km in our study may
indicate the existence of a blood lactate threshold. Below this
threshold, the changes in blood lactate concentration apparently
are not reflected in salivary concentration.
It is not known how long it takes for the salivary lactate to form
after the blood goes into the salivary gland, but this time did not
influence the results found in our experimental model, because the
blood and saliva collections were carried out simultaneously.
However, this is an important factor to consider when researchers
study the behavior of salivary lactate concentrations in progres-
sive tests whose stages lasts less than 3 min when the interval
among each sample is short and the lactate concentration could
not be in balance with plasma lactate.
Release of saliva is controlled by the autonomic nervous
system and sympathetic stimulation is responsible for the increase
of electrolytes release and re-absorption and for the increase in
protein release, mainly amylase (Denis et al., 1978; Calvo et al.,
1997). Exercise increases the sympathetic drive and promotes
increase in salivary protein concentration via humoral impulse
upon the salivary gland (Calvo et al., 1997). So, during exercise
and after long distance races, the salivary viscosity increases
(Housh et al., 1991; Ljungberg et al., 1997). Hence, this effect
may promote false results in substances observed in saliva,
leading to incorrect conclusions about exercise effects (Mack-
innon et al., 1993; Mackinnon and Jenkins, 1993; Mackinnon and
Hooper, 1994; Walsh et al., 2002; Gleeson et al., 2001; Laing et
al., 2005). Previous studies on salivary lactate did not take this
concern into consideration, but studies with hormones and IgA
showed the importance of those observations. Regarding salivary
lactate, we did not find many studies in the literature, but our study
showed a high and significant correlation between absolute and
relative lactate concentration in saliva (r = 0.994, p b 0.001 –
Table 1), showing that the effects observed in salivary lactate
concentration were caused by the exercise protocol and not by the
ratio of salivary lactate concentration to total saliva protein, as
observed in studies with hormones and immunoglobulins.
Therefore, we concluded that salivary lactate measured during
a long distance race (30 km) had high correlation with blood
lactate, similarly to what happens in progressive tests. Moreover,
the salivary lactate concentration is not influenced by salivary
viscosity.
Acknowledgements
We are thankful to the athletes who made this study possible
and who endured the inconvenience of the investigation. This
study was supported by FAPESP (# 01/13766-4).
References
Billat, L.V., 1996. Use of blood lactate measurements for prediction of exercise
performance and for control of training. Sports Méd. 22, 157–175.
Brackenbury, J.H., Vincent, R., Mitchell, M.A., 1987. Plasma glucagons and
energy substrate response of domestic fowl to treadmill exercise. Comp.
Biochem. Physiol., A 86, 209–212.
Calvo, F., Chicharro, J.L., Bandrés, F., Lucía, A., Pérez, M., Álvarez, J.,
Mojares, A.L., Vaquero, A.F., Legido, J.C., 1997. Anaerobic threshold
determination with analysis of salivary amylase. Can. J. Appli. Physiol. 22,
553–561.
Chicharro, J.L., Calvo, F., Alvarez, J., Vaquero, A.F., Bandrés, F., Legido, J.C.,
1995. Anaerobic threshold in children: determination from saliva analysis in
field tests. Eur. J. Appl. Physiol. 70, 541–544.
Chicharro, J.L., Lucía, A., Pérez, M., Vaquero, A.F., Ureña, R., 1998. Saliva
composition and exercise. Sports Med. 26, 17–27.
Chicharro, J.L., Pérez, M., Carvajal, A., Bandrés, F., Lucía, A., 1999. The
salivary amylase, lactate and electromyographic response to exercise. Jpn. J.
Physiol. 49, 551–554.
Clarkson, K., Kieffer, J.D., Currie, S., 2005. Exhaustive exercise and the cellular
stress response in rainbow trout, Oncorhynchus mykiss. Comp. Biochem.
Physiol., A 140, 225–232.
Coyle, E.F., Coggan, A.R., Hooper, M.K., Walters, T.J., 1988. Determinants of
endurance in well-trained cyclists. J. Appl. Physiol. 64, 2622–2630.
Dawes, C., 1981. The effects of exercise on protein and electrolytes secretion in
parotid saliva. J. Physiol. 320, 139–148.
Denis, A.R., Schneyer, L.H., Sucanthapree, C., Young, J.A., 1978. Actions of
adrenergic agonists on isolated excretory duct of submandibular glands. Am.
J. Physiol. 235, F548–F556.
Evans, D.L., Priddle, T.L., Davis, A.J., 2002. Plasma lactate and uric acid
responses to racing in pacing Standardbreds and relationships with
performance. Equine Vet. J. 34, 131–134 (Suppl).
 R.V.T. Santos et al. / Comparative Biochemistry and Physiology, Part B 145 (2006) 114–117
Gleeson, M., Bishop, N.C., Sterne, V.L., Hawkins, A.J., 2001. Diurnal variation
in saliva immunoglobulin A concentration and the effect of a previous day of
heavy exercise. Med. Sci. Sports Exerc. 33, S54.
Gobatto, C.A., de Mello, M.A., Sibuya, C.Y., de Azevedo, J.R., dos Santos, L.A.,
Kokubun, E., 2001. Maximal lactate steady state in rats submitted to swimming
exercise. Comp. Biochem. Physiol., A 130, 21–27.
Housh, T.J., Johnson, G.O., Housh, D.J., Evans, S.L., Tharp, G.D., 1991. The
effect of exercise at various temperatures on salivary levels of immuno-
globulin A. Int. J. Sports Med. 12, 498–500.
Laing, S.J., Gwynne, D., Blackwell, J., Williams, M., Walters, R., Walsh, N.P.,
2005. Salivary IgA response to prolonged exercise in a hot environment in
trained cyclists. Eur. J. Appl. Physiol. 93, 665–671.
Ljungberg, G., Ericsson, T., Ekblon, B., Birkhed, D., 1997. Saliva and marathon
running. Scand. J. Med. Sci. Sports 7, 214–219.
Lowry, O.H., Rosebrough, N.J., Farr, A.R., Randall, R.J., 1951. Protein
measurement with the Folin phenol reagent. J. Biol. Chem. 193, 265–275.
Mackinnon, L.T., Hooper, S., 1994. Mucosal (secretory) immune system
responses to exercise of varying intensity and during overtraining. Int. J.
Sports Med. 15, S179–S183.
Mackinnon, L.T., Jenkins, D.G., 1993. Decreased salivary immunoglobulins
after intense interval exercise before and after training. Med. Sci. Sports
Exerc. 25, 678–683.
Mackinnon, L.T., Ginn, E., Seymour, G.J., 1993. Decreased salivary
immunoglubulin A secretion rate after intense interval exercise in elite
kayakers. Eur. J. Appl. Physiol. 67, 180–184.
Mandel, I.D., 1990. The diagnostic uses of saliva. J. Oral Pathol. & Med. 19,
119–125.
McLellan, T.M., Cheung, K.S.Y., Jacobs, I., 1991. Incremental test protocol,
recovery mode and the individual anaerobic threshold. Int. J. Sports Med.
12, 190–195.
Minetto, M., Rainoldi, A., Gazzoni, M., Terzolo, M., Borrione, P., Termine, A.,
Saba, L., Dovio, A., Angeli, A., Paccotti, P., 2005. Differential responses of
serum and salivary interleukin-6 to acute strenuous exercise. Eur. J. Appl.
Physiol. 93, 679–686.
Ohkuwa, T., Itoh, H., Yamazaki, Y., Sato, Y., 1995. Salivary and blood lactate
after supramaximal exercise in sprinters and long-distance runners. Scand. J.
Med. Sci. Sports 5, 285–290.
117
Pérez, M., Lucía, A., Carvajal, A., Pardo, J., Chicharro, L.J., 1999.
Determination of the Maximal Steady State of Lactate (MLSS) in saliva:
an alternative to blood lactate determination. Jpn. J. Physiol. 49, 395–400.
Segura, R., Javierre, C., Ventura, J.L.L., Lizarraga, M.A., Campos, B., Garrido, E.,
1996. A new approach to the assessment of anaerobic metabolism:
measurement of lactate in saliva. Br. J. Sports Med. 30, 305–309.
Shannon, I.L., 1967. Effect of exercise on parotid fluid corticosteroids and
electrolytes. J. Dent. Res. 46, 608–610.
Sjödin, B., Jacobs, I., Svendenhag, J., 1982. Changes in the onset of blood
lactate accumulation (OBLA) and muscle enzymes after training at OBLA.
Eur. J. Appl. Physiol. 49, 45–57.
Stainsby, W.N., Brooks, G.A., 1990. Control of lactic acid metabolism in
contracting muscle and during exercise. In: Pandoff, K.B., Holloszy, J.O.
(Eds.), Exercise in Sports Science Review. Williams & Wilkins, Baltimore,
pp. 29–63.
Tegtbur, U., Busse, M.W., Braumann, K.M., 1993. Estimation of an individual
equilibrium between lactate production and catabolism during exercise.
Med. Sci. Sports Exerc. 25, 620–627.
Urhausen, A., Coen, B., Weiler, B., Kindermann, W., 1993. Individual anaerobic
threshold and maximum lactate steady state. Int. J. Sports Med. 14, 134–139.
Voltarelli, F.A., Gobatto, C.A., de Mello, M.A., 2002. Determination of
anaerobic threshold in rats using the lactate minimum test. Braz. J. Med.
Biol. Res. 35, 1389–1394.
Wakshlag, J.J., Snedden, K., Reynolds, A.J., 2004. Biochemical and metabolic
changes due to exercise in sprint-racing sled dogs: implications for
postexercise carbohydrate supplements and hydratation management. Vet.
Ther. 5, 52–59.
Walsh, N.P., Bishop, N.C., Blackwell, J., Wierzbicki, S.G., Montague, J.C.,
2002. Salivary IgA response to prolonged exercise in a cold environment in
trained cyclists. Med. Sci. Sports Exerc. 34, 1632–1637.
Wells, R.M, Baldwin, J., 2006. Plasma lactate and glucose flushes following burst
swimming in silver trevally (Pseudocaranx dentex: Carangidae) support the
“releaser” hypothesis. Comp. Biochem. Physiol., A 143, 347–352.
Zhang, P., Tokura, H., 1999. Thermoregulatory responses in humans during
exercise after exposure to two different light intensities. Eur. J. Appl.
Physiol. 76, 285–289.