Fatty acids, tall-oil, manganese salts

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vitro / ex vivo

Remarks:

Bioelution in simulated interstitial medium

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 December 2021 to 12 February 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

no guideline available

Principles of method if other than guideline:

Bioelution in simulated interstitial medium according to Standard Operating Procedure for the Bio-accessibility Testing Programme of Eurometaux (November 10, 2010).

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

The test item, Fatty acids, tall-oil manganese salts as received, was tested. Therefore, no further preparation of the test item at ECTX was needed.

Vehicle:

other: simulated interstitial fluid

Duration and frequency of treatment / exposure:

168 hours

Positive control reference chemical:

1001 mg/L Manganese standard

Bioaccessibility (or Bioavailability) testing results:

Bio-elution refers to the in vitro extraction methods used to measure the degree to which a substance is dissolved in simulated biological fluids. The resulting value is the “bio-accessibility” and is defined as the “fraction of a substance that is soluble under physiological conditions and therefore potentially available for absorption into systemic circulation”. Simulated biological fluids (e.g. gastric fluid, interstitial fluid, lysosomal fluid,…) represents relevant exposure routes. In this study, the inhalation route was investigated. Therefore, simulated interstitial fluid, as provided in the test guidance, was used to measure the release/dissolution of manganese from Fatty acids, tall-oil manganese salts.

The temperature of the sampled test solutions fluctuated between 36.8 °C and 37.0 °C, which was in line with the test conditions of 37 °C ± 1 °C.

The measured pH of the test medium at the start of the test was 7.43, i.e. within the specifications of pH 7.4 ± 0.2. At the start of the trial, the pH measured in the additional test item vessel (replica X) was 7.45, i.e. within the specifications of pH 7.4 ± 0.2. At the sampling points, the pH in the blank control vessel and positive control were between 7.41 and 7.44. The pH in the test vessels was between 7.43 and 7.58 at the 24 hours and 168 hours sampling points.

Measurements of the blank control vessels showed no manganese concentrations above the reporting limit of 20 μg/L Mn in the high salt loaded matrix.

The following observations could be made in the Positive Control item vessel containing a soluble form of manganese:
- The Positive Control item solution remained clear at the tested concentration level. No precipitation could be observed on the filter and in the Erlenmeyer flask at the end of the experiment.
- The recovery of the dissolved manganese concentrations in the positive control item was 100 % for Mn at the 24 hours sampling point and 97 % for Mn at the 168 hours endpoint, respectively and met the criterion (recovery: 100 % ± 10 %).

The following observations could be made in the test vessels with a loading of 2 g/L Fatty acids, tall-oil manganese salts:
- Undissolved test item (dark pieces) could be observed in the Erlenmeyer flasks at the end of the experiment.
- Following amounts of manganese were measured:
1. 24 hours sampling point:
An average dissolved manganese concentration in solution of 0.291 ± 0.047 mg/L Mn (corresponding to a dissolved manganese concentration per g test item of 0.145 ± 0.023 mg/g Mn) was measured after 24 hours of exposure to the simulated interstitial fluid (pH 7.4) with a between-vessel variation of 16 %.
Based on the manganese content of the test item (i.e. 8.9 % Mn) and the average dissolved manganese concentration, a manganese release of 0.16 % could be calculated at the 24 hours sampling point.
2. 168 hours endpoint:
An average dissolved manganese concentration in solution of 2.74 ± 0.52 mg/L Mn (corresponding to a dissolved manganese concentration per g test item of 1.36 ± 0.26 mg/g Mn) was measured after 168 hours of exposure to the simulated interstitial fluid (pH 7.4) with a between-vessel variation of 19 %, which met the <20 % quality criterion at the 168 hours endpoint.
Based on the manganese content of the test item (i.e. 8.9 % Mn) and the average dissolved manganese concentration, a manganese release of 1.5 % could be calculated at the 168 hours endpoint.

Conclusions:

During this study on Fatty acids, tall-oil manganese salts at a loading of 2 g/L in simulated interstitial fluid (pH 7.4), it was shown that for manganese, an average dissolved concentration in solution of 0.291 mg/L Mn (CVbetween-vessel = 16 %; N = 3) was found after 24 hours of extraction. This amount corresponds to a dissolved manganese concentration per g test item of 0.145 mg/g Mn or a manganese release of 0.16 % of the content.
At the 168 hours endpoint, an average value of 2.74 mg/L Mn (CVbetween-vessel = 19 %; N = 3) corresponding to a dissolved manganese concentration per g test item of 1.36 mg/g Mn was found or a manganese release of 1.5 % of the content.
The results are reliable since the test conditions stayed constant during the experiment and the recovery of the Positive Control met the criteria of acceptance for manganese.

Executive summary:

Bio-elution refers to the in vitro extraction methods used to measure the degree to which a substance (e.g., metal or mineral ion) is dissolved in artificial biological fluids. This study measured the release/ dissolution of manganese ions from Fatty acids, tall-oil manganese salts in a simulated interstitial fluid was measured. Simulated biological fluids represent relevant exposure routes. The resulting value is the “bio-accessibility” and is defined as the “fraction of a substance that is soluble under physiological conditions and therefore potentially available for absorption into systemic circulation”. 

The objective of this study was to obtain knowledge about the bio-elution characteristics of Fatty acids, tall-oil manganese salts in simulated interstitial fluid. This study has been conducted following the recommended Standard Operating Procedure (SOP) for the Bio-accessibility Testing Programme of Eurometaux (November 10, 2010). 

The extent of dissolution of Fatty acids, tall-oil manganese salts was tested in a simulated interstitial fluid at 37 °C and pH 7.4 during 168 hours (2 g/L loadings) at an agitation speed of 171 revolutions per minute (rpm). The bio-elution endpoint is based on the dissolved manganese (Mn) concentrations obtained after 24 and 168 hours of extraction. 

The measured pH of the test medium at the start of the test was 7.43, i.e. within the specifications of pH 7.4 ± 0.2. At the start of the trial, the pH measured in the additional test item vessel (replica X) was 7.45, i.e. within the specifications of pH 7.4 ± 0.2. At the sampling points, the pH in the blank control vessel and positive control were between 7.41 and 7.44. The pH in the test vessels was between 7.43 and 7.58 at the 24 hours and 168 hours sampling points. 

The temperature of the sampled test solutions, including the blank test vessel, was between 36.8 °C and 37.0 °C and corresponded to the required test conditions of 37 °C ± 1 °C. 

Measurements of the blank control vessels showed no manganese concentrations above the reporting limit of 20 μg/L Mn in the high salt loaded matrix. 

In the test item vessels with a loading of 2 g/L Fatty acids, tall-oil manganese salts at the 24 hours sampling point, the following average dissolved manganese concentration in solution of: 

• 0.291 ± 0.047 mg/L Mn (CVbetween-vessel = 16 %; N = 3), corresponding to a dissolved manganese concentration per g test item of 0.145 ± 0.023 mg/g Mn, was measured after 24 hours of exposure to the simulated interstitial fluid (pH 7.4). 

Based on the manganese content (i.e. 8.9 % Mn) in the test item and the average dissolved manganese concentrations in the test solutions, a release of: 

• manganese of 0.16 % of the content, could be calculated at the 24 hours sampling point. 

In the test item vessels with a loading of 2 g/L Fatty acids, tall-oil manganese salts at the 168 hours endpoint, the following average dissolved manganese concentration in solution of: 

• 2.74 ± 0.52 mg/L Mn (CVbetween-vessel = 19 %; N = 3), corresponding to a dissolved manganese concentration per g test item of 1.36 ± 0.26 mg/g Mn, was measured after 168 hours of exposure to the simulated interstitial fluid (pH 7.4). 

Based on the manganese content (i.e. 8.9 % Mn) in the test item and the average dissolved manganese concentrations in the test solutions, a release of: 

• manganese of 1.5 % of the content, could be calculated at the 168 hours endpoint. 

Reference 2

Endpoint:

basic toxicokinetics in vitro / ex vivo

Remarks:

Bioelution in simulated gastric medium

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 December 2021 to 12 February 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

no guideline available

Principles of method if other than guideline:

Bioelution in simulated gastric medium according to Eurometaux (2019): The Standard Operating Procedure for Bioelution Testing of Metals, Inorganic Metal Compounds, and Complex Metal-Containing Materials: Simulated Gastric Fluid (Eurometaux Brussels, November 02, 2019).

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

The test item, Fatty acids, tall-oil manganese salts as received, was tested. Therefore, no further preparation of the test item at ECTX was needed.

Vehicle:

other: simulated gastric fluid (0.032 N HCl)

Duration and frequency of treatment / exposure:

1 hour agitation and 1 hour to allow the sample to settle

Positive control reference chemical:

1001 mg/L Manganese standard

Bioaccessibility (or Bioavailability) testing results:

Bioelution refers to the in vitro extraction methods used to measure the degree to which a substance is dissolved in simulated biological fluids. The resulting value is the “bioaccessibility”, and is defined as the “fraction of a substance that is soluble under physiological conditions and therefore potentially available for absorption into systemic circulation”. Simulated biological fluids (e.g. gastric fluid, interstitial fluid, lysosomal fluid,…), represents relevant exposure routes. In this study, the oral route was investigated. Therefore, simulated gastric fluid, as provided in the SOP, was used to measure the release/dissolution of the elements of interest from Fatty acids, tall-oil manganese salts.

The temperature of the sampled test solutions including the Negative and Positive Control vessel at the 2-hours sampling point was between 36.9 °C and 37.1 °C and corresponded to the required test conditions of 37 °C ± 1 °C.

The measured pH of the test medium at the start of the test was 1.50, i.e. within the specifications of pH 1.5 ± 0.1. The pH measured in the additional test item vessel at the beginning of the trial was 1.50 for the 0.2 g/L and 2 g/L loadings, respectively, i.e. within the specifications of pH 1.5 ± 0.1. At the 2 hours endpoint of the test, the pH in the Negative Control vessel was 1.51 and 1.50 in the Positive Control. The pH in the test item vessels was between 1.50 and 1.51 for the vessels at the 0.2 g/L loadings and between 1.50 and 1.51 for the vessels at the 2 g/L loadings.

The Negative Control vessel showed no concentrations of manganese above the limit of quantification (i.e. accredited reporting limit) of 2 μg/L Mn.

The following observations could be made in the Positive Control item vessel containing a soluble form of manganese:
- The Positive Control item solution remained clear at the tested concentration level. No precipitation could be observed on the filter and in the Erlenmeyer flask at the end of the experiment.
- The recovery of the dissolved manganese concentration in the Positive Control item was 103 % and met the criterion (recovery: 100 % ± 10 %) as indicated in the SOP.

The following observations could be made in the test vessels with a loading of 0.2 g/L Fatty acids, tall-oil manganese salts:
- During the experiment (due to the shaking) the glass cover slips had turned upside down so that the test material was located between the bottom of Erlenmeyer flasks and the cover slip. It is not clear if and how this could have affected the results. Undissolved test item could be observed in the Erlenmeyer flasks at the end of the experiment.
- An average dissolved manganese concentration in solution of 0.401 ± 0.031 mg/L Mn (corresponding to a dissolved manganese concentration per g test item of 1.96 ± 0.12 mg/g Mn) was measured after 2 hours of exposure to the simulated gastric fluid (pH 1.5) with a between-vessel variation of 8 % which met the quality criteria at the 2 hours endpoint.
Based on the manganese content of the test item (i.e. 8.9 % Mn) and the average dissolved manganese concentration, a manganese release of 2.2 % could be calculated at the 2 hours endpoint.

The following observations could be made in the test vessels with a loading of 2 g/L Fatty acids, tall-oil manganese salts:
- During the experiment (due to the shaking) the glass cover slips had turned upside down so that the test material was located between the bottom of Erlenmeyer flasks and the cover slip. It is not clear if and how this could have affected the results. Undissolved test item could be observed in the Erlenmeyer flasks at the end of the experiment.
- An average dissolved manganese concentration in solution of 2.14 ± 0.16 mg/L Mn (corresponding to a dissolved manganese concentration per g test item of 1.07 ± 0.08 mg/g Mn) was measured after 2 hours of exposure to the simulated gastric fluid (pH 1.5) with a between-vessel variation of 7 % which met the quality criteria at the 2 hours endpoint.

Based on the manganese content of the test item (i.e. 8.9 % Mn) and the average dissolved manganese concentration, a manganese release of 1.2 % could be calculated at the 2 hours endpoint.

Conclusions:

During this study on Fatty acids, tall-oil manganese salts at a loading of 0.2 g/L in simulated gastric fluid (pH 1.5), it was shown that for manganese an average dissolved concentration in solution of 0.401 mg/L Mn (CVbetween-vessel = 8 %; N = 3) was found after 2 hours of extraction. This amount corresponds to a dissolved manganese concentration per g test item of 1.96 mg/g Mn or a manganese release of 2.2 % of the content.

During this study on Fatty acids, tall-oil manganese salts at a loading of 2 g/L in simulated gastric fluid (pH 1.5), it was shown that for manganese an average dissolved concentration in solution of 2.14 mg/L Mn (CVbetween-vessel = 7 %; N = 3) was found after 2 hours of extraction. This amount corresponds to a dissolved manganese concentration per g test item of 1.07 mg/g Mn or a manganese release of 1.2 % of the content.

The results are reliable since the test conditions stayed constant during the experiment. The recovery of Positive Control met the criteria of acceptance.

Executive summary:

Bioelution refers to the in vitro extraction methods used to measure the degree to which a substance (e.g., metal or mineral ion) is dissolved in artificial biological fluids. Simulated biological fluids represent relevant exposure routes. The release/dissolution of manganese ions from Fatty acids, tall-oil manganese salts in the simulated gastric fluid was measured. 

The resulting value is the “bioaccessibility”, and is defined as the “fraction of a substance that is soluble under physiological conditions and therefore potentially available for absorption into systemic circulation”. 

This study was conducted according to The Standard Operating Procedure for Bioelution Testing of Metals, Inorganic Metal Compounds, and Complex Metal-Containing Materials: Simulated Gastric Fluid (Eurometaux, November 2, 2019) in which the gastric medium will be prepared starting from a 0.1 M HCl solution. 

The extent of dissolution of Fatty acids, tall-oil manganese salts in the paste form (as received) was tested in a simulated gastric fluid at 37 °C and pH 1.5 for 2 hours (0.2 g/L and 2 g/L loadings) at an agitation speed of 100 revolutions per minute (rpm) for 1 hour followed by 1 hour of settling (without shaking). The bioelution endpoint is based on the dissolved manganese (Mn) concentrations obtained after 2 hours of extraction. 

The study was performed at ECTX. Analysis of the concentrations of dissolved manganese has been performed at WLN Business B.V. (The Netherlands), an ISO 17025 accredited laboratory, as delegated by ECTX. 

The measured pH of the test medium at the start of the test was 1.50, i.e. within the specifications of pH 1.5 ± 0.1. The pH measured in the additional test item vessel at the beginning of the trial was 1.50 for the 0.2 g/L and 2 g/L loadings, respectively, i.e. within the specifications of pH 1.5 ± 0.1. At the 2 hours endpoint of the test, the pH in the Negative Control vessel was 1.51 and 1.50 in the Positive Control. The pH in the test item vessels was between 1.50 and 1.51 for the vessels at the 0.2 g/L loadings and between 1.50 and 1.51 for the vessels at the 2 g/L loadings. 

The temperature of the sampled test solutions, including the Negative and Positive Control vessel was between 36.9 °C and 37.1 °C and corresponded to the required test conditions of 37 °C ± 1 °C. 

The Negative Control vessel showed no concentrations of manganese above the limit of quantification (i.e. accredited reporting limit) of 2 μg/L Mn. 

In the test item vessels with a loading of 0.2 g/L Fatty acids, tall-oil manganese salts, the following average dissolved manganese concentration in solution of: 

• 0.401 ± 0.031 mg/L Mn (CVbetween-vessel = 8 %; N = 3), corresponding to a dissolved manganese concentration per g test item of 1.96 ± 0.12 mg/g Mn, 

was measured after 2 hours of exposure to the simulated gastric fluid (pH 1.5). 

Based on the manganese content (i.e. 8.9 % Mn) in the test item and the average dissolved manganese concentrations in the test solutions, a release of: 

• 2.2 % of the content could be calculated at the 2 hours endpoint. 

 

In the test item vessels with a loading of 2 g/L Fatty acids, tall-oil manganese salts, the following average dissolved manganese concentration in solution of: 

• 2.14 ± 0.16 mg/L Mn (CVbetween-vessel = 7 %; N = 3), corresponding to a dissolved manganese concentration per g test item of 1.07 ± 0.08 mg/g Mn, 

was measured after 2 hours of exposure to the simulated gastric fluid (pH 1.5). 

Based on the manganese content (i.e. 8.9 % Mn) in the test item and the average dissolved manganese concentrations in the test solutions, a release of: 

• 1.2 % of the content could be calculated at the 2 hours endpoint. 

Description of key information

The dissolution of manganese from Fatty acids, tall-oil manganese salts after 2h in simulated gastric acid (pH 1.5) at 0.2 and 2 g/L loading is 2.2 and 1.2%, respectively, corresponding to 0.401 and 2.14 mg Mn/L.

The dissolution of Mn in simulated interstitial fluid (pH 7.4) at a loading of 2 g/L Fatty acids, tall-oil manganese salts is 0.16 and 1.5% after 24h and 168h, respectively, corresponding to 0.291 and 2.74 mg Mn/L.