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Uncertainty in Antibiotic Dosing in Critically Ill Neonate and Pediatric Patients: Can Microsampling Provide the Answers?

      Abstract

      Purpose

      With a decreasing supply of antibiotics that are effective against the pathogens that cause sepsis, it is critical that we learn to use currently available antibiotics optimally. Pharmacokinetic studies provide an evidence base from which we can optimize antibiotic dosing. However, these studies are challenging in critically ill neonate and pediatric patients due to the small blood volumes and associated risks and burden to the patient from taking blood. We investigate whether microsampling, that is, obtaining a biologic sample of low volume (<50 μL), can improve opportunities to conduct pharmacokinetic studies.

      Methods

      We performed a literature search to find relevant articles using the following search terms: sepsis, critically ill, severe infection, intensive care AND antibiotic, pharmacokinetic, p(a)ediatric, neonate. For microsampling, we performed a search using antibiotics AND dried blood spots OR dried plasma spots OR volumetric absorptive microsampling OR solid-phase microextraction OR capillary microsampling OR microsampling. Databases searched include Web of Knowledge, PubMed, and EMbase.

      Findings

      Of the 32 antibiotic pharmacokinetic studies performed on critically ill neonate or pediatric patients in this review, most of the authors identified changes to the pharmacokinetic properties in their patient group and recommended either further investigations into this patient population or therapeutic drug monitoring to ensure antibiotic doses are suitable. There remain considerable gaps in knowledge regarding the pharmacokinetic properties of antibiotics in critically ill pediatric patients. Implementing microsampling in an antibiotic pharmacokinetic study is contingent on the properties of the antibiotic, the pathophysiology of the patient (and how this can affect the microsample), and the location of the patient. A validation of the sampling technique is required before implementation.

      Implications

      Current antibiotic regimens for critically ill neonate and pediatric patients are frequently suboptimal due to a poor understanding of altered pharmacokinetic properties.
      An assessment of the suitability of microsampling for pharmacokinetic studies in neonate and pediatric patients is recommended before wider use. The method of sampling, as well as the method of bioanalysis, also requires validation to ensure the data obtained reflect the true result.

      Keywords

      Introduction

      Severe infection in neonate and pediatric patients is a significant cause of mortality worldwide.
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      Effective antibiotic therapy is essential to the resolution of an infection causing sepsis.
      However, the use of antibiotics in critically ill neonate and pediatric patients is poorly understood, with evidence suggesting that current dosing is frequently inadequate.
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      Meropenem Pharmacokinetics in Critically Ill Children.
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      Augmented renal clearance implies a need for increased amoxicillin-clavulanic acid dosing in critically ill children.
      Recommended neonate and pediatric antibiotic dosing regimens are often extrapolated from healthy adult data using basic empiric scaling factors, such as body weight or body surface area.
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      A Europe-wide point prevalence study that included 89 neonatal intensive care units from 21 countries found that 75% of vancomycin doses were below the recommendations.
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      • et al.
      High variability in the dosing of commonly used antibiotics revealed by a Europe-wide point prevalence study: implications for research and dissemination.
      Describing pharmacokinetic properties in a pediatric patient traditionally requires the sampling of 1 to 5 mL of whole blood, taken 5 to 8 times within a dosing interval, from a cannula or by venipuncture. There are challenges associated with performing pharmacokinetic studies in neonatal and some pediatric patients due to their small blood volumes and the psychological burden associated with blood sampling.
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      Studies have found that pediatric intensivists are in ethical conflict about performing potentially life-saving drug research, even while identifying them as ethically acceptable.
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      Yet, this information is important for optimizing the treatment of their patients.
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      • Moore W.
      • Chopra A.
      Meropenem Pharmacokinetics in Critically Ill Children.
      • De Cock P.A.
      • Standing J.F.
      • Barker C.I.
      • et al.
      Augmented renal clearance implies a need for increased amoxicillin-clavulanic acid dosing in critically ill children.
      • Jacqz-Aigrain E.
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      • van den Anker J.N.
      How to Optimize the Evaluation and Use of Antibiotics in Neonates.
      • Cies J.J.
      • Moore 2nd, W.S.
      • Dickerman M.J.
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      Pharmacokinetics of continuous-infusion meropenem in a pediatric patient receiving extracorporeal life support.
      Innovation in the quantitative analysis of clinical samples, led by improved sensitivity of analytical methods, has reduced blood sample volumes to “microsamples.”
      • Parker S.L.
      • Dorofaeff T.
      • Lipman J.
      • et al.
      Is there a role for microsampling in antibiotic pharmacokinetic studies?.
      Microsampling uses a low volume of sample (<50 μL), with some acquired by skin prick. Microsamples can require frozen storage or be dried for transport and storage.
      The aim of this paper was to review the current knowledge of antibiotic dosing in critically ill neonates and pediatric patients. From this, we investigated whether microsampling can improve opportunities to conduct pharmacokinetic studies and improve antibiotic dosing in these patients.

      Methodology

      We undertook a literature search to find relevant articles using the following search terms: sepsis, critically ill, severe infection, intensive care AND antibiotic, pharmacokinetic, p(a)ediatric, neonate. Specifically, these were articles that described pharmacokinetic properties and antibiotic concentrations in critically ill neonatal and pediatric patients. Additionally, we included papers relating to adult patients when they added important and relevant information. For microsampling, we performed a search using the following terms: antibiotics AND dried blood spots OR dried plasma spots OR volumetric absorptive microsampling OR solid-phase microextraction OR capillary microsampling OR microsampling. Databases searched include Web of Knowledge, PubMed, and EMbase.
      Of the 32 antibiotic pharmacokinetic studies performed on critically ill neonate or pediatric patients in this review, most of the authors identified changes to the pharmacokinetic properties in their patient group and recommended either additional investigations into this patient population or therapeutic drug monitoring to ensure antibiotic doses are suitable.

      Altered Pharmacokinetics in Critically ill Neonate and Pediatric Patients

      A number of physiologic changes occur within the body as a result of both sepsis and the medical interventions required for treatment. Roberts et al
      • Roberts J.A.
      • Abdul-Aziz M.H.
      • Lipman J.
      • et al.
      Individualised antibiotic dosing for patients who are critically ill: challenges and potential solutions.
      identified the effects of critical illness in adults that have the potential to affect antibiotic pharmacokinetic properties as altered hyperdynamic function, altered fluid balance, organ dysfunction, and organ support. Changes to patient pathophysiology can be described in terms of the alterations to the pharmacokinetic parameters of clearance and Vd. These changes can directly influence the antibiotic concentrations that are available to fight infection. A summary of the pharmacokinetic studies in critically ill neonate and pediatric patients described in this section is included in the Table.
      TableReview of literature for pharmacokinetic data on critically ill neonate and pediatric patients receiving antibiotics.
      Antibiotic ClassFirst AuthorStudy PopulationAntibioticSamples/ PatientAge, yDose, mg/kgFactors in StudyPatients, nPK Parameter
      Vd, L/kgt1/2, hCL, L/h/kgCmax, mg/LCmin, mg/LAUC, mg/h/L
      β-LactamBradley
      • Bradley J.S.
      • Sauberan J.B.
      • Ambrose P.G.
      • et al.
      Meropenem pharmacokinetics, pharmacodynamics, and Monte Carlo simulation in the neonate.
      Critically ill neonatesMeropenem3; unknown volume; various access
      Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      23 GA to 62 PA10 or 20NA370.402.90.104
      Cies
      Data from abstract only.
      • Cies J.
      • Moore W.
      • Chopra A.
      Meropenem Pharmacokinetics in Critically Ill Children.
      Critically ill pediatricsMeropenem0.25 to 929NA110.651.40.384
      Cies
      • Cies J.J.
      • Shankar V.
      • Schlichting C.
      • et al.
      Population Pharmacokinetics of Piperacillin/Tazobactam in Critically Ill Young Children.
      Critically ill pediatricsPiperacillin2–4; unknown volume0.75 to 675 to 106.4NA130.2491.390.299
      Cohen-Wolkowiez
      • Cohen-Wolkowiez M.
      • Watt K.M.
      • Zhou C.
      • et al.
      Developmental pharmacokinetics of piperacillin and tazobactam using plasma and dried blood spots from infants.
      Critically ill neonatesPiperacillin2–7; 0.2 mL/sample<32 GA to >32 PA80 every 8 hGA<32, PA<14120.425.30.05538.3
      GA<32, PA>1490.422.50.11634.6
      GA>32, PA<1480.422.80.10428.0
      GA>32, PA>3230.424.50.06531.1
      Kongthavonsakul
      • Kongthavonsakul K.
      • Lucksiri A.
      • Eakanunkul S.
      • et al.
      Pharmacokinetics and pharmacodynamics of meropenem in children with severe infection.
      Critically ill pediatricsMeropenem4; peripheral line 5 mL/sample4 to 1220 every 8 hNA140.200.286
      Calculated as /kg based on mean weight of patient population reported.
      Nehus
      • Nehus E.J.
      • Mizuno T.
      • Cox S.
      • et al.
      Pharmacokinetics of meropenem in children receiving continuous renal replacement therapy: Validation of clinical trial simulations.
      Critically ill pediatrics on CRRTMeropenem6–11; unknown volume5 to 2113.8 to 22 every 12 hNA70.354.33.7
      Calculated from mL/(min/1.73m2) using a mean surface area of 1.40 m2.
      80.1
      Nichols
      • Nichols K.
      • Chung K.
      • Knoderer C.A.
      • et al.
      Population Pharmacokinetics and Pharmacodynamics of Extended-Infusion Piperacillin and Tazobactam in Critically Ill Children.
      Critically ill pediatricsPiperacillin6; 0.5-mL/kg sample, 5 mL maximum0.75 to 11100 every 8 h, 4 h infusionNA120.430.22
      Tazobactam0.370.19
      Santos
      • Santos S.R.
      • Sanches-Giraud C.
      • Silva C.V.
      • et al.
      Pharmacokinetic-pharmacodynamic correlation of imipenem in pediatric burn patients using a bioanalytical liquid chromatographic method.
      Pediatrics burnsImipenem5; venous catheter 1 mL/sample1 to 90.5 g every 6 hNA60.231.450.138
      PenicillinDe Cock
      • De Cock P.A.
      • Standing J.F.
      • Barker C.I.
      • et al.
      Augmented renal clearance implies a need for increased amoxicillin-clavulanic acid dosing in critically ill children.
      Critically ill pediatricsAmoxicillin-clavulanic acid5; predefined maximum volume: 2.4 mL/kg0.08 to 1525 to 35 every 6 hAmoxicillin-50V1 0.13; V2 0.08; V3 0.160.26
      clavulanic acidV1 0.17; V2 0.140.17
      Muller
      • Muller A.E.
      • DeJongh J.
      • Bult Y.
      • et al.
      Pharmacokinetics of penicillin G in infants with a gestational age of less than 32 weeks.
      Critically ill neonatesPenicillin5; 0.2 mL/sample26 to 32 GA30Day 3 after birth200.45
      Calculated as /kg based on mean weight of patient population reported.
      3.90.086
      Calculated as /kg based on mean weight of patient population reported.
      FluoroquinoloneLeroux
      • Leroux S.
      • Turner M.A.
      • Barin-Le Guellec C.
      • et al.
      Pharmacokinetic Studies in Neonates: The Utility of an Opportunistic Sampling Design.
      Critically ill neonatesCiprofloxacin2–3; predefined maximum volume: 1.2 mL + scavenged samples<40 wk PA 10 every 12 h; >40 wk PA 10 every 8 hScheduled sampling60V1 1.28
      Calculated as /kg based on mean weight of patient population reported.
      0.330
      V2 0.84
      Calculated as /kg based on mean weight of patient population reported.
      Scavenged samplingV1 0.307
      Calculated as /kg based on mean weight of patient population reported.
      0.344
      V2 1.76
      Calculated as /kg based on mean weight of patient population reported.
      Lipman
      • Lipman J.
      • Scribante J.
      • Gous A.G.S.
      • et al.
      Pharmacokinetic profiles of high-dose intravenous ciprofloxacin in severe sepsis.
      Critically ill pediatricsCiprofloxacin6–9; 0.5 mL0.25 to 520A: 3–12 mo; days 0, 2, and 7202.06. 1.49, 2.053.67, 3.3, 4.233.82, 3.48, 4.646.08, 9.03, 5.810.21, 0.21, 0.1615.6, 19.2, 14.1
      B: 1–5 y; days 0, 2, and 71.44, 1.43, 1.762.84, 3.13, 2.826.65, 6.15, 8.167.38, 7.78, 6.380.14, 0.21, 0.1015.9, 18.0, 13.2
      CephalosporinAhsman
      • Ahsman M.J.
      • Wildschut E.D.
      • Tibboel D.
      • et al.
      Pharmacokinetics of cefotaxime and desacetylcefotaxime in infants during extracorporeal membrane oxygenation.
      Critically ill pediatrics on ECMOCefotaxime10; 0.7 mL0 to 0.7550–150 daily, every 6 hCefotaxime370.523.50.103
      Des-CTX3.145.40.417
      Olguin
      • Olguín H.
      • Asseff I.
      • Vieyra A.
      • et al.
      Effect of severity disease on the pharmacokinetics of cefuroxime in children with multiple organ system failure.
      Critically ill pediatricsCefuroxime11; unknown volume0.3 to 14100 every 8 hinfection, not unwell41.50.55116.4
      Severe sepsis, not intubated51.60.48121.6
      Severe sepsis, intubated63.11.87190.7
      Trang
      • Trang J.M.
      • Jacobs J.F.
      • Kearns G.L.
      • et al.
      Cefotaxime and desacetylcefotaxime pharmacokinetics in infants and children with meningitis.
      Critically ill pediatricsCefotaxime10 samples, IV cannula. 0.7 mL0.17 to 1250 every 6 hCefotaxime130.3610.80.289121.2212.7
      Des-CTX2.10.36321.682.4
      LipoproteinAkins
      • Akins R.L.
      • Haase M.R.
      • Levy E.N.
      Pharmacokinetics of daptomycin in a critically ill adolescent with vancomycin-resistant enterococcal endocarditis.
      Critically ill pediatricsDaptomycin8; unknown volume13Day 1: 6Day 110.0672.310.9183.00.01298.01
      Day 2–6: 8Day 610.0894.580.6196.92.70593.92
      Bradley
      • Bradley J.S.
      • Benziger D.
      • Bokesch P.
      • et al.
      Single-dose Pharmacokinetics of Daptomycin in Pediatric Patients 3-24 Months of Age.
      Complicated skin infection/bacteremiaDaptomycin5; 0.5 mL0.25 to 243–6 mo70.1285.10.02038.7215.0
      47–12 mo70.1355.50.02037.1219.3
      613–24 mo50.1224.40.02267.0281.5
      NitroimidazoleCohen-Wolkowiez
      • Cohen-Wolkowiez M.
      • Sampson M.
      • Bloom B.T.
      • et al.
      Determining Population and Developmental Pharmacokinetics of Metronidazole Using Plasma and Dried Blood Spot Samples From Premature Infants.
      Critically ill neonatesMetronidazole8; 0.2 mL1 to 82 PA15 loading, 7.5 every 12–24 h<14 d PA90.9624.30.0278.0
      ≥14 d PA150.9415.10.04211.1
      Cohen-Wolkowiez
      • Cohen-Wolkowiez M.
      • Ouellet D.
      • Smith P.B.
      • et al.
      Population Pharmacokinetics of Metronidazole Evaluated Using Scavenged Samples from Preterm Infants.
      Critically ill neonatesMetronidazole5; 0.3 mL + scavenged samples22 to 32 GA, 7 to 71 PA4.2–15.4 every 12 h<26 GA130.7120.50.024
      26–29 GA140.7118.60.026
      30–32 GA50.7116.70.029
      Suyagh
      • Suyagh M.
      • Collier P.S.
      • Millership J.S.
      • et al.
      Metronidazole Population Pharmacokinetics in Preterm Neonates Using Dried Blood-Spot Sampling.
      Critically ill neonatesMetronidazole2 to 10; unknown volume24 to 37 GA15 loading, 7.5 every 8–12 hNA320.75619.70.024
      GlycopeptideGous
      • Gous A.G.S.
      • Dance M.D.
      • Lipman J.
      • et al.
      Changes in vancomycin pharmacokinetics in critically ill infants.
      Critically ill pediatricsVancomycin4; days 2 and 82 to 41 GA10 every 6 hDay 2200.815.31.529.112.0
      Day 80.443.41.235.511.7
      Gomez
      • Gomez D.S.
      • Campos E.V.
      • de Azevedo R.P.
      • et al.
      Individualised vancomycin doses for paediatric burn patients to achieve PK/PD targets.
      Pediatrics burnsVancomycin4; + trough, 1-mL sample1 to 1140–60 every 6hNA130.412.42.78
      Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      552.8
      Amaker
      • Amaker R.D.
      • DiPiro J.T.
      • Bhatia J.
      Pharmacokinetics of vancomycin in critically ill infants undergoing extracorporeal membrane oxygenation.
      Critically ill neonatesVancomycin8; venous cannula, 0.3-mL sample volume37 to 42 GA15 or 20 very 8–18 hNA121.0616.90.15
      Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      Ciesd,
      • Cies J.
      • Moore W.
      • Nichols K.
      • et al.
      Population Pharmacokinetics of Vancomycin in neonates on Exra corporeal life support.
      Critically ill neonatesVancomycinNot specifiedNot specifiedNA130.494.91.93
      TriazoleWade
      • Wade K.C.
      • Wu D.
      • Kaufman D.A.
      • et al.
      Population pharmacokinetics of fluconazole in young infants.
      Critically ill neonatesFluconazole12; 0.3 mL23 to 40 GA, 0.14 to 12.6 PA3–12/doseNA551.00.015
      AminoglycosideBressolle
      • Bressolle F.
      • Gouby A.
      • Martinez J.-M.
      • et al.
      Population pharmacokinetics of amikacin in critically ill patients.
      Critically ill pediatricsAmikacin2–6; unknown volume0.5 to 1570–1500 mgNA360.40340.70.97
      Marik
      • Marik P.E.
      • Lipman J.
      • Kobilski S.
      • et al.
      A prospective randomized study comparing once- versus twice-daily amikacin dosing in critically ill adult and paediatric patients.
      Critically ill pediatricsAmikacin10; 0.5–2 mL, 2nd day; peak/trough alternate day arterial line<0.5 to 7020 every 12–24 h<0.5300.585.00.063
      Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      0.5–1300.52.90.068
      Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      15 every 12–24 h1–70400.333.50.051
      Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      Kopcha
      • Kopcha R.G.
      • Fant W.K.
      • Warden G.D.
      Increased dosing requirements for amikacin in burned children.
      Pediatrics burnsAmikacin4; unknown volume0.25 to 1810–15 every 6 hNA380.391.33.5
      Rengelshausen
      • Rengelshausen J.
      • Beedgen B.
      • Tsamouranis K.
      • et al.
      Pharmacokinetics of a netilmicin loading dose on the first postnatal day in preterm neonates with very low gestational age.
      Critically ill neonatesNetilmicin5; unknown volume22.9 to 32 GA5 loading; 1 every 24 h<24 wk50.9418.20.598.102.14141
      5 loading; 1 every 24 h24–27 wk71.0317.00.726.961.78122
      5 loading; 1.5 every 24 h>27 wk80.8817.50.627.952.00142
      Sherwin
      • Sherwin C.M.T.
      • Wead S.
      • Stockmann C.
      • et al.
      Amikacin population pharmacokinetics among paediatric burn patients.
      Pediatrics burnsAmikacin2; unknown volume0.6 to 174.9–22.3NA70Vc 0.24; Vp 0.570.0833.23.8
      Yu
      • Yu T.
      • Stockmann C.
      • Healy D.P.
      • et al.
      Determination of Optimal Amikacin Dosing Regimens for Pediatric Patients With Burn Wound Sepsis.
      Pediatrics burnsAmikacin1–2; unknown volume2 to 1013–20Burns701.14
      Data from abstract only.
      7.22322.3
      2 to 148–16No burns320.82
      Data from abstract only.
      5.3623.90.9
      Wagner
      • Wagner B.P.
      • Pfenninger J.
      Once-daily dosing of netilmicin in neonatal and pediatric intensive-care.
      Critically ill pediatricsNetilmicin2; unknown volume; arterial catheter28 GA to 4.6 PA3.5–6Normal renal666.2–19.50.1–2.5
      Impaired renal130.7–4.4
      CRRT = continuous renal replacement therapy; Des-CTX = desacetylcefotaxime; ECMO = extracorporeal membrane oxygenation; GA = gestational age, weeks; NA = not applicable; PA = postnatal age, days; PK = pharmacokinetic.
      low asterisk Umbilical artery catheters, umbilical venous catheters, peripherally inserted central catheters, antecubital vein venipuncture, or warmed-heel-stick sampling.
      Data from abstract only.
      Calculated as /kg based on mean weight of patient population reported.
      § Calculated from mL/(min/1.73m2) using a mean surface area of 1.40 m2.

      Clearance

      Generally, clearance occurs predominantly via the liver for lipophilic antibiotics (eg, fluoroquinolones) and predominantly via the kidney for hydrophilic antibiotics (eg, β-lactams, aminoglycosides).
      • Roberts J.A.
      • Lipman J.
      Pharmacokinetic issues for antibiotics in the critically ill patient.
      Hepatic and renal function therefore affect this important pharmacokinetic parameter.
      Altered renal function is common in critically ill pediatric patients and presents a challenge for effective antibiotic dosing. Clinically, drug clearance and renal function are often described by estimates of glomerular filtration rate (GFR). One method of estimating GFR is creatinine clearance, which does not describe the additional impacts of tubular secretion and reabsorption that, in sepsis, can be altered as a consequence of disease state and medical interventions. For antibiotics like β-lactams, which commonly undergo a high proportion of tubular secretion, this uncertainty can lead to inaccurate dosing. For pediatric patients, this is further complicated by the degree of maturation of renal function and age-related renal vascular changes.
      • Bartelink I.H.
      • Rademaker C.M.A.
      • Schobben A.
      • et al.
      Guidelines on paediatric dosing on the basis of developmental physiology and pharmacokinetic considerations.
      A study by De Cock et al
      • De Cock P.A.
      • Standing J.F.
      • Barker C.I.
      • et al.
      Augmented renal clearance implies a need for increased amoxicillin-clavulanic acid dosing in critically ill children.
      investigated serum and urinary creatinine and cysteine C as biomarkers of renal clearance to study critically ill children receiving amoxicillin and clavulanic acid and found cysteine C was a significant covariate influencing drug disposition. Furthermore, increased organ blood flow by cardiac output has been identified early in sepsis,
      • Di Giantomasso D.
      • May C.N.
      • Bellomo R.
      Vital organ blood flow during hyperdynamic sepsis.
      and this can affect renal blood flow, as can the use of vasopressors
      • Di Giantomasso D.
      • May C.N.
      • Bellomo R.
      Norepinephrine and vital organ blood flow during experimental hyperdynamic sepsis.
      and, consequently, clearance. Therefore, the description of the clearance of critically ill pediatric patients might require a suitable biomarker that adequately accounts for age-related changes to both renal clearance (that includes GFR, tubular secretion, and reabsorption) and cardiac output. Udy et al
      • Udy A.A.
      • Roberts J.A.
      • Lipman J.
      Clinical implications of antibiotic pharmacokinetic principles in the critically ill.
      recommend the use of urinary creatinine for estimating GFR in critically ill adults; this can be appropriate for pediatric patients also.
      Maturation of renal function begins at 9 weeks of gestation and is completed by 34 weeks of gestation, followed by postnatal changes in renal and intrarenal blood flow.
      • Bartelink I.H.
      • Rademaker C.M.A.
      • Schobben A.
      • et al.
      Guidelines on paediatric dosing on the basis of developmental physiology and pharmacokinetic considerations.
      Developmental changes to renal function are likely to affect the clearance of hydrophilic antibiotics. A study of levofloxacin, that is both hydro- and lipophilic, found children under 2 years of age can have a renal elimination of drugs that is twice as fast as adults.
      • Chien S.C.
      • Wells T.G.
      • Blumer J.L.
      • et al.
      Levofloxacin pharmacokinetics in children.
      Studies of meropenem in neonates
      • Bradley J.S.
      • Sauberan J.B.
      • Ambrose P.G.
      • et al.
      Meropenem pharmacokinetics, pharmacodynamics, and Monte Carlo simulation in the neonate.
      and critically ill infants and children
      • Cies J.
      • Moore W.
      • Chopra A.
      Meropenem Pharmacokinetics in Critically Ill Children.
      using current dosing regimens found increased clearance led to meropenem exposures that might be inadequate to reach some pharmacodynamic targets, such as that required for Pseudomonas aeruginosa.
      Augmented antibiotic clearance has been identified in fit, healthy adult patients with sepsis,
      • Udy A.A.
      • De Waele J.J.
      • Lipman J.
      Augmented renal clearance and therapeutic monitoring of beta-lactams.
      • Udy A.A.
      • Baptista J.P.
      • Lim N.L.
      • et al.
      Augmented renal clearance in the ICU: results of a multicenter observational study of renal function in critically ill patients with normal plasma creatinine concentrations*.
      • Udy A.A.
      • Morton F.J.
      • Nguyen-Pham S.
      • et al.
      A comparison of CKD-EPI estimated glomerular filtration rate and measured creatinine clearance in recently admitted critically ill patients with normal plasma creatinine concentrations.
      • Roberts J.A.
      • Roberts M.S.
      • Semark A.
      • et al.
      Antibiotic dosing in the ׳at risk׳ critically ill patient: Linking pathophysiology with pharmacokinetics/pharmacodynamics in sepsis and trauma patients.
      and can be present in neonate and pediatric sepsis patients. However, establishing an adequate definition of a normal range that accounts for age-related variability and maturation in neonate and pediatric patients can be challenging, but should be determined before significant studies into augmented antibiotic clearance are undertaken.
      A study by De Cock et al
      • De Cock P.
      • Standing J.F.
      • Barker C.I.S.
      • et al.
      Augmented Renal Clearance Implies a Need for Increased Amoxicillin-Clavulanic Acid Dosing in Critically Ill Children.
      found the observed population estimate for amoxicillin clearance in critically ill children is much higher than previously reported in critically ill adults. A standard dose of amoxicillin and clavulanic acid did not meet the expected treatment targets. There was a 32% failure rate in this study using amoxicillin and clavulanic acid for the empirical treatment of sepsis. Other studies of critically ill pediatric patients report clearance results of hydrophilic antibiotics, piperacillin and tazobactam and cefotaxime, that are similar to healthy uninfected patients
      • Cies J.J.
      • Shankar V.
      • Schlichting C.
      • et al.
      Population Pharmacokinetics of Piperacillin/Tazobactam in Critically Ill Young Children.
      or similar to adults.
      • Trang J.M.
      • Jacobs J.F.
      • Kearns G.L.
      • et al.
      Cefotaxime and desacetylcefotaxime pharmacokinetics in infants and children with meningitis.
      Ahsman et al
      • Ahsman M.J.
      • Wildschut E.D.
      • Tibboel D.
      • et al.
      Pharmacokinetics of cefotaxime and desacetylcefotaxime in infants during extracorporeal membrane oxygenation.
      found a lower cefotaxime clearance in critically ill pediatric patients on extracorporeal membrane oxygenation. However, this did not affect the attainment of pharmacokinetic target of time above the minimum inhibitory concentration.
      • Ahsman M.J.
      • Wildschut E.D.
      • Tibboel D.
      • et al.
      Pharmacokinetics of cefotaxime and desacetylcefotaxime in infants during extracorporeal membrane oxygenation.
      Ciprofloxacin is one of the few antibiotics studied in septic pediatric patients that undergoes hepatic metabolism.
      • Halilovic J.
      • Heintz B.H.
      Antibiotic dosing in cirrhosis.
      Lipman et al
      • Lipman J.
      • Scribante J.
      • Gous A.G.S.
      • et al.
      Pharmacokinetic profiles of high-dose intravenous ciprofloxacin in severe sepsis.
      found the clearance in pediatric patients under the age of 12 months was higher than for patients aged between 1 and 5 years. However, the AUC was lower than that seen in adult sepsis patients,
      • Lipman J.
      • Scribante J.
      • Gous A.G.S.
      • et al.
      Pharmacokinetic profiles of high-dose intravenous ciprofloxacin in severe sepsis.
      and the authors concluded that the ciprofloxacin dose might need to be increased in these patients to meet pharmacodynamic targets. The clearance of ciprofloxacin in critically ill neonates was lower in the Leroux et al
      • Leroux S.
      • Turner M.A.
      • Barin-Le Guellec C.
      • et al.
      Pharmacokinetic Studies in Neonates: The Utility of an Opportunistic Sampling Design.
      study than that found by Lipman et al in older critically ill pediatric patients.
      A population pharmacokinetic study by Cohen-Wolkowiez et al
      • Cohen-Wolkowiez M.
      • Sampson M.
      • Bloom B.T.
      • et al.
      Determining Population and Developmental Pharmacokinetics of Metronidazole Using Plasma and Dried Blood Spot Samples From Premature Infants.
      found clearance of metronidazole was significantly associated, and increased disproportionally, with covariates of maturation—a finding consistent with an antibiotic that is predominantly metabolized in the liver. The authors concluded a dosing strategy based on post-menstrual age outperformed current dosing guideline recommendations.
      • Cohen-Wolkowiez M.
      • Sampson M.
      • Bloom B.T.
      • et al.
      Determining Population and Developmental Pharmacokinetics of Metronidazole Using Plasma and Dried Blood Spot Samples From Premature Infants.
      Changes to antibiotic clearance in critically ill neonate and pediatric patients appear to be associated with suboptimal blood concentrations, which can be toxic or inadequate. This uncertainty of drug behavior in neonate and pediatric patients presents significant challenges for the development of optimal dosing regimens.

      Volume of Distribution

      Premature neonates have a high total body water (80%−90% of bodyweight), while fat content is low (10%−15% of bodyweight).
      • Bartelink I.H.
      • Rademaker C.M.A.
      • Schobben A.
      • et al.
      Guidelines on paediatric dosing on the basis of developmental physiology and pharmacokinetic considerations.
      Extracellular water falls from 45% of total body water in the full-term neonate to 20% in the adult.
      • McLeod H.L.
      • Relling M.V.
      • Crom W.R.
      • et al.
      Disposition of antineoplastic agents in the very young-child.
      Furthermore, changes to pediatric patients with a maldistribution of blood flow and fluid shifts (capillary leak syndrome, increased fluid volume) caused by sepsis and medical interventions can alter the Vd of hydrophilic drugs.
      • Roberts J.A.
      • Lipman J.
      Antibacterial dosing in intensive care - Pharmacokinetics, degree of disease and pharmacodynamics of sepsis.
      This increase in Vd has been shown in critically ill adult patients to alter pharmacokinetic parameters of half-life, Cmin, Cmax, and AUC.
      • Triginer C.
      • Izquierdo I.
      • Fernandez R.
      • et al.
      Gentamicin volume of distribution in critically ill septic patients.
      • Roberts J.A.
      • Paul S.K.
      • Akova M.
      • et al.
      DALI: defining antibiotic levels in intensive care unit patients: are current beta-lactam antibiotic doses sufficient for critically ill patients?.
      Therefore, dosing of antibiotics in pediatric patients with sepsis might need to account for both age-related volume changes and altered pathophysiology.
      Lipman et al
      • Lipman J.
      • Gous A.G.S.
      • Mathivha L.R.
      • et al.
      Ciprofloxacin pharmacokinetic profiles in paediatric sepsis: how much ciprofloxacin is enough?.
      reported that there is a difference in Vd for ciprofloxacin between infants with sepsis, with a higher Vd observed in patients younger than 12 months. A study of amikacin pharmacokinetics by Yu et al
      • Yu T.
      • Stockmann C.
      • Healy D.P.
      • et al.
      Determination of Optimal Amikacin Dosing Regimens for Pediatric Patients With Burn Wound Sepsis.
      also found the volume of distribution was increased in patients with burns injuries compared with those without (22.7 L vs 18.7 L; P < 0.01). Increased fluid infusions might explain, at least in part, the increased Vd of amikacin observed among burn patients.
      • Yu T.
      • Stockmann C.
      • Healy D.P.
      • et al.
      Determination of Optimal Amikacin Dosing Regimens for Pediatric Patients With Burn Wound Sepsis.
      Both studies identified that an increase in dose was likely to lead to improved pharmacodynamic target attainment rates, although further clinical evaluations are suggested.
      • Yu T.
      • Stockmann C.
      • Healy D.P.
      • et al.
      Determination of Optimal Amikacin Dosing Regimens for Pediatric Patients With Burn Wound Sepsis.
      A pharmacokinetic study by Bradley et al
      • Bradley J.S.
      • Benziger D.
      • Bokesch P.
      • et al.
      Single-dose Pharmacokinetics of Daptomycin in Pediatric Patients 3-24 Months of Age.
      of daptomycin in pediatric patients found greater renal clearance and volume of distribution compared with adults, resulting in a decreased half-life and reduced exposure. As daptomycin is associated with nerve toxicity, the authors recommended alterations to dosing intervals and time of infusion to address exposure.
      The Vd is also affected by the protein-binding capacity of an antibiotic and availability of binding sites. Plasma concentrations of albumin and total proteins increase from birth to adulthood and, as a consequence of this, neonates and infants 1 to 3 years can be at risk of increased exposure to unbound antibiotic.
      • Sethi P.K.
      • White C.A.
      • Cummings B.S.
      • et al.
      Ontogeny of plasma proteins, albumin and binding of diazepam, cyclosporine, and deltamethrin.
      This can be of significance to antibiotics that are highly protein bound or have a small Vd in adults.
      • Bartelink I.H.
      • Rademaker C.M.A.
      • Schobben A.
      • et al.
      Guidelines on paediatric dosing on the basis of developmental physiology and pharmacokinetic considerations.
      • Kimura T.
      • Sunakawa K.
      • Matsuura N.
      • et al.
      Population pharmacokinetics of arbekacin, vancomycin, and panipenem in Neonates.
      Increased unbound antibiotic concentrations can present as an increased Vd. The clinical relevance of this change, as well as changes to Vd as a consequence of altered pathophysiology and age-related volume changes, requires further study. Poorly understood pharmacokinetic changes lead to uncertainty of how to effectively dose antibiotics in critically ill neonate and pediatric patients. A new approach to pharmacokinetic research is required to address these gaps in knowledge.

      Application of Microsampling to Critically ill Pediatric Patients

      Microsampling from skin prick or from scavenged point-of-care-testing (POCT) samples are the least invasive sampling options for use in a pharmacokinetic study. For most neonate and pediatric patients, these samples can be collected easily. Peripheral samples can be impossible or challenging for patients experiencing septic shock, and this option might not be available. Neonate and pediatric pharmacokinetic studies usually obtain samples ranging from 0.3 to 0.6 mL. Rather than relying on larger volumes of sample, microsamples can still be obtained from scavenged POCT. A study comparing scheduled pharmacokinetic sampling with scavenged sampling and applying the results to population pharmacokinetic modeling found similar predictive performance between each of the models.
      • Leroux S.
      • Turner M.A.
      • Barin-Le Guellec C.
      • et al.
      Pharmacokinetic Studies in Neonates: The Utility of an Opportunistic Sampling Design.
      Implementing microsampling into a pharmacokinetic study requires consideration of whether a wet or dry or plasma or whole blood sample is the most suitable. These decisions are based on multiple factors and are discussed in the section on Considerations for Study Design Using Microsampling.

      Microsampling Techniques and Application to Antibiotics

      Microsampling methods currently used clinically for pediatric patients include dried blood spots (DBS) for neonatal screening of metabolic and inherited diseases, and capillary microsampling is used for small-volume POCT for capillary glucose, capillary ketones, and blood gases.
      • Boyd R.
      • Leigh B.
      • Stuart P.
      Capillary versus venous bedside blood glucose estimations.
      • Khan Z.H.
      • Samadi S.
      • Sadeghi M.
      • et al.
      Prospective study to determine possible correlation between arterial and venous blood gas values.
      • Kuwa K.
      • Nakayama T.
      • Hoshino T.
      • et al.
      Relationships of glucose concentrations in capillary whole blood, venous whole blood and venous plasma.
      • Zavorsky G.S.
      • Cao J.
      • Mayo N.E.
      • et al.
      Arterial versus capillary blood gases: A meta-analysis.
      Newer forms of microsampling hold potential for the quantitation of drugs in biologic samples, and these include dried plasma spots (DPS), volumetric absorptive microsampling (VAMS), solid-phase micro-extraction (SPME), and plasma preparation technologies (PPT). Capillary microsampling has the potential to expand its current use from POCT into the quantitative analysis of drugs.
      DBS are typically prepared by applying a small volume of blood, obtained by thumb or heel prick, to absorbent paper, which is then dried. For a quantitative analysis, either the whole blood spot or a sub-punch is analyzed. Variability in hematocrit has been found to affect the reliability of the resulting concentrations of DBS samples. Variable hematocrit affects the spot size, homogeneity of the sample, and recovery
      • Lawson A.J.
      • Bernstone L.
      • Hall S.K.
      Newborn screening blood spot analysis in the UK: influence of spot size, punch location and haematocrit.
      of the laboratory extraction, and can affect the reliability of the method of detection (termed the matrix effect).
      • Jager N.G.
      • Rosing H.
      • Schellens J.H.
      • et al.
      Determination of tamoxifen and endoxifen in dried blood spots using LC-MS/MS and the effect of coated DBS cards on recovery and matrix effects.
      Therefore, variable hematocrit from individuals can affect the precision and accuracy of the concentration result. Hematocrit varies widely in early infancy, particularly at birth, where normal values can range between 42% and 65%.
      • Jopling J.
      • Henry E.
      • Wiedmeier S.E.
      • et al.
      Reference ranges for hematocrit and blood hemoglobin concentration during the neonatal period: data from a multihospital health care system.
      This variability decreases with 95% ranges of 26.8% to 37.6% at 2 months (n = 119), 29.7% to 38.3% at 5 months (n = 93), and 31.2% to 39.1% at 13 months (n = 42).
      • Hinchliffe R.F.
      • Bellamy G.J.
      • Bell F.
      • et al.
      Reference intervals for red cell variables and platelet counts in infants at 2, 5 and 13 months of age: a cohort study.
      Thereafter, hematocrit rises to adult levels of 36% to 50%.
      • Adeli K.
      • Raizman J.E.
      • Chen Y.
      • et al.
      Complex biological profile of hematologic markers across pediatric, adult, and geriatric ages: Establishment of robust pediatric and adult reference intervals on the basis of the Canadian health measures survey.
      As De Kesel et al
      • De Kesel P.M.
      • Sadones N.
      • Capiau S.
      • et al.
      Hemato-critical issues in quantitative analysis of dried blood spots: challenges and solutions.
      report, there is a clear need to define the hematocrit interval at which the impact of the hematocrit on the accuracy of the analytical result is acceptable. Measures to control for the hematocrit effect on DBS include avoiding the problem (applying the blood volumetrically or using DPS), minimizing the problem (by preparing calibration standard samples in blood with hematocrit close to the range of the patient), or compensating for the problem (by predicting the hematocrit based on an endogenous biomarker, such as potassium.
      • De Kesel P.M.M.
      • Capiau S.
      • Stove V.V.
      • et al.
      Potassium-based algorithm allows correction for the hematocrit bias in quantitative analysis of caffeine and its major metabolite in dried blood spots.
      • Capiau S.
      • Stove V.V.
      • Lambert W.E.
      • et al.
      Prediction of the hematocrit of dried blood spots via potassium measurement on a routine clinical chemistry analyzer.
      DBS has been used as a microsampling tool for the analysis of ceftriaxone
      • Page-Sharp M.
      • Nunn T.
      • Salman S.
      • et al.
      Validation and Application of a Dried Blood Spot Ceftriaxone Assay.
      ; ertapenem
      • la Marca G.
      • Giocaliere E.
      • Villanelli F.
      • et al.
      Development of an UPLC-MS/MS method for the determination of antibiotic ertapenem on dried blood spots.
      ; linezolid
      • la Marca G.
      • Giocaliere E.
      • Villanelli F.
      • et al.
      Development of an UPLC-MS/MS method for the determination of antibiotic ertapenem on dried blood spots.
      • Vu D.H.
      • Bolhuis M.S.
      • Koster R.A.
      • et al.
      Dried Blood Spot Analysis for Therapeutic Drug Monitoring of Linezolid in Patients with Multidrug-Resistant Tuberculosis.
      ; metronidazole
      • Cohen-Wolkowiez M.
      • Sampson M.
      • Bloom B.T.
      • et al.
      Determining Population and Developmental Pharmacokinetics of Metronidazole Using Plasma and Dried Blood Spot Samples From Premature Infants.
      • Suyagh M.
      • Collier P.S.
      • Millership J.S.
      • et al.
      Metronidazole Population Pharmacokinetics in Preterm Neonates Using Dried Blood-Spot Sampling.
      ; moxifloxacin
      • Vu D.H.
      • Koster R.A.
      • Alffenaar J.W.C.
      • et al.
      Determination of moxifloxacin in dried blood spots using LC-MS/MS and the impact of the hematocrit and blood volume.
      ; rifampicin and clarithromycin
      • Vu D.H.
      • Koster R.A.
      • Bolhuis M.S.
      • et al.
      Simultaneous determination of rifampicin, clarithromycin and their metabolites in dried blood spots using LC-MS/MS.
      ; ramoplanin
      • Ewles M.F.
      • Turpin P.E.
      • Goodwin L.
      • et al.
      Validation of a bioanalytical method for the quantification of a therapeutic peptide, ramoplanin, in human dried blood spots using LC-MS/MS.
      ; fluconazole, voriconazole, and posaconazole
      • Reddy T.M.
      • Tama C.I.
      • Hayes R.N.
      A dried blood spots technique based LC-MS/MS method for the analysis of posaconazole in human whole blood samples.
      • van der Elst K.C.M.
      • Span L.F.R.
      • van Hateren K.
      • et al.
      Dried Blood Spot Analysis Suitable for Therapeutic Drug Monitoring of Voriconazole, Fluconazole, and Posaconazole.
      ; netilmicin and gentamicin
      • Tawa R.
      • Matsunaga H.
      • Fujimoto T.
      High-performance liquid chromatographic analysis of aminoglycoside antibiotics.
      ; and piperacillin and tazobactam.
      • Barco S.
      • Risso F.M.
      • Bruschettini M.
      • et al.
      A validated LC-MS/MS method for the quantification of piperacillin/tazobactam on dried blood spot.
      Hofman et al
      • Hofman S.
      • Bolhuis M.S.
      • Koster R.A.
      • et al.
      Role of therapeutic drug monitoring in pulmonary infections: use and potential for expanded use of dried blood spot samples.
      identified DBS as a potentially feasible tool for dosing individualization, using therapeutic drug monitoring, for a wide range of antibiotics in the treatment of pulmonary infections. Antibiotic prescribing for critically ill patients recommends the use of therapeutic drug monitoring to ensure appropriate pharmacokinetic targets are met in a patient group that can experience dramatic and intra-individual fluctuations in physiology.
      • Denny K.J.
      • Cotta M.O.
      • Parker S.L.
      • et al.
      The use and risks of antibiotics in critically ill patients.
      Capillary microsamples are prepared by collecting whole blood in a plastic or glass capillary tube (that may contain anticoagulant).
      • Jonsson O.
      • Villar R.P.
      • Nilsson L.B.
      • et al.
      Validation of a bioanalytical method using capillary microsampling of 8 µl plasma samples: application to a toxicokinetic study in mice.
      Plasma is then obtained by centrifuging the capillary tube. Capillary microsampling has been used as a blood sampling tool for the quantitative analysis of moxifloxacin.
      • Bowen C.L.
      • Licea-Perez H.
      • Karlinsey M.Z.
      • et al.
      A novel approach to capillary plasma microsampling for quantitative bioanalysis.
      DPS are prepared in a manner similar to DBS, with the application of plasma rather than whole blood. DPS eliminates some of the problems associated with variable hematocrit being applied to the absorbent paper. DPS can be prepared using capillary microsampling. DPS has been used as a microsampling tool for the quantitative analysis of linezolid,
      • Baietto L.
      • D׳Avolio A.
      • Ariaudo A.
      • et al.
      Development and validation of a new UPLC-PDA method to quantify linezolid in plasma and in dried plasma spots.
      fosfomycin,
      • Parker S.L.
      • Roberts J.A.
      • Lipman J.
      • et al.
      Quantitative bioanalytical validation of fosfomycin in human whole blood with volumetric absorptive microsampling.
      daptomycin,
      • Baietto L.
      • D׳Avolio A.
      • Pace S.
      • et al.
      Development and validation of an UPLC-PDA method to quantify daptomycin in human plasma and in dried plasma spots.
      trimethoprim, and sulfamethoxazole.
      • Gonzalez D.
      • Melloni C.
      • Poindexter B.B.
      • et al.
      Simultaneous determination of trimethoprim and sulfamethoxazole in dried plasma and urine spots.
      Recent advances in microsampling include VAMS devices. The VAMS device is an absorbent polymeric tip that wicks up a fixed volume (10 μL) of whole blood by capillary action.
      • Spooner N.
      • Denniff P.
      • Michielsen L.
      • et al.
      A device for dried blood microsampling in quantitative bioanalysis: overcoming the issues associated blood hematocrit.
      The tip is then dried in preparation for transport and storage. VAMS has been validated as a microsampling tool for the quantitative analysis of fosfomycin.
      • Parker S.L.
      • Roberts J.A.
      • Lipman J.
      • et al.
      Quantitative bioanalytical validation of fosfomycin in human whole blood with volumetric absorptive microsampling.
      PPT are single-step technologies in which whole blood is applied to a spot on a card. The sample passes through a membrane removing cells and allowing plasma to be collected on a disc as a standard volume.
      • Sturm R.
      • Henion J.
      • Abbott R.
      • et al.
      Novel membrane devices and their potential utility in blood sample collection prior to analysis of dried plasma spots.
      The plasma thus becomes a DPS. PPT has not been applied to the analysis of antibiotics.
      Finally, a form of microsampling that holds potential for future development is SPME. SPME devices use stainless-steel fibers on which the drug is absorbed.
      • Prosen H.
      • Zupančič-Kralj L.
      Solid-phase microextraction.
      • Schubert J.K.
      • Miekisch W.
      • Fuchs P.
      • et al.
      Determination of antibiotic drug concentrations in circulating human blood by means of solid phase micro-extraction.
      The drug is desorbed from the fiber for analysis. SPME has been used in proof-of-concept studies for the analysis of linezolid.
      • Schubert J.K.
      • Miekisch W.
      • Fuchs P.
      • et al.
      Determination of antibiotic drug concentrations in circulating human blood by means of solid phase micro-extraction.
      • Olszowy P.
      • Szultka M.
      • Nowaczyk J.
      • et al.
      A new way of solid-phase microextraction fibers preparation for selected antibiotic drug determination by HPLC-MS.
      This technology has not been reported for sampling direct from humans.

      Considerations for Study Design Using Microsampling

      Decisions for using microsampling in a pharmacokinetic study can be based on a “contingency approach,” as described by Parker et al.
      • Parker S.L.
      • Dorofaeff T.
      • Lipman J.
      • et al.
      Is there a role for microsampling in antibiotic pharmacokinetic studies?.
      This approach considers the impact of the properties of the antibiotic, as well as the type and geographical location of the patients.

      Properties of the Antibiotic

      Total versus Unbound Measurement of Antibiotics

      Once an antibiotic is absorbed, its distribution is influenced by its binding capacity, organ perfusion, organ size, and the permeability of tissues.
      • Rowland M.
      • Tozer T.N.
      The portion of the antibiotic that is unbound is responsible for the pharmacologic activity (drug that is not bound to either plasma proteins or red blood cells). Often, microsampling only allows for measuring total concentrations of drug.
      For antibiotics that do not exhibit high levels of binding, for example, fosfomycin,
      • Zeitlinger M.A.
      • Sauermann R.
      • Traunmuller F.
      • et al.
      Impact of plasma protein binding on antimicrobial activity using time-killing curves.
      pharmacokinetic studies are usually performed based on the total concentration of the antibiotic. If an antibiotic is bound to plasma proteins or red blood cells, but the binding is fixed (does not vary with concentration or between patients), for example, cefazolin,
      • Wong G.
      • Briscoe S.
      • Adnan S.
      • et al.
      Protein Binding of β-Lactam Antibiotics in Critically Ill Patients: Can We Successfully Predict Unbound Concentrations?.
      total concentrations can be used and a factor applied to the pharmacokinetic calculations to represent the bound proportion. When antibiotic binding alters across a concentration range that results from the therapeutic dose, or it varies between patients, for example, ceftriaxone
      • Schleibinger M.
      • Steinbach C.L.
      • Topper C.
      • et al.
      Protein binding characteristics and pharmacokinetics of ceftriaxone in intensive care unit patients.
      or linezolid,
      • D.E. W.
      • A. S.
      • J.L. K.
      • et al.
      Determination of Tissue Penetration and Pharmacokinetics of Linezolid in Patients with Diabetic Foot Infections Using In Vivo Microdialysis.
      both total and unbound concentrations are measured.

      Stability

      The stability of an antibiotic in its biologic matrix (ie, plasma, whole blood, urine) and in its format as a microsample, requires assessment. This is an essential element of a quantitative bioanalytical validation (see section on Validation Requirements). Before commencing a pharmacokinetic study, the stability of the microsample is assessed for storage and sample handling conditions. Some antibiotics have been found to have limited stability in whole blood and when dried and stored as microsamples at room temperature, including piperacillin
      • Barco S.
      • Risso F.M.
      • Bruschettini M.
      • et al.
      A validated LC-MS/MS method for the quantification of piperacillin/tazobactam on dried blood spot.
      and fosfomycin.
      • Parker S.L.
      • Lipman J.
      • Roberts J.A.
      • et al.
      Quantitative bioanalytical validation of fosfomycin in human whole blood with volumetric absorptive microsampling (VAMS).

      Distribution in the Body

      The concentration of drugs can exhibit a marked blood sampling site dependence,
      • Chiou W.L.
      The phenomenon and rationale of marked dependence of drug concentration on blood-sampling site - implications in pharmacokinetics, pharmacodynamics, toxicology and therapeutics.
      this can be between arterial and venous sampling, or between central-line sampling and peripheral (toe or finger) sampling. A bridging study that correlates a peripheral microsample with arteriovenous samples can provide sufficient data on the suitability of the sampling site.

      Type of Patient: Critically Ill Neonate and Pediatric Patients

      Critically ill neonate and pediatric patients can commonly experience anemia and hemodilution from medical interventions.
      • Vincent J.L.
      • Baron J.F.
      • Reinhart K.
      • et al.
      Anemia and blood transfusion in critically ill patients.
      As discussed in the section on Microsampling Techniques and Application to Antibiotics, hematocrit can affect the viscosity of a whole blood sample. Some whole blood microsamples, for example, DBS, from these patients can produce unreliable antibiotic concentrations if they are calculated relative to calibration standards prepared using blood with a different level of hematocrit.
      Similarly, neonate and pediatric patients can experience changes in albumin concentrations. Lower albumin concentrations can be caused by hemodilution or hypoalbuminemia due to liver or kidney failure, and higher albumin concentration from dehydration. Changes to albumin concentrations can affect the viscosity of a microsample for both whole blood and plasma.
      Alterations to hemostasis have been identified in adult patients with a clinical diagnosis of sepsis.
      • Stief T.W.
      • Ijagha O.
      • Weiste B.
      • et al.
      Analysis of hemostasis alterations in sepsis.
      Hemostasis can provide challenges to sampling pediatric patients. Hypercoagulability can prevent effective blood absorption by the microsample substrate, such as DBS, PPT, or VAMS. The capillary action required to collect a capillary microsample might be limited. Scavenged POCT samples might still be available for collection of plasma.
      Pediatric patients are frequently administered anticoagulants, such as heparin, to prevent tissue hypoxygenation and to attenuate organ damage and dysfunction.
      • Cornet A.D.
      • Smit E.G.M.
      • Beishuizen A.
      • et al.
      The role of heparin and allied compounds in the treatment of sepsis.
      Lithium heparin has been found to cause a suppression or enhancement of signal in LC-MS/MS.
      • Mei H.
      • Hsieh Y.S.
      • Nardo C.
      • et al.
      Investigation of matrix effects in bioanalytical high-performance liquid chromatography/tandem mass spectrometric assays: application to drug discovery.
      This can produce a falsely high or low concentration result for a patient sample.

      Patient Location

      Many neonate and pediatric patients with sepsis will be treated in intensive care units or emergency departments in hospital. Pediatric patients in a metropolitan location might receive oral antibiotics prescribed by a general practitioner before transfer to hospital. In rural or remote locations, intravenous antibiotics can be administered, with the patient then transferred to hospital. Patients in impoverished locations might be administered intravenous antibiotics in hospitals with limited facilities.
      Where patients are in remote or rural locations, the requirement for samples to be easily transported without the need for complex sample manipulation is paramount. Dry microsamples (such as DBS, PPT, and VAMS) are well suited to this and would allow shipping between collaborating sites.
      Collection of a sample in close proximity to a laboratory for sample preparation or analysis, such as a metropolitan hospital with an onsite research facility, offers the researcher a greater number of choices in sample preparation, with both wet and dry samples suitable for analysis.
      Microsampling has already been used to analyze the efficacy of antimalarial therapy in children in developing countries.
      • Ashley E.A.
      • Stepniewska K.
      • Lindegardh N.
      • et al.
      Comparison of plasma, venous and capillary blood levels of piperaquine in patients with uncomplicated falciparum malaria.
      • Ursing J.
      • Eksborg S.
      • Rombo L.
      • et al.
      Chloroquine is grossly under dosed in young children with malaria: implications for drug resistance.
      Interestingly, the results of the study by Ursing et al
      • Ursing J.
      • Eksborg S.
      • Rombo L.
      • et al.
      Chloroquine is grossly under dosed in young children with malaria: implications for drug resistance.
      found that dosing of chloroquine was inadequate.

      Validation Requirements

      The application of microsampling to a pharmacokinetic study requires the conduct of comprehensive quantitative bioanalytical validation.
      European Medicines Agency
      Guideline on bioanlaytical method validation.
      Food and Drug Administration
      Guidance for Industry Bioanalytical Method Validation.
      Most of the testing is performed before the collection of clinical samples. This prescriptive testing process ensures the resulting concentrations of the antibiotic tested reflects the original sample.
      A quantitative bioanalytical validation includes testing for accuracy, precision, linearity, recovery, matrix effects, stability, and an incurred sample reanalysis. The US Food and Drug Administration’s Draft Guidance for Industry Bioanalytical Method Validation provides direction on the validation of DBS, including stating that for DBS, correlative studies (or “bridging studies”) with traditional sampling should be performed.
      Food and Drug Administration
      Draft Guidance for Industry Bioanalytical Method Validation.

      Conclusions

      Infection in critically ill neonate and pediatric patients is a major health problem. Current antibiotic regimens for critically ill neonate and pediatric patients are frequently suboptimal due to a poor understanding of altered pharmacokinetic properties. An assessment of the suitability of microsampling for pharmacokinetic studies in neonate and pediatric patients is recommended before wider use. The method of sampling, as well as the method of bioanalysis, also requires validation to ensure the data obtained reflects the true result.

      Conflicts of Interest

      The authors have indicated that they have no conflicts of interest regarding the content of this article.

      Acknowledgments

      J.A.R is a recipient of an Australian National Health and Medical Research Council Fellowship (APP1048652). We wish to acknowledge funding from the Australian National Health and Medical Research Council for Project Grants (APP1044941, APP1062040) and Centre of Research Excellence (APP1099452).
      Dr. Dorofaeff: First author. Compiled review and table, drafting of manuscript and final approval of manuscript. Ms. Bandini: Second author. Contributed to drafting of manuscript and final approval of manuscript. Prof. Lipman: Third author. Contributed to the conception of the manuscript and final approval of manuscript. Dr. Ballot: Fourth author. Contributed to the conception of the manuscript and final approval of manuscript. Prof. Roberts: Fifth author. Contributed to the conception of the manuscript, revising it critically for important intellectual content and final approval of manuscript. Dr. Parker: Senior author. Contributed to the conception of the manuscript, revising it critically for important intellectual content and final approval of manuscript.

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