Median Effective Dose of Esketamine Combined with Remifentanil for Anesthesia Induction in Painless Colonoscopy

Introduction

Esketamine is the dextrorotatory enantiomer of ketamine, which is a racemic mixture consisting of equal parts of the dextrorotatory (S-ketamine) and levorotatory (R-ketamine) forms.¹ Compared to ketamine, esketamine demonstrates a greater potency in binding to N-methyl-D-aspartate (NMDA) receptors and other targets, resulting in more effective sedation and analgesia. These enhanced effects are associated with a significantly shorter recovery time compared to ketamine.^2–4 Consequently, esketamine is better suited to clinical settings where rapid postoperative recovery, shorter hospital stays for outpatient procedures, and efficient medical management are essential.

Ketamine is the only intravenous general anesthetic with strong analgesic properties, primarily providing sedation and analgesia. However, its analgesic effects are often supplemented with other general anesthetics, such as propofol, in both research and clinical practice.⁵ This raises the question of whether combining ketamine with other potent analgesics could fully harness its analgesic potential. The combination of esketamine and remifentanil is supported by complementary pharmacodynamic mechanisms. Esketamine exerts potent sedative and analgesic effects via non‑competitive antagonism at NMDA receptors, inhibiting central nociceptive transmission. Remifentanil, a selective μ‑opioid receptor agonist, provides rapid, dose‑dependent analgesia without significant sedation. Their distinct molecular targets and synergistic analgesic action allow for effective pain suppression with potentially reduced individual drug doses, minimizing adverse effects while maintaining hemodynamic stability. This mechanistic synergy provides a pharmacodynamic basis for their combined use in procedural sedation and analgesia.And a preliminary study conducted by our research team explored this idea, applying a combination of esketamine and remifentanil for anesthesia induction during painless gastroscopy. The results showed the feasibility, safety, and efficacy of this regimen.⁶

Building on this preliminary research, this investigation aimed to determine both the median effective dose (ED₅₀) and the 95% effective dose (ED₉₅) of esketamine combined with remifentanil for anesthesia induction during painless colonoscopy. Given the significant differences in stimulation intensity, pain intensity, and pain characteristics between gastroscopy and colonoscopy, this study further analyzed the ED₅₀ and ED₉₅ specific to anesthesia induction during painless colonoscopy. The objective was to offer additional insights and references for clinical practice.

Data and Methods

General Data

This study adopted a sequential design.This study was approved by the ethics committee of Northern Jiangsu People’s Hospital (Approval number: 2023ky062) and registered in the Chinese Clinical Trial Registry (Registration number: ChiCTR2300069712). Patients undergoing painless colonoscopy at the Endoscopy Center of Subei People’s Hospital of Jiangsu province in April 2023 were screened for this study.

Inclusion criteria were as follows: Both male and female participants; aged between 18 to 60 years; body mass index (BMI) of 18 to 30 kg/m²; American Society of Anesthesiologists (ASA) physical status I or II; patients undergoing colonoscopy alone, with normal cognitive function and no severe communication barriers; patients who were well informed about the study and volunteered to participate.

Exclusion criteria were as follows: Patients undergoing gastroscopy, digestive tract polyp resection, foreign body removal from the digestive tract, or other digestive tract diagnostic or treatment procedures via an endoscope in addition to colonoscopy; Patients with a history of alcohol or drug abuse; Patients with a history of long-term oral use of benzodiazepines or similar medications for insomnia or other conditions (duration > 1 month);

Criteria for removing patients from the study: Patients experiencing temporary changes to the colonoscopy procedure, such as the resection of multiple colonic polyps; Patients requesting to withdraw from the study midway.

Study Methods

Preoperative Preparation, Anesthesia, and Colonoscopy

Before the procedure, patients attended preoperative consultations at the Department of Gastroenterology or other relevant departments and received a colonoscopy order. At the anesthesia clinic, patients underwent pre-anesthesia evaluations, including risk assessments and necessary preoperative examinations. Screening and informed consent discussions were conducted and patients were assigned to the corresponding groups after signing informed consent forms.

In accordance with standard preoperative preparation for colonoscopy, patients ingested polyethylene glycol electrolyte powder for bowel cleansing and adhered to routine preoperative fasting protocols. Upon arrival at the endoscopy center as scheduled, participants completed routine preparations, including changing into appropriate attire, establishing intravenous access, positioning, and continuous cardiac monitoring. Nasal oxygen was administered at a flow rate of 2 to 4 L/min. All colonoscopy procedures were performed by the same highly experienced endoscopist following standard operating procedures.

Management of intraoperative adverse events: If systolic blood pressure is less than 90 mmHg or the blood pressure drop exceeds 20% of the baseline value, norepinephrine 8 μg or phenylephrine 100 μg should be slowly injected intravenously. If the heart rate is less than 50 beats per minute, atropine 0.5 mg should be injected intravenously. If the blood oxygen saturation is less than 95%, the patient’s head should be appropriately hyperextended to open the airway, or the angle of the mandible should be appropriately stimulated to stimulate breathing.

Protocol for Anesthesia Induction and Sequential Administration

Anesthesia induction was performed in accordance with the established protocol, and colonoscopy began after the patient’s eyelash reflex had subsided.

(1) Based on the previous research foundation of our research group and related studies, the initial dose for anesthesia induction was determined as: Esketamine (Batch number: 221018BL, Jiangsu Hengrui Pharmaceuticals Co., Ltd.) was administered at a dose of 0.1 mg/kg,^7,8 combined with remifentanil (Batch number: 20A02151, Yichang Humanwell Pharmaceutical Co., Ltd.) at a dose of 0.3 μg/kg.⁹

(2) Sequential administration: According to the anesthesia induction protocol, the remifentanil dosage was fixed, while the esketamine dosage was adjusted using a sequential dosing method. Specifically, consecutive esketamine doses followed a 1:1.2 ratio, with each dose incrementally increased relative to the preceding one. The esketamine doses administered were 0.100 mg/kg, 0.120 mg/kg, 0.144 mg/kg, 0.173 mg/kg, 0.207 mg/kg, 0.249 mg/kg, and 0.299 mg/kg.

For each participant, if a positive response was observed at a given dose, the subsequent participant received the adjacent lower dose. Conversely, if a negative response was noted, the next participant received the adjacent higher dose. The participant immediately preceding the first instance of a positive response was designated as participant number 1.

The sample size for the study was determined by identifying at least seven inflection points, defined as transitions from a negative to a positive response between consecutive doses. Each inflection point involved two participants: one exhibiting a negative response followed by another with a positive response. Data from cases containing at least seven such consecutive negative-to-positive inflection points were included in the final analysis.

Criteria for Positive Response and Remedial Measures for Negative Response

A positive response was defined by the simultaneous fulfillment of the following six criteria. Failure to meet any one of the six criteria classified the response as negative:

(1) Modified observer’s assessment of alertness/sedation (MOAA/S) score: An MOAA/S score of ≤ 1 within one minute following anesthesia induction was required. The MOAA/S scale grades are as follows:

Grade 5: Full alertness with normal response to verbal stimuli.

Grade 4: Slow response to normal verbal stimuli.

Grade 3: Response only to loud and/or repeated verbal stimuli.

Grade 2: Response only to mild prodding or gentle shaking.

Grade 1: Response only to painful stimuli (squeezing the trapezius muscle).

Grade 0: No response to painful stimuli (squeezing the trapezius muscle).

(2) Time to reach adequate depth of anesthesia ≤ 2 minutes, defined as the duration from the completion of anesthesia induction dosing to the MOAA/S score ≤ 1.

(3) Body movement grade: A body movement grade ≤ 1 was necessary during the passage of the colonoscope’s working end through the sigmoid colon. The grading scale for body movement is as follows:¹⁰

Grade 1: Patients may experience mild involuntary limb movements that do not interfere with the endoscopy procedure and do not require the administration of additional anesthetics.

Grade 2: Patients may grip the endoscope, disrupting the endoscopist’s procedure, necessitating the administration of additional anesthetics to manage the situation.

Grade 3: Patients may display agitation and an inability to cooperate, requiring the examination to be halted and anesthesia deepened before proceeding.

(4) Esketamine dosing frequency: Fewer than five esketamine doses were administered within any 15-minute interval before the working end of the colonoscope passed through the sigmoid colon.

(5) Adverse event severity: No adverse events of Grade III or higher occurred before the working end of the colonoscope fully passed the sigmoid colon. Adverse events grades were defined as follows:¹¹

Grade I: Mild adverse events without clinical symptoms or only mild symptoms with no need for treatment.

Grade II: Moderate adverse events: Requiring minimal, local, or non-invasive treatment and potentially limiting daily activities, with the use of age-appropriate assistive devices (eg., cooking, shopping, making phone calls).

Grade III: Severe adverse events: The presence of serious diseases or medically significant symptoms that are not immediately life-threatening but result in hospitalization or prolonged hospitalization, or cause non-bedridden disability with limitations in activities of independent daily living (eg., bathing, dressing, undressing, eating, using the toilet, taking medication).

Grade IV: Life-threatening conditions requiring urgent treatment.

Grade V: Fatality due to adverse events.

For study participants with a negative response, anesthesiologists independently chose appropriate anesthetic agents, such as propofol or etomidate, based on the individual’s condition to administer supplemental anesthesia and ensure that the participants underwent colonoscopy at an adequate depth of anesthesia.

Observation Indicators

The observation indicators included the incidence of positive responses, the total number of cases with positive responses, the number of negative-to-positive inflection points observed, the esketamine dosages administered during anesthesia induction, and detailed documentation of any adverse events.

Statistical Analysis

Data were analyzed using SPSS 20.0.Data conforming to a normal distribution were expressed as mean ± standard deviation (), while non-normally distributed data were presented as median (interquartile range,IQR). In the correlation analysis, the chi-square test was used to assess the association between gender and the positive/negative groups, with the effect size expressed by Cramer’s V value. Age and BMI were non-normally distributed data, and the Mann–Whitney U-test was adopted to analyze their association with the grouping, with the effect size represented by the correlation coefficient r value. For the logistic regression analysis, a binary logistic regression model was constructed with the positive/negative grouping as the binary dependent variable (negative = 0, positive = 1) and gender (male = 1, female = 0), age, and BMI as independent variables, so as to analyze the predictive effect of demographic characteristics on the grouping. The variance inflation factor (VIF) was used to test for multicollinearity, the area under the receiver operating characteristic curve (AUC) was applied to evaluate the predictive efficacy of the model, and McFadden R² was used to assess the goodness of fit of the model. All tests were two-tailed, with a test level of α=0.05. The ED₅₀, ED₉₅, and corresponding 95% CI of esketamine combined with remifentanil for painless colonoscopy anesthesia induction were determined using Probit regression analysis. And the sample size was verified through the power analysis of the association between demographic characteristics and positive/negative response grouping.

Results

General Data of the Study Participants

A total of 23 study participants were included in this study, with the following distribution: 13 males and 10 females. The participants had an age range of 44.727 ± 6.971 years, a height range of 168.173 ± 5.990 cm, and a body weight range of 65.478 ± 7.248 kg. The BMI was 23.125 ± 1.974 (Table 1). In terms of the ASA physical status, 13 participants were classified as Grade I, and 10 were classified as Grade II.

Table 1 General Information of the Subjects

Sequential Administration

The first positive response was observed in the 4th study participant following the initial dosing. The participant immediately preceding this 4th participant was designated as participant number 1. By the conclusion of the sequential trial, a total of 23 participants had been enrolled, yielding 9 negative-to-positive inflection points in total. The number of inflection points exceeded the minimum statistical requirement of 7 inflection points for the sequential dosing method, indicating that the results were reliable. (Figure 1).

Figure 1 A sequential diagram showing the use of esketamine in combination with remifentanil for anesthesia induction during a painless colonoscopy.

Power Analysis of Sample Size

The power analysis of the association between demographic characteristics and anesthetic response grouping showed the following results:For the association between age and positive/negative grouping: the effect size was r=0.720, the actual power was 1-β=0.89 (≥0.8), and the minimum required sample size was 18 cases; For the association between BMI and positive/negative grouping: the effect size was r=0.614, the actual power was 1-β=0.83 (≥0.8), and the minimum required sample size was 20 cases;For the binary logistic regression with gender, age, and BMI as independent variables: the model’s AUC=0.765, the actual power was 1-β=0.86 (≥0.8), the minimum required sample size was 21 cases, and there was no multicollinearity (VIF < 1.1);For the association between gender and positive/negative grouping: the effect size was Cramer’s V=0.000, indicating no association. (Table 2).

Table 2 Correlation Analysis of Demographic Characteristics with Positive/Negative Groups

In conclusion, the 23 cases meet the sample size requirements of this study.

Anesthetic Efficacy and Adverse Events

A total of 11 study participants exhibited a positive response, with a duration of (60.750 ± 4.288) seconds to reach the target depth of anesthesia. One participant experienced body movement during the passage of the colonoscope through the sigmoid colon, which was scored as Grade 1, prompting an additional dose of esketamine. The anesthesia induction dose of esketamine for the 11 participants was (0.190 ± 0.0219) mg/kg. Two Grade I adverse events, specifically tachycardia ([Heart rate] HR > 100 beats/min), were observed in 2 cases before the working end of the colonoscope fully passed through the sigmoid colon, the incidence rate of adverse events was 18.18%.

A total of 12 study participants exhibited a negative response, with an esketamine induction dose of (0.231 ± 0.0271) mg/kg. Among these 12 participants, 11 did not reach the target depth of anesthesia, as indicated by an MOAA/S score > 1 after the administration of anesthesia induction. Of these 11 participants, 8 scored 4, 1 scored 3, and 2 scored 2 on the MOAA/S scale. In the case of the remaining participant, although the target depth of anesthesia with a MOAA/S score ≤ 1 was achieved, the participant experienced body movement of Grade 2 or above during the passage of the colonoscope through the sigmoid colon. As a result, propofol was intravenously administered as a supplemental anesthesia, allowing the procedure to proceed.

Correlation and Logistic Regression Analyses of Demographic Characteristics with Positive/Negative Groups

A total of 23 patients were enrolled in this study, including 12 cases (52.2%) in the negative group and 11 cases (47.8%) in the positive group.

Results of correlation analysis:The chi-square test was used to analyze the association between gender and the positive/negative groups, and the results showed no statistically significant difference in gender distribution between the two groups (χ²=0.000, P=1.000) with Cramer’s V=0.000, indicating no significant association between gender and grouping. The Mann–Whitney U-test was adopted to evaluate the association between age and grouping, revealing no statistically significant difference between the two groups (U=95.000, P=0.078) with an effect size of r=0.720, which suggested no significant association between age and grouping. For the association between BMI and grouping, no statistically significant difference was found between the two groups (U=81.000, P=0.372) with an effect size of r=0.614, demonstrating no significant association between BMI and grouping (Table 2).

Results of Logistic regression analysis:Results of the full-variable model showed that the model had a moderate predictive ability (AUC=0.765) with a general goodness of fit (McFadden R²=0.186). The AUC of 5-fold cross-validation was 0.667±0.264, indicating a moderate stability of the model. Univariate logistic regression analysis revealed that gender had no significant predictive effect on grouping (OR=1.167, 95% CI: 0.224–6.081, P=0.855, AUC=0.519); age had no significant predictive effect on grouping (OR=0.348, 95% CI: 0.110–1.099, P=0.072, AUC=0.720); BMI had no significant predictive effect on grouping (OR=0.536, 95% CI: 0.215–1.341, P=0.183, AUC=0.614). Multicollinearity test showed that the VIF values of all variables were < 1.1, indicating no multicollinearity issue and the model results were reliable (Table 3).

Table 3 Results of Logistic Regression Analysis of Demographic Characteristics with Positive/Negative Groups

In conclusion, there was no statistically significant association between the demographic characteristics of the patients in this study and the positive and negative groups.

The ED50 and ED95 of Esketamine Combined with Remifentanil for Painless Colonoscopy Anesthesia Induction

The ED₅₀ and ED₉₅ of esketamine combined with remifentanil for painless colonoscopy anesthesia induction were found to be 0.209 mg/kg (95% CI: 0.178–0.248 mg/kg) and 0.259 mg/kg (95% CI: 0.231–0.478 mg/kg), respectively.

Discussion

Ketamine is unique among intravenous general anesthetics due to its potent analgesic properties, providing both sedation and analgesia. These dual effects make ketamine a viable option for primary general anesthesia and an essential choice in opioid-free anesthesia protocols. However, its sympathetic-stimulation effects present a notable limitation, particularly in adult anesthesia. While ketamine can depress myocardial function, it also leads to significant increases in blood pressure and heart rate, placing an added strain on the circulatory system. This limitation restricts its standalone use in adults. The conventional approach to adult general anesthesia involves combining ketamine with other anesthetic agents for intravenous general anesthesia. This combination harnesses ketamine’s analgesic effects while mitigating its sympathetic stimulation agents that suppress the circulatory system. Propofol is commonly used as the sedative in the “ketamine + sedative” combinations due to its strong sedation and circulatory suppression, which counteract ketamine’s excitatory cardiovascular effects.^5,12,13 Additionally, agents such as etomidate, remimazolam, or others have also been reported in various “ketamine + sedative” regimens to broaden options and adapt to specific clinical needs.^14–17

Esketamine is twice as potent as ketamine in terms of anesthetic, analgesic, and hypnotic effects, requiring only half the dose of ketamine to achieve the same anesthetic outcome. Furthermore, esketamine has higher bioavailability, a shorter elimination half-life, and enables more precise control over anesthesia depth. Consequently, esketamine facilitates a faster recovery with an improved quality of awakening.^2–4

Esketamine serves as an improved drug option for clinicians and a promising tool for innovative research on the clinical application of ketamine-based drugs. The concept of “awake analgesia” offers valuable insight in this regard. The aim of awake analgesia is to achieve a state of sedation where patients remain conscious but experience a significant reduction or complete elimination of pain and other distressful sensations. This state is achieved by combining potent analgesics with minimal sedative use, summarized by the principle “strong analgesia + relatively weak sedation.” Such an approach enables effective pain management while allowing for a reduction in general anesthesia dosages, thereby minimizing related adverse reactions.

The “ketamine + sedative,” combination, particularly when paired with remifentanil, optimizes ketamine’s analgesic properties. Esketamine, with its advantages of a shorter elimination half-life, more controllable anesthesia, faster awakening, and improved recovery quality, is especially effective in this combination. Additionally, remifentanil, known for its independence from hepatic and renal metabolism, constant elimination half-life, and lack of drug accumulation, complements esketamine by enhancing its efficacy. This synergy not only fulfills the goal of “strong analgesia + relatively weak sedation” but also reduces drug dosage, minimizes circulatory fluctuations caused by esketamine, and promotes faster awakening and superior recovery quality.

In summary, the combination of esketamine and remifentanil represents a highly promising approach for the innovative use of ketamine-based drugs, offering significant clinical benefits.

Guided by the concept of “strong analgesia + relatively weak sedation,” our research team previously investigated the feasibility, efficacy, and safety of combining esketamine with remifentanil.⁶ At the same time, the ED₅₀ and ED₉₅ of esketamine combined with remifentanil for painless gastroscopy anesthesia induction were examined. Considering the significant differences between colonoscopy and gastroscopy in terms of procedure methods, accessed digestive tract, stimulation level, pain intensity and nature, and procedure duration, ED₅₀ and ED₉₅ of this combination for painless colonoscopy anesthesia induction were further examined in this study, along with an analysis of its safety and efficacy, building upon previous research findings.

The findings from this study confirmed that the combination of esketamine and remifentanil for anesthesia induction in painless colonoscopy successfully achieved the predetermined depth of anesthesia based on the positive response criteria. Furthermore, this combination resulted in only two Grade I adverse events (tachycardia), underscoring its safety and efficacy for use in colonoscopy anesthesia induction.

This study employed more rigorous criteria for assessing anesthetic efficacy than those typically applied in clinical practice. These criteria include target anesthetic depth, duration required to reach the target anesthetic depth, and an innovative criterion of body movement during the passage of the colonoscope through the sigmoid colon, which was used to define successful anesthesia induction in the context of colonoscopy.

Target Depth of Anesthesia and Duration to Reach Target Depth

In mainland China, the target depth of anesthesia for painless digestive endoscopy typically involves deep sedation and anesthesia.^18,19 In 2018 alone, more than 5 million painless digestive endoscopies were performed in mainland China.²⁰ As a result, there is a significant demand for effective anesthesia duration, rapid awakening, and high turnover. Given these conditions, this study defined the target depth of anesthesia as deep sedation and anesthesia, specifically aiming for a MOAA/S score ≤ 1, measured 1 minute after completing anesthesia induction. The duration required to achieve the target depth of anesthesia was set at ≤ 2 minutes.

Using Patient Body Movement During the Passage of the Colonoscope Through the Sigmoid Colon as a Criterion for Successful Colonoscopy Anesthesia Induction

Anesthesia plays a crucial role in ensuring patient cooperation by promoting a quiet and steady state during procedures, which is essential for medical staff, especially endoscopists. Anesthesia induction is deemed ineffective if patient movement interferes with the endoscopic procedure. Successful anesthesia induction not only indicates adequate anesthesia depth but also ensures that the patient remains stable, without significant movements that could disrupt the procedure, even under the initial stimulus of the endoscope. The strongest stimulus at the start of a gastroscopy procedure occurs when the endoscope passes through the pharynx. Therefore, successful passage through the pharynx signifies successful gastroscope entry. No equivalent standard exists for colonoscopy. The stimulus when the colonoscope passes through the anus and rectum is minimal, and even conscious patients generally do not experience significant discomfort. Consequently, our team proposed an innovative and more stringent criterion for successful anesthesia induction: the colonoscope’s passage through the sigmoid colon without significant body movement, categorized as ≤ Grade 1 movement.

The results of this study showed that the ED₅₀ and ED₉₅ of esketamine combined with remifentanil for painless colonoscopy anesthesia induction were 0.209 mg/kg and 0.259 mg/kg, respectively. Two cases of Grade I adverse events occurred before the working end of the colonoscope passed through the sigmoid colon, both involving tachycardia (HR > 100 beats/min). In our previous study on painless gastroscopy anesthesia induction, the ED₅₀ and ED₉₅ of esketamine combined with remifentanil for were 0.215 mg/kg and 0.254 mg/kg, respectively.⁶ Additionally, two cases of Grade I adverse events occurred, including one case of tachycardia (HR > 100 beats/min) and one case of significant postoperative fatigue. These results yielded similar ED₅₀ and ED₉₅ values for esketamine combined with remifentanil between painless gastroscopy and colonoscopy anesthesia induction. Furthermore, both studies confirmed the safety and feasibility of this anesthesia induction regimen. However, both studies also showed tachycardia (HR > 100 beats/min) as an adverse event, highlighting that circulatory function fluctuations caused by esketamine remain a critical issue. On the other hand,we also observed similarities in the esketamine dosage in this study with those in other relevant research. For example, in the study by Liu et al,²¹ the dosage of esketamine for painless colonoscopy was 02 mg/kg combined with 1 mg/kg of propofol, while in the study by Liu et al²² on anesthesia for endoscopic ultrasound (EUS), the dosage was 0.25 mg/kg of esketamine combined with 1.5–2 mg/kg of propofol. The esketamine dosages of 0.2~0.25 mg/kg (either as monotherapy or in combination) in relevant studies are highly consistent with that in our study, suggesting that 0.2~0.25 mg/kg is an optimal dosage range of esketamine for sedation and anesthesia in endoscopic procedures. This dosage can achieve effective sedation across different drug combination regimens and various endoscopic procedures, demonstrating good clinical universality. This consistency in dosage also confirms the stable sedative efficacy of esketamine within this range, which can serve as a core reference for the clinical selection of esketamine dosage in endoscopic examinations.

This Study Has Several Limitations

(1) All procedures were performed by a highly experienced endoscopist, whose skills may have been significantly more advanced than those of a mid-level endoscopist. As a result, patients under the care of the highly experienced endoscopist experienced significantly less traction and stimulation from the colonoscope. This could have led to an underestimation of the esketamine dosage, potentially affecting the calculated ED₅₀ and ED₉₅ values. This factor may partly explain the similar ED₅₀ and ED₉₅ outcomes for esketamine combined with remifentanil in both painless gastroscopy and colonoscopy. Future research should involve diverse patient populations, endoscopists with varying levels of experience, and hospitals of different grades to enable a more precise analysis.

(2) The ED₅₀ and ED₉₅ values of anesthesia induction drugs in this study were determined using the sequential method, with a small sample size. Larger, well-powered randomized controlled trials are needed to fully evaluate the efficacy and postoperative recovery associated with this new drug combination.

(3) The inherent limitations of the sequential method may have led to imprecision in the determination of ED₅₀ and ED₉₅ values.

In summary, the ED₅₀ and ED₉₅ for esketamine combined with remifentanil for inducing anesthesia in painless colonoscopy were 0.209 mg/kg (95% CI 0.178–0.248 mg/kg) and 0.259 mg/kg (95% CI 0.231–0.478 mg/kg), respectively.