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Biosensors and Bioelectronics

journal homepage: www.elsevier.com/locate/bios

Lactate biosensing: The emerging point-of-care and personal healthmonitoring

Fahmida Alama, Sohini RoyChoudhurya, Ahmed Hasnain Jalala, Yogeswaran Umasankarb,Shahrzad Forouzanfara, Naznin Aktera, Shekhar Bhansalia, Nezih Palaa,⁎

a Department of Electrical and Computer Engineering, Florida International University, Miami, FL 33174, United Statesb Biomolecular Sciences Institute, Florida International University, Miami, FL 33199, United States

A R T I C L E I N F O

Keywords:LactateWearableLactate oxidaseLactate dehydrogenaseInvasive sensingNon-invasive sensing

A B S T R A C T

Lactate plays a crucial role in the anaerobic metabolic pathway of humans. In situations of oxygen deficit, itsproduction increases; leading to several life-threatening conditions such as hemorrhage, respiratory failure,trauma or ischemia from lactate acidosis. Lactate level detection and point-of-care (POC) monitoring in a fast,accurate and non-invasive manner is ultimately important for many health care applications. Optical andelectrochemical techniques are employed in lactate sensing to achieve high sensitivity and selectivity, minia-turization, portability, simplicity, and low cost. To improve the selectivity and sensitivity, two important en-zymes, lactate oxidase (LOx) and lactate dehydrogenese (LDH) are employed. Conventional methods for lactatedetection are not fast enough to be used in point-of-care or personal health monitoring settings. Moreover, theexisting point-of-care lactate sensing tools follow invasive or partially invasive sampling protocols such as fingerpricking. In this review, a comprehensive overview of different lactate biosensing devices is presented.Particularly, the state-of-the-art and prospects of wearable, non-invasive lactate sensing from different biofluidsare discussed.

1. Introduction

Lactic acid (LA) is the end-product of anaerobic breakdown ofglucose in tissues. It exists in cells and is transported to the liver, whereit is oxidized back to pyruvate (PA) (Blomkalns, 2006) and eventuallyconverted back to glucose via the Cori cycle. During tissue oxygenationdeficits, LA is produced with consumption of energy. In such persistentoxygen debt situations, lactic acidosis ensues (Stacpoole et al., 1994).LA exists in two optical isomeric forms: L-lactate and its mirror image,D-lactate (Kowlgi and Chhabra, 2015). Blood lactate level in unstressedpatients at rest is reportedly ~ 0.5–1mM whereas patients critically illhave concentrations> 4mM. The plasma lactate levels in the body il-lustrate a finely tuned interplay of parameters influencing the delicatelactate balancing system; as its viability in the human body is within anarrow pH range, between 7.2 and 7.4 (Valenza et al., 2005). Hy-perlactatemia is defined as persistent mild to moderate (2–4mM)concentrations without metabolic acidosis, while lactic acidosis ischaracterized with persistent increased levels (usually> 5mM) in as-sociation with metabolic acidosis (Rathee et al., 2016). Lactate innormal human subjects is removed very quickly at a rate of 320mM/L/h, mostly by liver metabolism and re-conversion of lactate back to PA.

Excess lactate leads to tissue hypoperfusion with increased anaerobicmetabolism (Jones and Puskarich, 2009). While hyperlactatemia canoccur during adequate tissue perfusion, intact buffering systems, andadequate tissue oxygenation; lactic acidosis occurs with increasedtransient metabolic demand (e.g., post-seizure lactic acidosis) or majormetabolic dysregulation (Garcia-Alvarez et al., 2014). Several etiologiescan be responsible for lactic acidosis, most commonly circulatoryfailure and hypoxia. Evidence suggests increased morbidity and mor-tality for patients with persistently elevated lactate. Excess lactate inthe body also causes hemorrhage, respiratory failure, trauma, seizures,ischemia, renal issues, hepatic disease, sepsis, tissue hypoxia, shock,seizures, sepsis, blood loss, anemia, hematoma, malignancy, diabetes,heart diseases, ischemia, hypoxemia, meningitis, renal failure (Nakaoet al., 1982) and even arthritis.

Lactate monitoring is of prime importance to diagnose and evaluatethe health concerns related to lactate acidosis and those which occur inoxygen deficit situations (where the lactate levels in the body increasebeyond the accepted values). These include systemic healthcare inmilitary and high-risk personnel (Goodman et al., 1999; White, 2015),clinical emergencies, sports, and general medicine. For example, sportsmedicine requires monitoring lactate for training and performance

https://doi.org/10.1016/j.bios.2018.06.054Received 4 June 2018; Accepted 26 June 2018

⁎ Corresponding author.E-mail address: npala@fiu.edu (N. Pala).

Biosensors and Bioelectronics 117 (2018) 818–829

Available online 28 June 20180956-5663/ © 2018 Elsevier B.V. All rights reserved.

T

endurance evaluation. This evaluation can be an indicator of the phy-sical training level of an athlete (Tsai et al., 2007), as high blood lactatelevels can lead to a drop in the blood pH level causing exhaustion.Inadequate oxygen supply at childbirth may also tend towards acidosiswith increase lactate levels in the fetus blood. Lactate and pH levels infetal scalp blood are used as indicators of hypoxic or acidotic distressduring labor in the fetus (Heinis et al., 2011). Elevated lactate levels inthe body can also be because of bacterial or fungal infection in anyclosed cavity of the body like joints, meninges, or pleura. Lactate, beinga major cause of acidification in a microenvironment of cancer cells andtumors, if uncontrolled, may lead to tumors (Walenta et al., 2000).Lactate also plays an imperative role in brain metabolism providingindications of a cerebral stroke or trauma in the brain (Cureton et al.,2010). Therefore, a laboratory-based testing system is not enough todiagnose and monitor these life staking diseases and conditions. Con-tinuous lactate monitoring could be a possible solution to early diag-nosis and improve the quality of life. Though recent advances in con-tinuous monitoring systems have been made, a dearth exists inwearable, minimal invasive options. Finger-pricks are invasive meansto assess an individual's pathophysiology. Laboratory-based analyticalinstrumentation is not amenable for field-based use. Recently, novelfabrication concepts with robust sensing technologies are being im-plemented on textiles for highly-sensitive chemical or optical analyses.Wearable sensors in this new generation have the potential to renderthe wearer's health information in real time (Chuang et al., 2010).

This article presents an extensive review of the current state of theart and the changes in lactate sensors from different biofluids, addres-sing the pros and cons of each; moving towards non-invasive detectionand continuous monitoring in real time. Commercially available Lactatesensors are also discussed with an overview of the requirements yet tobe met in detection.

2. Bio-catalysts for lactate biosensing

Biosensing is a mechanism to sense bio-compounds through mon-itoring the physical, chemical or biological changes that they causewith their presence. Coupling enzymes as biocatalysts to amplify signalsis an important paradigm in bio-sensing. Enzymes are among the mostefficient forms of natural catalysts and easily available. Enzyme-basedbiosensors are biocompatible and provide simple, direct, rapid responsewith high specificity. Enzymes and substrates usually have com-plementary structures that bond to form a complex reactive site, whichstabilizes the transition reaction state or lowers activation energy in thereaction.

Two different enzymes that are widely used for lactate assays areLOx and LDH. They consume the electro-reactive reagent to generate anelectro-reactive species that can be monitored to correlate the responsewith the analytes’ concentrations. The physical, biological and chemicalproperties of these enzymes are discussed below. Utilization of nano-materials in lactate biosensors is known to have notably improved thereaction involved with enhanced electron transfer (ET) (Pundir et al.,2016).

LOx catalyzes the aerobic oxidation of lactate to PA, releasing hy-drogen peroxide (H2O2). It is then oxidized at the electrode surface,restoring the former concentration of oxygen, providing a proportionalresponse (Pfeiffer et al., 1997). This is depicted in the equations below:

L-lactate + LOx → pyruvate + LOx red (1)

LOx red + O2 → LOx ox + H2O2 (2)

LOx is available from different bacterial sources like Pedi coccus,Aerococcus viridians, and Mycobacterium smegmatis, also being pre-sent in viruses and cellular organisms. Based on different sources, theiractivity and stability may differ with pH and temperature. Being amember of the flavin mononucleotide (FMN) family, it employs FMN asa cofactor to catalyze the oxidation of hydroxyl acids in its reactions

involving glycolate oxidase, LOx, L-lactate monooxygenase, flavocyto-chrome b2, long chain a- hydroxyl acid oxidase and L-mandelate dehy-drogenase. It is also an oxidoreductase enzyme which uses oxygen asoxidant on the substrate to generate oxygenase. Depending on thesource, this enzyme is known to be stable within a pH range of 4–9 (VanHaeringen, 1981).

LDH plays an important role in cellular respiration converting glu-cose to adenosine triphosphate (ATP) for our cells, a crucial step inenergy production. It is an enzyme that catalyzes lactate to PA, throughits cofactor, Nicotinamide adeninedinucleotide (NAD+).

+ ⎯ →⎯⎯⎯ + ++ +NAD Lactate NADH Pyruvate H

LDH (3)

This conversion is a crucial step in energy production in humancells. There are two ways of measuring the amount of lactate throughLDH in oxygen deficit situations; either through oxidation of NADHwith PA or through the reduction of NAD+. One unit of enzyme isknown to catalyze the oxidation of NADH or reduction of NAD+ at1 µmol min−1, under standard conditions.

There are reportedly five different isozymic forms of LDH, dis-tinguished by slight differences in their structure. These isozymes canbe separated using NAD+ and NADH. Depending on the source, thisenzyme is known to be stable over a relatively narrower pH range of5–8 with optimum activity shown around 7.2–7.4. LOx attracts atten-tion of researchers in the design of a biosensor, eliminating the diffi-culties involved in oxidation of co-enzymes (NADH). Carbon-basednanomaterial is extensively explored for lactate detection using thisenzyme, besides other nanostructures of Pt, Au or Zn. Nanostructures ofPt, Au, C and Zn, as catalysts for electrochemical oxidation, are knownto increase the electro-activity and detection sensitivity over a broadphysiologically relevant range. Their use in the design of biosensors canquantitatively determine lactate in human plasma.

3. Sensing techniques

Electrochemical and optical sensors offer considerable promiseowing to their miniaturized form factor and low cost. There exist someminiaturized lactate bio-sensing devices in sports medicine for athletesand some which can optically detect the analyte with an electronic readout of the lactate concentration in the body. Efforts however are still onfor minimal invasive techniques in biomedicine for continuous mon-itoring and diagnosis (Salazar et al., 2012).

3.1. Electrochemical techniques for lactate sensing

Electrochemical biosensors can be an attractive means for selectivedetection of lactate through direct translation of enzymatic reactionsinto electrical signals in presence of its specific enzyme (Koyun et al.,2012). The fundamental operation of this technique depends on (i)electron transfer mechanism and (ii) enzyme immobilization technique.These two steps are integral to electrochemical sensing of lactate andare described below.

In electrochemical biosensors, biocatalysts are immobilized onto asupporting substrate close to the electrode surface. Their response de-pends on the immobilization technique employed, the nature of thebiocatalyst used and the other adsorbed species through ET, mediators,and additives. Depending on the nature of the support and stability ofthe biomolecule, several techniques for immobilizing the enzyme canbe used. These include, (i) physical adsorption, (ii) entrapment behind adialysis membrane or within a polymeric film, (iii) covalent couplingthrough a cross-linking agent, and (iv) incorporation within the bulk ofa carbon composite matrix (Scouten et al., 1995). The framework of ETcan perform selective and specific reactions (Dong and Wang, 2002). InET, electrons are transferred from active site of the enzyme to theelectrode (Habermüller et al., 2000). Since this mechanism between thebio-molecule and the electrodes is sluggish on planar electrodes, the

F. Alam et al. Biosensors and Bioelectronics 117 (2018) 818–829

819

immobilization medium should be engineered to provide enhancedelectron transport. Different strategies have been used to constructenzymatic sensor electrodes to facilitate direct ET process or mediator-modified enzymes in L-lactate detection. Protein mediated ET is afundamental phenomenon in cellular processes. The redox enzyme actsas an electro-catalyst, facilitating ET between the electrode and thesubstrate (Ghindilis et al., 1997). Such biosensors are known to offerimproved selectivity since their operating potential is closer to theredox potential of the enzyme and hence less exposed to interferingreactions. Several enzymes can undergo direct catalysis through direct(mediator-less) ET. This process is associated with: (i) ET from theelectrode to the substrate molecule, or vice versa, via the active site ofthe enzymes (i.e. LDH and LOx as shown in Fig. 1a and b); and (ii)catalytic nature of the whole process. ET between the active site of theenzyme and the transducer surface through the cross-linker, is shown inFig. 1c and d. It can thus be concluded that in mediator-less bio-electro-catalysis, the electrode itself is a substrate for the reaction; and theenzymatic transformation on the electrode is not a separate reaction.On the other hand, diffusional electron mediators (Montagné andMarty, 1995) are employed as charge transporters between active redoxcenter of the enzyme and transducer. Different catalysts and polymer-based mediators are integrated with the electrode to eliminate inter-ference from the redox reaction. Examples include redox dye

compounds transition metal compound-based mediators polymeric orperm selective membranes (Bridge et al., 2007). These have been foundto enhance sensor performance with linearity, lower detection limit,enhanced stability, sensitivity, selectivity, and superior electron trans-port. Mediators and matrices in combination with a recognition layer ofnanoparticles (NPs) have been found to repel interfering electro-activespecies, resulting stable quantification of lactate. Cross-linkers (asshown in Fig. 1c and d) have also been seen to exclude interference,providing greater stability, preventing fouling, and protecting the en-zyme structure (Groegel et al., 2011). The mechanism of mediated ETfor both enzymes (LDH and LOx) for lactate are shown in Fig. 1e and f.

In electrochemical sensing mechanism two- or three-electrode sen-sing platforms are widely used (Jalal et al., 2017) for detection ofanalytes (shown in Fig. 2a) in which the reference electrode (commonlymade from Ag/AgCl), is kept at proximity to the working electrode tomaintain a stable and known potential (Pasha et al., 2016). Theworking electrode serves as a transducer, while the counter electrodeestablishes a path to pass the current due to the potential changes at theworking electrode. These sensors types are composed of a chemical(molecular) recognition system and a physicochemical transducer. Re-cognition systems react with the unknown analyte whereas the trans-ducer converts the chemical response into an electrical signal. Fig. 2adepicts a standard three electrode cell system with a commercial glassy

(e)

(b)(a)

(f)

(c) (d)

LOx

Lactate Pyruvate

e-

Redox mediator

O2

electrode

Reduction

Oxidation

LDH

Lactate Pyruvate

e-

Redox mediator

O2

electrode

Reduction

Oxidation

LOx

Lactate Pyruvate

e-

O2

electrode

LDH

Pyruvate

e-

electrode

O2

Lactate

LDH

Lactate

e-

O2

electrodeCross linker

Pyruvate

LOx

Lactate

e-

O2

electrodeCross linker

Pyruvate

Fig. 1. Schematic of direct electron transfer in (a) LOx, (b) LDH, (c) LOx with cross-linker and (d) LDH with cross-linker; and for mediated electron transfer in (e) LDHand (f) LOx.

F. Alam et al. Biosensors and Bioelectronics 117 (2018) 818–829

820

carbon electrode (GCE) modified with the prepared catalyst as workingelectrode. A Pt wire, and an Ag/AgCl electrodes are used as counter andreference electrodes respectively (Zhang et al., 2018). The CV plot re-presents response of a Pt/HCS (hollow carbon sphere) in 0.1M PBS (pH= 7.4) and in the presence and absence of H2O2. From the response,two small cathodic peaks are observed at − 0.06 and − 0.36 V, cor-responding to the reduction of Pt oxide and defective sites of HCS. Thecommon approaches for measurement are: cyclic voltammetry (CV),amperometry and potentiometry.

The response of electrochemical lactate biosensors resulting from aredox reaction in the presence of electroactive species depends linearlyon the concentrations of the analyte being measured, when a constantpotential is applied between the working and reference electrodes. Forexample, a saturated Ag/AgCl was employed as a reference electrode tocontrol the potential on the working electrode with immobilized en-zyme for a steady and reproducible potential to be maintained on theworking electrode with the flow of current in between the working andcounter electrodes (Romero et al., 2008). Similarly, CV analysis is alsoused for the characterization and calibration of lactate sensing. As thereduction of oxygen causes interference, oxidation of H2O2 is preferredfor lactate recognition. In the presence of LOx, L-lactate oxidized to PAand produces H2O2. H2O2 further reduced or oxidized either and gen-erates current proportionally to the L-lactate concentration. Sensitivity

of such sensors can be improved by acclimated mixed bacteria andpalygorskite co-modified oxygen electrode. For instance, sweat is de-posited on screen-printed carbon electrodes (SPCEs) functionalizedwith Fe3+ in the case of which, the response depends on the non-equilibrium potential difference of the Fe (III)/Fe (II) redox couple(Lewenstam, 2011). Fig. 2b (i) depicts calibration curve of the potentialdifference of the Fe (III)/Fe (II) couple versus the concentration oflactate from 0 to 250mM (after 60 s of UV irradiation). The inlet ofFig. 2b (iii) reports the trend of the lactate excretion rate (LER), innanomoles per square centimeter per minute (nmol cm−2 min−1) as afunction of the exercise time, showing the possible effects of lactatedilution are negligible (Onor et al., 2017).

Electrochemical biosensors are attractive due to their high sensi-tivity, wide linear range, and rapid response and hold a leading positionamong the presently available biosensor systems. However, signal re-duction from fouling agents, interference from chemicals present in thesample biofluid, poor coupling of biochemical recognition materials,and electrochemical transducers affect the performance of electro-chemical biosensors.

3.2. Optical techniques for lactate sensing

Optical detection relies on photon transfer, eliminating the need of

Enzyme film

Electrode material

Insulator

Potentiostat

Working ElectrodeCounter Electrode

Reference Electrode

(a)

(b)

(c)

(i) (ii) (iii)

Fig. 2. (a) Electrochemical set-up for lactate measurement and CV curves of Pt/HCS-modified GC electrode in 0.1M PBS (pH = 7.4) solution with the absence andpresence of 500 µM H2O2 at a scan rate of 50mV s−1 (Zhang et al., 2018) (b) (i) Calibration curves of the potential difference of the Fe(III)/Fe(II) couple versus theconcentration of lactate (0–250mM), (ii) Error bars represents the standard deviation of N=5 measurements performed using 5 different electrodes, open circlerefer to the potential value at t= 0 irradiation time (iii) Monitoring of sweat lactate during 24min of cycling exercises in 3 different subjects while changing the workintensity and during the successive cool down (Onor et al., 2017) (c) Chemiluminescence based Lactate Sensors (Biswas et al., 2017).

F. Alam et al. Biosensors and Bioelectronics 117 (2018) 818–829

821

Table1

Lactatesensorsco

mpa

rison:

detectionmecha

nism

andpe

rforman

ce.

Type

ofEn

zyme

Type

ofbios

enso

rTy

peof

bodily

fluid

Inva

sive

/non

-inva

sive

Sens

itivity

Limit

ofDetec

tion

(LOD)

Med

iator/Non

-med

iator

Immob

ilizationtech

niqu

eReferen

ces

LOx

Electroc

hemical

Sweat

Non

-inva

sive

–0.2mM

–Cov

alen

tcross-lin

king

Tur-Garcíaet

al.(20

17)

LOx

Electroc

hemical

–inva

sive

32nA

mm

−2mM

−1

2µM

UV-cross-link

erCross-lin

king

Weltinet

al.(20

14)

LOx

Electroc

hemical

Sweat

Non

-inv

asive

644.2nA

mM

−1

–Te

trathialfulvalen

e(TTF

)med

iator

Cross-link

ing

Jiaet

al.(20

13a)

LOx

Electroc

hemical

Sweatan

dbloo

d–

36.8

mAM

−1cm

−2

11µM

–Cov

alen

tBind

ing

Lamas-Ardisan

aet

al.

(201

4)LO

xElectroc

hemical

Sweat

Non

-inv

asive

256nA

cm−2mM

−1

–Glutaraldeh

yde(G

A)

Cross-link

ing

Abrar

etal.(20

16)

LDH

Optical

Sweat

Non

-inv

asive

–8.9×

10−

12mol

L−1

––

Cai

etal.(20

10)

LOx

Electroc

hemical

Saliv

aNon

-inv

asive

–0.01

mM

Prussian

blue

(PB)

Electrod

eposition

Kim

etal.(20

14)

LOx

Optical

Saliv

aNon

-inv

asive

–5×

10−

4M

–Cross-link

ing

Balle

staClave

ret

al.(20

09)

LOx

Electroc

hemical

Tear

Minim

um-inv

asive

53µA

cm−

2mM

−1

–GA

Cross-link

ing

Thom

aset

al.(20

12)

LOx

Electroc

hemical

––

45µA

cm−

2mM−

1–

Line

arpo

ly(ethyle-

nimine)

(LPE

I)Crosslin

king

Hicke

yet

al.(20

16)

LOx

Electroc

hemical

Serum

Inva

sive

172.9µA

cm−

2mM

−1

0.3µM

–ElectrostaticInteraction

Uzu

nogluet

al.(20

16)

LOx

Electroc

hemical

––

15.6

µAcm

−2mM

−1

12µM

–Adsorption

Zhao

etal.(20

14)

LOx

Optical

Serum

Inva

sive

–4.5×

10−

8M

––

Huet

al.(20

14)

LDH

Electroc

hemical

Serum

Inva

sive

–55

0µM

MB-Reine

ckeSa

ltCross-link

ing

Pian

oet

al.(20

10)

LDH

Electroc

hemical

Serum

Inva

sive

–0.1µM

–Electrod

eposition

Batraet

al.(20

16)

LDH

Electroc

hemical

Serum

Inva

sive

7.67

µAmM

−1

5µM

–Cov

alen

tBind

ing

Teym

ourian

etal.(20

12)

LDH

Optical

Serum

Inva

sive

0.40

FmM

−1

0.02

3mM

–Cross-link

ing

Liet

al.(20

17)

LOx

Electroc

hemical

––

42.42µA

cm−

2mM

−1

60µM

–Electrod

eposition

Tuet

al.(20

16)

LOx

Electroc

hemical

Plasma

Inva

sive

–0.25

µM–

Cov

alen

tBind

ing

Dag

aran

dPu

ndir

(201

7)LO

xElectroc

hemical

––

4.54

µAcm

−2mM

−1

18.3

µMMesop

orou

ssilic

aElectrop

olym

eric

entrap

men

tSh

imom

uraet

al.(20

12)

LOx

Electroc

hemical

Bloo

dInva

sive

0.53

7µA

M−1

0.8µM

Muc

in/album

inhy

drog

elCross-link

ing

Rom

eroet

al.(20

10)

LDH

Electroc

hemical

––

49.7

mV/d

ecad

e2×

10−

7M

–Cov

alen

tBind

ing

Lupu

etal.(20

07)

LDH

Electroc

hemical

Bloo

dInva

sive

3.46

µAcm

−2mM

−1

100µM

Meldo

la'sBlue

(MB)

Cross-link

ing

Pereiraet

al.(20

07)

LDH

Electroc

hemical

Bloo

dInva

sive

571.19

µAmM

−1

0.00

4µM

–Electrostaticinteraction

Nesak

umar

etal.(20

13)

LOx

Optical

–Non

-inv

asive

–0.5mM

–Adsorption

Rod

aet

al.(20

14)

F. Alam et al. Biosensors and Bioelectronics 117 (2018) 818–829

822

transcutaneous connection. Optical lactate sensors reported in the lit-erature are mostly into two major categories: (i) electro-chemilumi-nescence based lactate sensors and (ii) fluorescence based lactate sen-sors.

In electro-chemiluminescence (ECL) sensors, photons produced byrelaxation of excited molecules in an electrochemically initiated reac-tion are collected and analyzed. The photon intensity is directly pro-portional to the concentration of one or all the reactants involved in theelectrochemical reaction (Marquette et al., 2000). This technique hasbeen used for enzymatic systems of LOx /peroxidase/lumino (Girottiet al., 1991), and LDH/NADH/bioluminescent with different types ofoptodes (Xiao et al., 2017). Light emission occurs in chemiluminescencetransducers without any excitation, as the energy is released in thechemical reaction in presence of O2 or oxidation of H2O2. Such bio-sensors rely on the chemiluminescent reaction between H2O2 and lu-minol production in the enzymatic reactions.

When lactate is oxidized under catalysis of immobilized LDH and PAoxidase (PyOD) employing NAD+ as co-enzyme, electroactive H2O2 isproduced, which enhances the ECL of luminol to assist in detection. Asimple, efficient and versatile biosensing platform has been reportedthat capable of multiplexed detection of lactate; along with glucose andcholine, developed from ECL, emitted on the same pole from luminoland in situ generated H2O2 (Cai et al., 2010). The sensing bipolarelectrode (BPEs) arrays were successfully applied to determine glucose,lactate and choline in the ranges of 0.01–1mM, 0.01–1mM and0.02–5mM providing LOx as 7.57 μM, 8.25 μM and 43.19 μM, respec-tively from corresponding enzymes. Although chemiluminescence lacksselectivity and depends on environmental conditions such as tempera-ture and pH, it offers simple, low-cost instrumentation, lower detectionlimits, wide dynamic range, higher sensitivity and selectivity with goodspatial control and low signal to noise ratio.

Fluorescence based biosensors rely on either the intensity of theemitted light or the lifetime of fluorescence after a period of excitation.Fluorescence spectroscopy is one of the most useful analytical tools inbioanalysis and it causes little or no damage to the host system. Manyfluorescent probes, such as europium coordination complexes (Ferreret al., 1990), Amplex, quantum dots (Jin et al., 2010), gold nanoclusters(He et al., 2006), and cationic conjugated polymers (Hu et al., 2014),have been developed for the detection of H2O2 (Li et al., 2017). A de-sign for the synthesis of quantum-dot(QD)-hydrogel-based fluorescentprobe for bio-sensing and bioimaging of extracellular lactate has beenreported (Zhang et al., 2016). Their approach provided high selectivityand sensitivity using fluorescence resonance energy transfer mechanismby surface engineering the destabilized QDs sol with Nile Blue. Anotherhydrogel-based enzymatic lactate sensor has been reported as shown inFig. 2c (Biswas et al., 2017), which included enzymatic microsensorembedded in a hydrogel matrix. The encapsulated LOx acted as thebioactive component while phosphorescent metalloporphyrin was usedas the optical transducer. The permeability of lactate into the sensingsystem was controlled by coated polyelectrolyte multilayers on thesensor surface. The average sensitivity was determined to be11 ± 0.90% dL mg−1. The sensor is highly stable and was capable toperform without any loss till 20 cycles (shown in Fig. 2c). Holographiclactate sensors were also explored with synthetic receptors for con-tinuous monitoring of L-lactate in in-vivo biosensing (Felicity et al.,2006). Changes in replay wavelength of these sensors characterizeswelling behavior of the biofluid as a function of L-lactate concentra-tion. With L-lactate, acidic boronate receptor (3-APB) was seen to dis-play increased holographic response. An enzymatic sensor based onLDH and nanoscale optical fiber with high spatial resolution has beenreported to monitor lactate in cancer cells (Zheng et al., 2010). It de-monstrated the ability of the sensor to distinguish from abnormal ex-tracellular lactate levels in cancer cells. Near-infrared light can passthrough several centimeters of tissue, with the appropriate choice offluorophore in detection. The molecules can be excited, investigatingemissions from outside the body (Lakowicz, 2006), providing a

potential for non-invasive sensing. Special fluorescence techniques canprovide information about the structure and micro-environment ofmolecules; and their change in response to analyte variations can bemonitored. However, this mechanism has its disadvantages of rapidphoto-bleaching of fluorescent organic dyes conjugated to the biomo-lecules of interest; and a potential loss in activity of biomolecules on thebio-chemical conjugation with the fluorescent dye.

Different lactate sensing techniques and their performance aresummarized in Table 1.

4. Lactate detection protocols

Detection of lactate can be performed through either invasive ornon-invasive mechanisms from relevant bio-fluids such as blood, in-terstitial fluid, saliva, tear, breath and sweat. The lactate concentrationfor a healthy person ranges in blood, interstitial fluid, saliva, tear fluid,breath and sweat are 0.5–2mM, 1.9–2.2 mM, 0.1–2.5 mM, 1–5mM,13.5–22 µM and 10–25mM respectively.

4.1. Invasive detection techniques

Blood is the major biofluid for invasive lactate detection. L-Lactatelevels in blood raises as high as 12mM (Goodwin et al., 2007) duringexercise from the healthy subject at rest. Blood lactate concentration innormal unstressed individuals lies between 0.5 and 1mM; and thosewith critical illnesses have concentrations below 2mM (Scrutton,1976).

Blood, a primarily biofluid for measuring lactate needs a specialprotocol for sample collection to prevent changes in lactate during andafter the blood sample is invasively drawn for measurement. Samplesshould be analyzed immediately on drawing, as concentrations canincrease by 70% within 30min at room temperature through glycolysis(Tietz et al., 2006). The sampling bio-matrix is known to affect thebiosensor response, which is also affected by the functionalizationthrough membranes or films to prevent interference. Electrochemicalsensors require sample preparation protocols which ensure: (i) removalof electrochemically active compounds which can act as interefrents(e.g. uric acid, ascorbic acid and dopamine can be removed fromserum/plasma (Brahim et al., 2001) (ii) adjustment of ionic strengthand temperature (Zhang et al., 2008) since variable ionic strengthswhich influence measurements; (iii) removal of other surface foulingcompounds from plasma proteins, lipids, or other biochemical compo-nents from the matrix (Botasini et al., 2013); and (iv) adjustment of pH(Henderson, 1982). An automated lactate analyzer based on electro-enzymatic method with continuous blood sampling through a catheterwas developed to monitor lactate concentrations in blood and to detectsmall changes in lactate concentrations during physical exercise,(Shimojo et al., 1991). Determination of lactate in human serum sam-ples indicated that the fabricated sensors can be used to continuouslymonitor levels in real time. In an invasive optical sensing from blood,sensing of analytes can enzymatically generate hydrogen peroxide.Successful applications to this approach has been seen using humanserum samples as well. An implantable dual sensing biochip was de-signed for lactate sensing (Guiseppi-Elie, 2011). This bio-transducerwas employed for monitoring stress levels during long term space ex-ploration, shown in Fig. 3a. In the sensor, bioactive hydrogels wereencapsulated in a micro-disc electrode array with electro-polymeriza-tion of pyrrole within it. The bio-chip was biocompatible and can beoperated in a linear dynamic range for the lactate assay till 30-foldincrease of the concentration with high operational stability.

4.2. Commercially available products for invasive lactate sensing

The three most commonly used automated analyzers in laboratoriesare: (i) Biosen analyzers and (ii) Yellow Spring Instruments (YSI) 2300Stat analyzer, and (iii) EBIO Plus analyzer which have price tags in the

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range of $3 K-$10 K. These POC devices follow enzymatic electro-chemical principle for lactate measurement. For example, Biosen ana-lyzers measures lactate from whole blood, plasma and serum and pro-vides response in 20–25 s up to 160 results / hour. The samples remainstable up to 5 days if refrigerated. Substantial progress led to the

development of commercial hand-held blood lactate sensors, such asLactate-Pro (LP) in Fig. 3b. These sensors can be in the size of a creditcard and capable of quickly measuring lactate with only a small sampleof blood. Only a 5 µL blood sample is required and serves speedymeasurements in 60 s with high performance in a small size. Lactate

Fig. 3. (a) The ECC MDEA 5037 dual responsive biotransducer and associated 2-channel wireless potentiostat. Cell A glucose responsive hydrogel and cell B lactateresponsive hydrogel (Guiseppi-Elie, 2011) and Comparison of performances of (b) lactate-Pro I, (c) Lactate Scout I, (d) lactate Plus I, (Tanner et al., 2010) (e) EBIOPlus vs. Lactate-Pro (Baldari et al., 2009) (f) Accuaport vs. Lactate-Pro, (g) ABL700 vs. Lactate- Pro, and (h) YSI 2300 Stat vs. Lactate-Pro (Pyne et al., 2000).

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Scout (LS from Sports Resource Group, Inc.) in Fig. 3c, have been va-lidated in the sport settings using standardized testing protocols andmethodologies, making them common for lactate threshold (LT) testing.LS is used by clinicians around the world for easy, reliable measure-ments in medicine. This device needs 0.2 µL of blood providing resultsin 10 s 250 results can be stored on this device which features a stop-watch and counts downtimes for measures. It operates in temperaturesfrom 5 to 45 °C up to 85% humidity. Lactate Plus (L+) Meter is anothercompact, hand-held device for POC quantitative measurements inblood. With Nova lactate strips, L+ can be used in sports medicine toassess physical performance and establish optimum training regimens.This measures blood lactate in 13 s using a small blood drop. This re-sults in a virtually painless, yet fast and accurate test. StatStrip is ahandheld, point-of-care lactate sensing system that provides rapid re-sponse in minimal turnaround time (13 s) requiring smallest wholeblood sample (0.6 µL), and is easily operable. This low-cost devicemakes detection practical and affordable for the user.

Most sensors rely on blood bio-matrix for accurate measures.However, their intrusive nature poses a hindrance for patients impedinginformation acquisition desired for diverse biomedical applications. InFig. 3b- d, LP, LS, and L+ were assessed. The suitability of the LP and L+ as accurate analyzers are supported by strong correlations (r= 0.98and r= 0.988) and limits of agreement ≤ 2.1 mM. The correlationcoefficient of LS was found 0.91. This study revealed that the LP and L+ performed better reliability, precision and accuracy compared to LSalthough the LS also demonstrated relatively good reliability (Tanneret al., 2010). Similarly, the correlations between the Lactate-Pro withdifferent lactate measurement tools: EBIO Plus (Fig. 3e), Accusport(Fig. 3f), ABL 700 Series Acid-Base analyzer (Fig. 3g), and YSI 2300(Fig. 3h) were estimated r= 0.984, r= 0.97, r= 0.98, r= 0.99, re-spectively. The level of agreement between the Lactate-Pro and otheranalyzers was generally less than± 2.0mM over the physiologicalrange of 1.0–18.0mM (range of mean difference: −0.06 to 0.52mM).The higher correlation coefficients and lower limit of agreements sup-port the reliability of the existing devices.

4.3. Noninvasive detection techniques

Continuous analyte monitoring is critical of physical sensor devicesfor lactate sensing (Hammock et al., 2013). Measurement from otherbio-matrices is gaining interest recently for alternative non-invasiveanalysis. Although many sampling sites are available for non-invasivesensing, most efforts have concentrated on tear, saliva and sweat andskin interstitial fluid.

4.3.1. SweatLactate in sweat can be a biomarker for different pathological dis-

orders. Sweat lactate is a function of sweat gland energy metabolism.Under anaerobic conditions a rise in sweat lactate concentration canreduce sweat gland activity (Rassaei et al., 2014). The levels of lactatemeasured in human sweat may vary up to 25mM depending on theparts of the body, stimulation intensity and mode, sweat rate and en-vironment. (Jia et al., 2013a). Also, it varies with respect to gender andage. Its increased levels with sweat rate can be an indicator of tissuecompromise. Lactate concentrations have been observed to be elevatedespecially at the beginning of a sweating period, whether elicited bythermos-genic simulation of sweat glands or heavy workout (Weinerand Van Heyningen, 1952). It has also been studied that with sweat re-absorption in the ducts, with the water content in it getting reabsorbed;lactate remains in the ducts after a period of sweating, simulatinggreater lactate production in the beginning of a sweating period ascompared to later.

Conventional sampling methods used for sweat collection such asband-aid like device with combination of nitrocellulose membrane andfilter paper. Nitrocellulose promotes absorption of sweat but is as wellable to pass water through the filter paper during evaporation

(Imamura et al., 1994). Similarly, a liquid filled glass cup attached onthe skin surface (Mitsui, 1997) used to apply for sweat collection,however, its prolonged collection time was not feasible for wearableapplication. Contrarily, microfluidic platforms are getting popular forthe extraction, capture, and analysis of sweat for point-of-care appli-cation. Such a platform is coupled with soft hydrogels which allows tointerface with dermal layer of the skin to facilitate collection of sweat.Triggering techniques such as iontophoresis further promotes extrac-tion of sweat. Excreted sweat can be preserved through disposablegauzes, absorbent pads, Parafilm-M pouches, etc. (Choi et al., 2018)

4.3.2. SalivaSaliva is a convenient biofluid to measure lactate level in the study

of anaerobic metabolism (Segura et al., 1996). Beyond certain workoutintensity and correlating accumulation of blood lactate (anaerobicthreshold), a saliva threshold is reported to exist, defining the pointwhere salivary α-amylase and electrolytes rise above their baselines(Chicharro et al., 1998). Lactate in saliva has high correlation withblood lactate with the ratio of ~1: 4 (Santos et al., 2006), which sets itslevel in human saliva ~ 0.2mM (Guilbault et al., 1995). Thus, salivacan be used as a key diagnostic tool in clinical and point of care settingsto provide lactate level along with other valuable information about thebiochemical, metabolic and functional status of an individual. Salivasampling has the advantages of sparing repeated hassles of skin or veinpuncturing over venous or capillary sampling making it more con-venient. Saliva is a biomatrix from where sampling involves less pre-treatment. These merits attract the attention of researchers for in vitrodiagnostics.

4.3.3. TearLactate is one of the principal bio-analytes in the basal tear film with

a typical concentration of 2–5mM in the tear fluids that provides asecure analytical environment for noninvasive monitoring. Biomarkerslike urea, ascorbic acid and L-lactate in the tear film can be used forcontinuous, non-invasive surveillance of certain health conditions in apatient (Van Haeringen and Glasius, 1977) (Baeyens and Gurny, 1997).Formation of lactate has been found to be around 0.50 µmol/h in intactcorneas and half of the same value in deepithelialized corneas. Lactate,originating generally from the corneal epithelium is about four to tentimes higher in tears than serum (Waterman, 1991). L-lactate travelsfrom the corneal epithelium to stroma through lactate–proton ex-change, spreading further to endothelium and crossing into tear film(Giasson and Bonanno (1994)). LDH in tears still has not appeared to beuseful for diagnosing retinoblastoma although different approaches forthe suppression of interfering species in the tear film are evaluated.

4.4. State of the art in noninvasive sensing

There has been fast increasing research and development effort fornon-invasive POC sensing methods and tools especially for their po-tential for integrating with wearable sensor platforms. One promisingapproach for noninvasive lactate sensing is epidermal sweat analysis. Anoninvasive tattoo-based lactate sensor fabricated on a flexible sub-strate was demonstrated (Jia et al., 2013a). The sensor was functiona-lized with LOx for the continuous and real time monitoring of lactatelevels in human sweat as shown in Fig. 4a. The resulting temporallactate profiles reflect the changes in the production of sweat lactateupon varying the exercise intensity with a linearity up to 20mM ofconcentration. Fig. 4a (middle) represents real-time response of lactatefrom the tattoo biosensor during cycling. Red dots represent the lactateconcentrations in sweat whereas Fig. 4a (right) shows the in vitro ca-libration curve at 37 °C (inset shows the reported amperometric re-sponse to different lactate concentrations up to 30mM with 5mM in-crements. An organic electrochemical transistor (OECT) sensor with anionogel solid-state electrolyte on a flexible biosensor was presented forthe first time to detect lactate in a relevant physiological range as

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shown in Fig. 4b (left) (Khodagholy et al., 2012). The sensor can beused as a wearable bandage-type sensor, which can be worn duringexercise or health monitoring, allowing sweat to diffuse into the sensorwith consequent detection of the lactate analyte. An ECL based lactatebiosensor with a detection limit of 8.9× 10−12 mol/L was observed forreal sweat samples (Cai et al., 2010). The biosensor was assessed fordetecting lactate from sweat of athletes during workout. Fig. 4b depictsthe normalized response of this transistor (as a function of lactate

concentration between 10 and 100mM), which shows detection in therelevant physiological range of sweat, suggesting its application in sportscience and healthcare.

Lately, other types of noninvasive POC sensors with lightweight andflexible nature have been studied for wearable platforms (Labroo andCui, 2013). For instance, contact lenses provide a safe, noninvasiveplatform for collection of biological information eliminating samplingprocedures. A minimally invasive contact lens- based lactate sensor was

Fig. 4. (a) Tattoo lactate biosensor with sensing characteristics (Jia et al., 2013a) (b) Conformal OECT on forearm and normalized response of the OECT vs. lactateconcentration (Khodagholy et al., 2012) (c) Contact lens based L-lactate sensor and its performance for monitoring of l-lactate levels in tear fluid (Thomas et al.,2012) (d) Eyeglasses biosensor system, which integrates a wireless circuit board along the arms of the spectacles and two electrochemical sensors for lactate andpotassium on to the nose-bridge pads, monitoring of sweat lactate (top-left) and calibration curves for the lactate biosensor and enzyme-free control sensor before on-body tests (top- right) (Sempionatto et al., 2017) (e) mouth-guard biosensor with sensing characteristics (Kim et al., 2014).

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fabricated for monitoring lactate levels in tear fluid for personalhealthcare and assessment as shown in Fig. 4c (left) (Thomas et al.,2012). The platinum sensing structures were functionalized by cross-linkage of LOx with GA and bovine serum albumin, and coated withmedical grade polyurethane for detecting lactate well within the phy-siological range. Its dual sensor design also was reported to have hadminimal interference with a Nafion membrane and functional at tem-peratures close to that of the eyes. Fig. 4c (right) depicts the sensorresponse as function of L-lactate concentration. The current increasesproportionally for single sensors between 0 and 5mM L-lactate, withsufficient resolution and accuracy for clinical measurements. Havingbeen built on a polymer substrate, this also provides good biocompat-ibility with the eyes to be worn like a regular contact lens. Differentisoenzymatic forms of PA, which is produced by glycolysis and con-verted to lactate by LDH; are present in a contact lens and under normalaerobic conditions it has been observed that around 70% of the glucoseis converted to lactate. In an alternative approach, an amperometriclactate biosensor connected to nose-bridge pads was designed as shownin Fig. 4d (Sempionatto et al., 2017). This can be interfaced with anelectronic backbone on spectacle glass arms and can continually senselactate for a few hours. It's positioning on separate nose pads minimizedcross-talk and enables isolated fabrication and replacement. It can re-portedly transmit data wirelessly through Bluetooth to a remote mobilehost device for data analysis and visualization. Here, the nose-bridgepads are interchangeable with multifunctional sensing applications.Different authors have reported the possibility of determining theanaerobic threshold through saliva lactate concentration in incrementalprotocol and exercise using cycle ergometer verifying high correlationsbetween that in blood and in saliva (Pérez et al., 1999). An ampero-metric lactate sensor from where lactate concentration levels were de-termined from the saliva was demonstrated during workout in real-timeand noninvasive manner (Schabmueller et al., 2006). Saliva weredrawn due to the capillary effect to expedite the detection of lactate.This approach allowed for portability, with permanent real-time mea-surements and rapid response time. Another wearable metabolitesensor was reported for detecting lactate in saliva, incorporating aprintable enzymatic electrode on a flexible PET (polyethylene te-tephthalate) substrate on a mouth-guard as in Fig. 4e (left) (Kim et al.,2014). This biosensing system was designed on a printable PB trans-ducer using poly-orthophenylenediamine (PPD)/lactate-oxidase (LOx).This non-invasive mouth-guard sensor targets continuous monitoring oflactate enabling specific detection with high stability and sensitivity.Fig. 4e (middle) indicates response for increasing concentrations oflactate in 0.1mM increments (b-k) in PBS. Its calibration (inset) ex-hibits high linearity (slope, 0.553 µAmM−1; correlation coefficient,0.994). Fig. 4e (right) indicates that the sensor responds favorably tochanges in lactate (b-f). A disposable electro-chemiluminescent bio-sensor for saliva lactate through enzymatic recognition was demon-strated based on LOx (Ballesta Claver et al., 2009). It provided responsefrom 10−5 to 5×10−4 M with a detection limit of 5×10−6 M andprovided an RSD of 3.30%. This saliva based lactate, prepared by hy-brid materials of ZnO NPs and multiwalled carbon nanotubes (MWCNT)in a glassy carbon electrode surface and then modified them with LOxand, show good repeatability, quick detection time (20 s) and versati-lity.

4.5. Commercially available products for noninvasive lactate sensing

A commercially available wearable lactate threshold (WLT) sensoris BSXinsight, (BSX Athletics, Texas, USA), as shown in Fig. 5a, which isbased on portable NIR (near infrared) light emitting diode (LED) device(Driller et al., 2016). It was marketed as the first wearable lactatethreshold sensor claiming to determine lactate threshold levels duringworkout, without the need for invasive blood sampling that costs in therange of $299 US- $419 US. This non-invasive and portable device al-lows athletes and coaches to monitor their lactate levels with minimal

expertise. Kenzen Patch (http://kenzen.com) is a commercially avail-able lactate sensor (as shown in Fig. 5b) in the form of a reusable smart-patch which can adheres to the skin, to analyze certain electrolytes likeLA and glucose from sweat, along with pH. This information is pro-cessed and wirelessly sent to the user's phone, notifying them know oftheir energy stores.

5. Challenges and future trends of wearable non-invasive lactatebiosensors

5.1. Challenges of wearable lactate biosensors

Numerous challenges still exist for wearable non-invasive lactatesensors. Resiliency, long-term stability, selectivity, sensitivity, lowerdetection limits, power consumption and biocompatibility are issueswhich still need to be addressed. Epidermal sensors face mechanicaldeformations during ergonomic movements which increases when thewearer is under intense physical strains. Such challenges arise withtextile-based sensors, where textiles can be subject to immense me-chanical, chemical, and heat degradations (Bandodkar and Wang,2014). Extensive efforts need to be taken to withstand such severestresses over rigorous usage. Such efforts can greatly benefit from theconsiderable advances that have been made in flexible electronics(Rogers et al., 2010). Researchers exploring wearable electrochemicalsensors look toward addressing such resiliency issues cost effectively.Mechanical stretchability and resiliency have been improved sig-nificantly in a textile-based bio-fuel cell lactate sensor where it showedstable output after 100 cycles of 100% stretching. High surface area ofCNT leads also to enhance stable adsorption of the LOx and 1, 4-naphthoquinone. Printable tattoo-based electrochemical sensors havemajor challenges of calibration, fouling and related offsets and stabilityand on-body durability under mechanical stress (Windmiller et al,2012). Such devices have limited stretchability which should be im-proved in connection to new low-cost large scale fabrication techni-ques. Leaching of reagents and need for re-calibration are some of theother challenges that researchers will face when developing long-lasting tattoo-based electrochemical devices. Stability and selectivity ofenzymatic sensors are crucial parameters for POC monitoring affectingsensor's reliability. Multiple parameters, like pH, ionic strength, dif-ferent interfering analytes, and operational conditions (e.g. tempera-ture, humidity, applied voltage) can alter the sensor's performance in amulti-analyte system. Additionally, other intrinsic or extrinsic proper-ties of the sensors, such as structure alteration, phase conversion, poi-soning, degradation, bulk diffusion or interference, electrode fouling,irreversible nonspecific adsorption may cause hysteresis and affect thestability of the biosensors (Heikenfeld et al., 2018). To overcome thestability issue, pre-treatment or pre-calibration is mandatory in many oflactate sensors before they move to operation mode (Parrilla et al.,2016). Incorporation of stabilizing agents with the selective bio-re-ceptors can further improve stability of the sensors (Jolly et al., 2016).In the presence of electrolytes (e.g. Na+, K+), selective and simulta-neous detection of lactate and glucose have been executed (Gao et. al,2016). To improve selectivity, multi-sensors array can provide accuracyand precision to predict specific concentration of lactate through mu-tually exclusive sensors’ data (Bandodkar et al., 2016). The sensitivityand limit of detection can be enhanced in many folds through inclusionincorporating nanomaterials (Taghdisi et al., 2015), target recyclingand signal amplification. However, their potential toxicity of nanoma-terials requires prior consideration. Lactate sensing may require strin-gent bio-affinity protocols, with their continuous monitoring requiringreceptor regeneration, achieved by subjecting the sensor to harsh con-ditions which may not be biocompatible with ergonomic variations.Another issue marring the introduction of such a wearable bio-affinitysensor is its ability to detect ultra-low limits. Findings in nano-elec-tronics, wearable-biofuel cells (Jia et al., 2013b), piezoelectric energyharvesting (Wang and Song, 2006), and thin-film batteries may help

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address the issues of power consumption that still exist. Chemi- sensorsrequire direct contact with biofluids; while associated passive electro-nics need sealing from the aqueous surroundings. Satisfying these cri-teria, while maintaining the aesthetics of the device on a miniaturized,wearable platform is daunting.

5.2. Future trends of wearable lactate biosensors

Efforts to devise ultrasensitive sensors for lactate provide insights toclinical diagnosis and health monitoring. Enzymatic electrochemicalsensing techniques are predominant among others due to their im-proved sensitivity, lower detection limits, simpler fabrication, simpleuse, portability, reliability and reasonable cost. Nevertheless, more re-search is required to fulfill demands of continuous monitoring, whichmay include implantables in blood vessels (intravascular) or under theskin (subcutaneous), which would avoid requirement of sample dilu-tion. These sensors however face issues with biocompatibility and re-sponse time. Nano-technological advancements offer support for min-iaturization and development of semi-invasive or noninvasive sensing.Nanomaterials (metal-oxides, SWCNTs/MWCNTs), due to their ad-vantages of large surface-to-volume ratio, provide an improved surfacefor enzyme immobilization (RoyChoudhury et al., 2016). Moreover,nanomaterials like ZnO nanoflakes and nanowires with high isoelectricpoint (IEP) values, high surface reaction activity, high catalytic effi-ciency and strong adsorption abilities provide additional advantages(Alam et al., 2018).

Wearable electrochemical sensing garners attention of large in-dustrial players in bringing breakthroughs through continuous mon-itoring of biomarkers in the sector of health and wellness. With theentry of companies like Google, NovioSense, OrSense, and Electrozyme,the nascent field of wearable and non-invasive electrochemical sensorsis expected to grow rapidly, with new innovative devices. Prospects ofwearable biosensing are immense in personalized health monitoring;expecting to provide benefits of cost effective solutions for self-man-agement of diseases. With such personalized remote health-monitoringdevices, patients will be granted improved and convenient healthanalysis. Certain non-invasive sensing devices that expected to be in-troduced soon, include those for monitoring catecholamines and anti-oxidants (Choy et al., 2001) in tears; cortisol (Vasudev et al., 2013) and

pathogens (Tabak, 2007) in saliva; or amino acids and pathogens(Zelada-Guillén et al., 2012) in sweat/ISF. Fusion of several wearablesensors should lead to non-invasive multi-analyte sensing. Integrationof radio-frequency identification or Bluetooth devices will enablewireless transmission of the processed data to cellphones/computers ina user-friendly manner providing comprehensive assessment of thewearer's health status, as per stress and performance. With the cap-ability of continuous monitoring, data processing by deep learning andartificial intelligence tools will become more and more important forfast and accurate interpretation of the collected data. Increasing cloud-based data storage and processing services will improve the efficiencyand accelerate widespread adoption of wearable sensing and healthmonitoring platforms.

6. Summary and conclusion

We provided an extensive overview of the technological progress inL lactate detection, particularly through wearable sensors. After brieflycovering the clinical importance of the level of human lactate level, itsinteractions with two enzymes, LOx and LDH, were explained as theyare commonly employed in lactate sensing. Existing invasive and non-invasive techniques of detection from various bio-fluid matrices havebeen discussed among which electrochemical methods have been re-ported to show more promise in simple, direct analysis, using enzymespartaking in its metabolic pathway. A comparison has been drawn onthe detection mechanisms and sensing performances of the variousexisting lactate biosensors. Advancements in nano-scale materials andprocessing techniques greatly benefited the lactate sensing just likeother sensors, electronic and photonic devices. These new materials andtechniques increased the performance of the sensors and allowed use ofpreviously dismissed bio-fluid matrices such as sweat and tear anddemonstration of novel non-invasive sensing methods such as tattooand contact lens integrated sensors. However, there still are challengesyet to be overcome to achieve better selectivity, stability, reliability andlifetime for wearable sensing platforms. Rapid expansion of mobiledevices and services integrated to them are expected to attract in-creasing investment in wearable health and wellness monitoring plat-forms which will most likely include integrated lactate level detectorsas well. These integrated efforts will transform the future of personal

(a)

(b)

Fig. 5. (a) The wearable lactate threshold sensor (WLT) (Driller et al., 2016) (b) Kenzen Patch (http://kenzen.com).

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healthcare in a way never imagined before.

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