(S)-7-[[2-O-6-deoxy-α-L-mannopyranosyl)-β-D-glucopyranosyl]oxy]-2,3-dihydro-5-hydroxy-2-(3-hydroxy-4-methoxyphenyl)-4H-1-benzopyran-4-one

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Objective of study:

absorption

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Neohesperidin was determined by UHPLC-MS/MS in rat plasma after a single administration of Poncirus trifoliata extract to 6 rats.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

(S)-7-[[2-O-6-deoxy-α-L-mannopyranosyl)-β-D-glucopyranosyl]oxy]-2,3-dihydro-5-hydroxy-2-(3-hydroxy-4-methoxyphenyl)-4H-1-benzopyran-4-one

EC Number:

236-216-9

EC Name:

(S)-7-[[2-O-6-deoxy-α-L-mannopyranosyl)-β-D-glucopyranosyl]oxy]-2,3-dihydro-5-hydroxy-2-(3-hydroxy-4-methoxyphenyl)-4H-1-benzopyran-4-one

Cas Number:

13241-33-3

Molecular formula:

C28H34O15

IUPAC Name:

(S)-7-[[2-O-6-deoxy-α-L-mannopyranosyl)-β-D-glucopyranosyl]oxy]-2,3-dihydro-5-hydroxy-2-(3-hydroxy-4-methoxyphenyl)-4H-1-benzopyran-4-one

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: The powdered P. trifoliata fruits (100 g) were extracted by refluxing with water (1 : 15, w/v) for 2 h.2 The aqueous extract was filtered, concentrated under reduced pressure by using a rotary evaporation apparatus, and lyophilized to give a powder (15.6 g). The powder was suspended in 0.5% CMC–Na solution to get a concentration equivalent to 0.156 g/ml of P. trifoliata extract.
- Purity: Neohesperidin, NE: 0.604 mg/mL.
- Hesperidin, HE: 1.61 mg/mL
- Hesperidin Methyl chalcone, HMC: 0.378 mg/mL.
- Poncirin, PO: 2.06 mg/mL.
- Phellopterin, PH: 0.196 mg/mL.

Radiolabelling:

no

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Laboratory Animal Center of China Medical University (Shenyang, China).
- Weight at study initiation: 230 ± 20 g

ENVIRONMENTAL CONDITIONS
- All animals were kept in conformity with the European Community guidelines for the use of experimental animals (86/609/EEC).

Administration / exposure

Route of administration:

oral: unspecified

Vehicle:

other: 0.5% CMC–Na solution

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: lyophylized p. trifoliata extract was suspended in 0.5% CMC–Na solution to get a concentration equivalent to 0.156 g/mL of extract (0.604 mg/mL neohesperidin).

Duration and frequency of treatment / exposure:

single dose.

Doses / concentrations

Dose / conc.:

6.04 mg/kg bw/day (nominal)

Remarks:

Neohesperidin in plant extract. Dose of P. trifoliata extract of 1.56 g/kg (equivalent to 16.1, 6.04, 3.78, 20.6, and 1.96 mg/kg of HE, NE, HMC, PO, and PH, respectively).

No. of animals per sex per dose / concentration:

6 males

Control animals:

no

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: blood, plasma.
- Time and frequency of sampling: Serial blood samples (approximately 0.3 mL) were collected through the oculi chorioideae vein in heparinized tubes at 0, 5, 15, and 30 min, and then at 1, 1.5, 2, 3, 5, 8, 12, and 24 h.
- Other: Plasma was separated by centrifugation (2500 x g, 10 min), and stored at -20 ºC until analysis.

ANALYTICAL METHOD:
- Separation method: UHPLC. UHPLC system (Dionex UltiMate 3000) with a cooling autosampler and column oven and a TSQ Quantum Ultra mass spectrometer (Thermo Scientific, San Jose, USA) with an electrospray ionization (ESI) interface.
- Conditions: Agilent XDB C18 column (100 x 2.1 mm and 1.8 mm). The mobile phase consisted of water containing 0.1% formic acid (solvent A) and acetonitrile (solvent B). The gradient elution program
was set as follows: 20 / 45% B (v/v) at 0–5.0 min; 45 / 90% B at 5.0–5.5 min, 90% B at 5.5–6.5 min; 90 / 20% B at 6.5–7.0 min; 20% B at 7.0–10.0 min. The flow rate was set at 0.35 mL/min and injection volume was 2 mL using a partial loop mode for sample injection. The temperatures of the column and autosampler were maintained at 30 and 10ºC, respectively.
- Detection method: MS/MS. Mass spectrometric detection was performed using an ESI source via polarity switching between positive and negative (for NE) ionization mode; m/z 609.2 / 301.0 was used as quantifier for NE.
- Limits of detection and quantification: LOQ = 2.0 ng/ml for NE.
- Extraction recovery (indicate if results are corrected or not for recoveries): 88.4 - 93.9%. Results not corrected.
- Validation: Method validation was performed according to the guidelines set by the United States Food and Drug Administration (FDA) for bioanalytical method validation. Linear range for NE was 2.0 - 250 ng/ml, r = 0.9962, accuracy 99.5%, RSD = 2.5%.

Results and discussion

Main ADME results

Type:

absorption

Results:

quick absorption, detection in plasma 5 min after oral administration.

Toxicokinetic / pharmacokinetic studies

Details on absorption:

These compounds were quickly absorbed into the body and were detected 5 min after oral administration.

Toxicokinetic parameters

Toxicokinetic parameters 1

Key result

Test no.:

#1

Toxicokinetic parameters:

Tmax: 1.18 ± 0.57 h

Toxicokinetic parameters 2

Key result

Test no.:

#1

Toxicokinetic parameters:

Cmax: 19.1 ± 3.6 ng/mL

Toxicokinetic parameters 3

Key result

Test no.:

#1

Toxicokinetic parameters:

half-life 1st: 4.17 ± 1.04 h

Toxicokinetic parameters 4

Key result

Test no.:

#1

Toxicokinetic parameters:

AUC: 79.2 μg/(L·h)

Remarks:

to the last measurable concentration

Toxicokinetic parameters 5

Key result

Test no.:

#1

Toxicokinetic parameters:

AUC: 94.8 μg/(L·h)

Remarks:

to infinity

Metabolite characterisation studies

Metabolites identified:

not measured

Applicant's summary and conclusion

Conclusions:

The substance was quickly absorbed and found in plasma 5 minutes after oral administration, reaching a maximum plasma concentration of 19.1 ± 3.6 ng/mL, the half-life of elimination (T1/2) was 4.17 ± 1.04 h.

Executive summary:

A pharmacokinetic study was performed after a single oral administration of Poncirus trifoliata extract to 6 male Sprague-Dawley rats, containing 0.604 mg/mL of neohesperidin, which resulted in a dose of 6.04 mg/kg bw of neohesperidin. Neohesperidin content was determined in rat plasma by means of a validated UHPLC-MS/MS method. The substance was quickly absorbed and found in plasma 5 minutes after oral administration. The relevant toxicokinetic parameters obtained were the following: The time of maximum plasma concentration (Tmax) and maximum plasma concentration (Cmax) were 1.18 ± 0.57 h and 19.1 ± 3.6 ng/mL, the half-life of elimination (T1/2) was 4.17 ± 1.04 h, and the area under the curve to the last measurable concentration (AUC0–t) and to infinity (AUC) were 79.2 and 94.8 mg/Lh.