Distribution of quetiapine between serum and whole blood in therapeutic drug-monitoring specimens

Abstract

Quetiapine use is on the rise, leading to a corresponding increase in acute intoxications, some of which have fatal outcomes. When assessing whole-blood quetiapine concentrations during forensic autopsies, interpretations are primarily based on toxicity data from studies of serum concentrations. To our knowledge, there are only two previous studies that have attempted to establish the ratio between whole blood and serum quetiapine concentrations with limited populations and high variability of results. Paired specimens of whole blood and serum from 16 quetiapine users recruited from the Psychiatric Clinic, St. Olav University Hospital were analyzed using LC–MS-MS. Quetiapine concentrations in both matrices were determined and compared. The mean blood:serum ratio of quetiapine was 0.74 (standard deviation (SD) = 0.05, 95% confidence interval (CI) 0.71–0.76, P < 0.001), range 0.66–0.85. Simple linear regression showed strong linear correlation between quetiapine concentrations in the two matrices (B = 0.774, P > 0.001, r = 0.999). Our results imply that quetiapine occurs at lower concentrations within erythrocytes than in plasma. This is most likely due to a high degree of plasma protein binding. Other factors which may influence the distribution of quetiapine between these compartments are solubility, metabolism and passive or active efflux mechanisms. We did not observe any covariation between blood:serum ratios and serum concentrations. Quetiapine was consistently present at lower concentrations in whole blood than in serum. If so inclined to, a conversion factor of ∼0.7 may be considered for extrapolation of concentrations from serum to whole blood, at least in cases with therapeutic quetiapine concentration levels.

Introduction

Quetiapine is the most commonly used antipsychotic drug in Norway having undergone a sharp increase in use over the past 20 years compared to other atypical antipsychotics. Similar trends have also been reported from several other countries (1–6). Throughout this period, quetiapine ingestion—accidental and intentional—has become the second most common cause of contact with the Norwegian Poisons Information Centre (4.6%), exceeded only by paracetamol intake (14.6%) (7), and quetiapine is now a common finding in toxicological screens performed at our Department, both in specimens from living and deceased subjects.

In therapeutic drug monitoring and diagnostics of acute poisonings, serum or plasma is usually sampled. Reference values for therapeutic and toxic concentrations are thus mainly based on plasma/serum measurements. In forensic casework, however, and in particular autopsy cases, whole blood is sampled (8). Drug distribution can exhibit significant variations between plasma/serum and whole blood, with these discrepancies often being unpredictable and specific to each substance (9). Reference values for both whole blood and plasma/serum are therefore desirable. For many substances, such combined reference values are scarce or lacking (10). In these cases, knowledge of the distribution between plasma/serum and whole blood can be useful.

To our knowledge, only two studies have previously examined the correlation between quetiapine concentrations in whole blood and serum/plasma. Fisher et al. (11) compared 17 paired plasma and whole-blood specimens from therapeutic drug monitoring and found a mean blood:plasma ratio of 0.67 (range 0.36–0.83), whereas Saito et al. (12) compared 9 paired serum and whole blood specimens from suspected intoxications, and reported a blood:serum ratio range of 0.5–1.1 (no mean provided) and increasing ratio with increasing quetiapine concentrations.

In this study, we investigated the ratio of quetiapine concentrations in serum and whole-blood specimens collected simultaneously in a therapeutic drug-monitoring setting.

Materials and methods

Sample collection

Participants were recruited from the Psychiatry Clinic, St. Olav University Hospital including both inpatients and outpatients. The study included participants who met the following inclusion criteria:

1. Informed consent from patients with capacity to consent.

2. Therapeutic, physician-prescribed use of quetiapine, regardless of indication or dosage.

3. Referral to therapeutic drug monitoring of quetiapine, regardless of indication or dosage.

Paired specimens of serum and ethylenediaminetetraacetic acid (EDTA)-anticoagulated whole blood were collected from 16 different patients aged 18–57 years (mean 34), 4 males and 12 females. After sampling, specimens intended for serum production were stored at room temperature for 30–60 min followed by centrifugation. Specimens were stored at 4–8°C for a maximum of 66 h prior to transportation to the Department of Clinical Pharmacology via St. Olav University Hospital’s central sample reception. Following arrival at the Department of Clinical Pharmacology, specimens were stored at a temperature of 4–8°C for a maximum of 24 h prior to analysis.

Ethics

The study has been approved by the Regional Committee for Medical and Health Research Ethics (approval number 2019/561), and data acquisition has been permitted by the Data Protection Officer of St. Olav University Hospital.

Specimen collection constituted no additional medical intervention on the participants. Sample identification numbers were deleted upon data extraction for processing, so that participants were completely anonymized in the study.

Analysis

Specimens were processed and analyzed using high-performance liquid chromatography—tandem mass spectrometry with liquid–liquid extraction. All sample pairs were analyzed on the same day and work-up and analysis were performed using the same instruments and reagent batches for each pair.

Specimens were spiked with a 400 µg/L quetiapine-d₈-fumarate internal standard prepared in an aqueous solution of methanol, followed by protein precipitation using −20°C acetonitrile, then centrifugated. The resulting supernatants were transferred to a collection well plate and passed through a positive pressure processor. Filtered extracts were injected on an UPLC–MS-MS instrument and analyzed. Technical specifications for sample work-up and UPLC–MS-MS settings are described in detail in previous publications (13, 14). The methods have been rigorously validated for determination of quetiapine concentrations in whole blood and serum and are routinely utilized by the nationally accredited laboratory at the Department of Clinical Pharmacology, St. Olav University Hospital in forensic autopsy cases and therapeutic drug monitoring, respectively.

Data processing

Statistical significance of means and differences of means were estimated using one-sample and independent-sample t-tests with two-sided P-values. Normality was tested visually with a Q–Q plot and numerically with the Shapiro–Wilkes test for normality. Linear regression was performed with a simple linear regression model. Degree of linear correlation was estimated using Pearson’s R. Statistical analyses and graphs were produced in IBM SPSS Statistics v 29.0.0.0.

Results

Quetiapine concentrations in serum and whole blood and the concentration ratio between the two matrices are presented in Table I. Specimen pairs are numbered chronologically in the order they were received and analyzed.

Table I.

Whole blood and serum concentration levels of quetiapine

Specimen pair (sex, age in years)	Serum concentration	Whole-blood concentration (µg/L)	Blood:serum ratio
1 (F, 24)	83	58	0.70
2 (F, 41)	543	410	0.76
3 (M, 28)	150	105	0.71
4 (M, 50)	14	10	0.73
5 (F, 42)	61	42	0.69
6 (F, 25)	5.0	4.2	0.85
7 (F, 18)	28	20	0.74
8 (F, 23)	10	6.9	0.69
9 (F, 28)	9.2	7.3	0.79
10 (F, 38)	5.4	3.8	0.71
11 (F, 30)	8.4	6.1	0.73
12 (M, 42)	14	11	0.81
13 (F, 40)	5.4	4.2	0.79
14 (F, 21)	15	10	0.69
15 (F, 57)	56	38	0.68
16 (M, 31)	161	118	0.74

Serum and whole-blood concentrations and blood:serum concentration ratio of quetiapine in paired therapeutic drug-monitoring specimens.

The mean blood:serum ratio of quetiapine was 0.74 (standard deviatiation (SD) = 0.05, 95% Confidence interval (CI) 0.71–0.76, P < 0.001), range 0.68–0.85. Ratios were shown to be normally distributed using a Q–Q plot (data not shown) and the Shapiro–Wilkes test for normality (P = 0.085).

Linear regression showed strong linear correlation between quetiapine concentrations in the two matrices (B = 0.753, P > 0.001, r = 0.999). The correlation between serum and whole blood concentration levels of quetiapine is displayed graphically as a cluster plot with regression line in Figure 1.

Figure 1.

Cluster plot of serum and whole-blood concentrations of quetiapine in paired whole blood and serum specimens with regression line.

The difference of mean blood: serum ratios between male and female participants was 0.01 (95% CI −0.75–0.05, P = 0.68). The correlation between participant age and blood:serum concentrations was r = 0.07 (P = 0.81).

Discussion

Quetiapine was consistently present at lower concentrations in whole blood compared to serum, indicating that the concentration of quetiapine contained within or bound to the membranes of circulating hematocytes—of which erythrocytes make up the entire volume—is lower than in the remaining blood fractions. The gender and age of participants showed no significant covariance with quetiapine blood:serum concentration ratios. Twelve participants (75%) were female and only two participants were aged ≥50 years at the time of specimen collection.

The ability of a drug to cross cell membranes is dependent on its volume of distribution, and thus its lipophilicity. Quetiapine is poorly water-soluble and should diffuse easily into erythrocytes (15). Also, as a lipophilic substance, one would anticipate higher concentrations of quetiapine in whole blood, given the higher lipid content of this matrix (16). However, it is important to note that quetiapine has a high affinity for binding to plasma proteins, which may contribute to its distribution not aligning solely with solubility factors. Another possible explanation is that quetiapine is effluxed from or metabolized within the erythrocyte itself, thus reducing the relative concentration in that compartment, but, to our knowledge, this has not been reported as an elimination route for quetiapine.

In the previous study by Saito et al. (12), quetiapine concentrations in whole blood and serum specimens from nine acute overdose cases were investigated. This study showed larger variability in the blood:serum ratio (0.5–1.1) than our material, and also increasing ratio with increasing quetiapine concentrations, intersecting a ratio of 1 at a serum concentration of ∼2500 µg/L. Increasing relative concentrations of quetiapine in the cellular fraction of the blood could suggest saturation of potential binding proteins in serum which yields an elevated free fraction of quetiapine. As stated by the authors, this could indicate that whole-blood concentrations better reflect the unbound fraction of quetiapine, and thus toxicity, at higher concentration levels.

Our results display very high degree of linear correlation and consistent ratios between the two matrices. This could be attributed to the fact that our study encompasses patients with therapeutic serum concentrations of quetiapine (mean 98 µg/L, range 5–543 µg/L) at steady state, whereas Saito et al. included overdose cases with serum quetiapine concentrations ∼2500 µg/L in which all patients were transported to the hospital within 3 h of suffering an overdose (12). In the Saito et al. study, blood:serum ratios for cases with therapeutic concentrations of quetiapine were estimated to be close to 0.6, corresponding quite closely to our results. Therapeutic serum concentrations of quetiapine are commonly asserted to lie between 50 and 500 µg/L (17–19)—although the recommended dosing range is sometimes greatly exceeded for individual patients (20)—and apply to specimens collected ∼12 h after last intake of the drug when using immediate release dosage formulations, and ∼24 h after last intake of extended release formulations. When samples are collected shortly after ingestion, absorption from the gastrointestinal tract is still occurring and there may thus be a larger fraction of quetiapine which has not yet been bound to circulating proteins. Hypotension and associated renal impairment may affect both unbound and bound fractions of quetiapine, whereas myoglobinemia due to seizures and increased synthesis of acute phase proteins as seen in severe overdoses could provide additional binding sites—these phenomena have all been reported in severe quetiapine intoxications (21, 22). Furthermore, acute quetiapine intoxication frequently occurs concomitantly with other drug intoxications, and drug–drug interactions could influence the apparent concentration levels and distribution of quetiapine. In other words, specimens collected in the setting of an acute overdose may display blood:serum ratios distinct from those seen in physiologically stable patients with well-distributed and declining quetiapine concentrations. These effects could occur independently of the proposed binding protein saturation.

In the study by Fisher et al., a mean blood:plasma ratio of 0.67 (range 0.35–0.83) was reported. This study also investigated therapeutic drug-monitoring specimen pairs and the mean ratio was similar to that found in our material, although the range was considerably wider. Fisher et al. used EDTA plasma as opposed to serum in our study and the study by Saito et al. However, there is tradition for using the terms “serum” and “plasma” interchangeably and haphazardly, particularly in clinical practice (23). The primary mechanism of EDTA anticoagulant in blood work is irreversible chelation of metal ions, which inhibits the induction of coagulation by calcium ions (24). For some drug molecules, the presence of EDTA may interfere with the stability or protein binding in the sample by means of pH alteration or chelation of compounds involved in drug protein binding, respectively. Such interference has not been reported for quetiapine and is considered unlikely. This is corroborated by the similar results of the EDTA plasma study of Fisher et al. and the present study, in which no added anticoagulant was present in the analyzed specimens.

Regardless of the mechanism(s) causing the uneven distribution of quetiapine between whole blood and serum, the results imply that hematocrit influences the blood:serum distribution of quetiapine, and that, an assumption of a 1:1 correlation between whole blood and serum concentrations is fallacious. Preferably, more toxicity data on quetiapine should be generated in whole blood for use in forensic toxicology casework. If using serum reference values for this purpose, one may consider a conversion factor of ∼0.7. However, the findings of Saito et al. indicate that this factor should not be extrapolated to cases with serum quetiapine concentrations higher than those reported in the present study, as the relationship between blood and serum concentrations may exhibit nonlinearity in acute intoxications with high concentration levels.

A key limitation to the present study is the absence of quetiapine concentrations that greatly exceed therapeutic concentrations. Also, the sample size was limited to 16 paired specimens, so the validity of extrapolating results from the present study can be questioned as much as for previous studies. Moreover, measurements of hematological parameters such as hematocrit and serum protein concentrations were not available. The inclusion of such parameters may have allowed a better interpretation and understanding of the underlying mechanisms. In particular, interindividual differences in the blood:serum distribution could possibly be illuminated. However, such differences were not pronounced in our material.

It would be of interest to use the presented results to assume the relationship between quetiapine concentrations in perimortem serum and postmortem whole blood. However, one must keep in mind that this study utilized sample pairs collected simultaneously in vivo. Physiological, microbiological and chemical postmortem phenomena may influence the quetiapine distribution in blood to an extent that renders such assumptions highly insecure.

Future perspectives

Future research into the distribution of quetiapine in whole blood and serum should attempt to bridge the gap between the apparently linear ratio in the present study and the shift towards nonlinearity at higher concentrations observed by Saito et al. Such studies should also attempt to account for paraclinical parameters such as hematocrit and plasma protein, which may influence this distribution. Populations with prescribed quetiapine doses higher than the recommended range should be studied to identify whether ratio nonlinearity is an inherent function of high quetiapine concentrations or due other physiological or preanalytical factors which are specific to acute intoxications. Regions which utilize plasma rather than serum samples would benefit from having similar studies conducted comparing plasma and whole blood.

Conclusion

In this material of therapeutic drug-monitoring specimens, quetiapine was consistently present at lower concentrations in whole blood than in serum. If so inclined to, a conversion factor of ∼0.7 may be considered for extrapolation of concentrations from serum to whole blood, at least in cases with therapeutic quetiapine concentration levels.

Data availability

The data underlying this article will be shared upon reasonable request to the corresponding author.

Funding

The project was funded entirely by the Faculty of Medicine and Health Sciences at the Norwegian University of Science and Technology, Trondheim, Norway.

Conflict of interest

None of the authors have declared any conflicts of interest of relevance to the contents of this article, financial or otherwise.