Fatty acids, tall-oil, manganese salts

Dissociation constant

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

dissociation constant

Remarks:

dissociation of salts into individual ions

Type of information:

experimental study

Adequacy of study:

key study

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 followed

Principles of method if other than guideline:

zero-sink test that measures the dissociation of the metal carboxylate substance in two contrasting test media (low and neutral pH) based on the free metal ion concentration in the presence of a selective adsorbent that acts as a zero-sink for the metal cation

GLP compliance:

no

Dissociating properties:

yes

No.:

#1

Temp.:

20 °C

Remarks on result:

other: 101±<1% dissociation after 1h in simulated gastric fluid (pH 1.5)

No.:

#2

Temp.:

20 °C

Remarks on result:

other: 102±2% dissociation after 1h in simulated interstitial fluid (pH 7.4)

Simulated gastric fluid 

Solution composition

The measured metal concentration of the reference compound and metal carboxylate (MC) in the gastric medium is given in Table 5. The MC is not completely dissolved in the gastric fluid after 12h, the percentage dissolved material at 1 g compound/L is 46 ± 1%, which is higher than in pure water (Table 2). The reference compound is completely dissolved (> 100%) in the gastric fluid. 

Table 5 Measured total metal concentration (C_M) dissolved during 12h in the simulated gastric fluid and percent solubility (f_sol) ± standard deviation. For the metal carboxylate (MC), the nominal concentrations are the expected concentrations in solution based on the metal content of the compound and using the exact weighted mass of the compounds. 

	Measured	Nominal
Compound	CM	CM	fsol
	μM	μM	%
Reference salt
MnCl2.4H2O	32.1 ± 1.2	30	107 ± 4
	3233 ± 24	3000	108 ± 1
Metal Carboxylate
Tall oil, Mn	592 ± 25	1296	46 ± 1

Free metal ion concentration measurement 

For the reference salts (MnCl₂.4H₂O) the Mn concentration in solution decreased to < 2% of the initial concentration after 1h due to sorption of the metal on the resin (Table 6). The measured Mn concentration in all diluted test solutions is still above the quantification limit (LOQ) of the ICP-MS analysis. The concentration in solution is the free metal in equilibrium with the resin. Binding of Mn to the resin is instantaneous since no differences are observed between measurements at 1h, 6h and 24h. The strong binding of the metal to the resin at the composition and pH of the gastric medium is in agreement with speciation calculations using Visual Minteq. The model predicts an equilibrium Mn concentration in solution that corresponds well with the measured concentration (Table 6, reference salts). 

The initial concentration Mn of the MC after 12h dissolution in gastric medium, but before addition of the zero sink resin is within the concentration range of the reference samples. The Mn concentration in solution of the MC also decreased to < 1% of the initial concentration after 1h due to sorption of the metal on the resin. The MC undergoes a dissociation reaction and exists in equilibrium with its dissociation products. The dissociated metal reacts with the resin. The metal concentration in solution of the MC is thus, in theory, the free metal in equilibrium with the zero-sink and undissociated MC. The free metal concentration is calculated from the reference compounds and is after 1h equal to 11.5 ± 0.5 μM Mn. This approximates the total measured metal concentration in solution of 4.6 ± 0.2 μM Mn. After 1h, the calculated dissociated fraction of metal carboxylate is 101 ± < 1%. Therefore, this test indicates that the MC dissociates fully in the test media representing gastric fluid. The metal concentration in solution for the reference salt and MC remained constant in time after 1h. This indicates that the MC dissociates instantaneously (< 1h) in the test media relevant for gastric fluid. 

Table 6 Measured metal concentration in solution at the start (C_M,i) and after equilibration (C_M,e) and concentration sorbed on the zero sink (C_M,s) ± standard deviation at different time points for the reference salts and the metal carboxylate (MC) dissolved in the gastric medium. For the MC, the equilibrium free metal concentration C_M²⁺ is calculated from the reference compounds and used to calculate the dissociated fraction (f_diss). The C_M²⁺ values of the reference compounds are also calculated with ion exchange principles using Visual Minteq. 

		Measured	Measured	Calculated	Predicted	Calculated
Compound	time	CM, i	CM,e	CM,s	CM^2+	CM^2+	fdiss
	h	μM	μM	μmol/g	μM	μM	%
Reference salt	0	32.1 ± 1.2
MnCl2.4H2O	1		0.22 ± 0.01	3.19 ± 0.11	0.25
30 μM	6		0.22 ± 0.01	3.19 ± 0.12	0.25
	24		0.22 ± < 0.01	3.19 ± 0.12	0.25
Reference salt	0	3233 ± 24
MnCl2.4H2O	1		63.1 ± 1.2	317 ± 3	28.4
3000 μM	6		61.9 ± 1.3	317 ± 3	28.4
	24		63.4 ± 0.6	317 ± 2	28.4
Metal Carboxylate	0	592 ± 25
Tall oil, Mn	1		4.6 ± 0.2	58.8 ± 2.5		11.5 ± 0.5	101 ± < 1
	6		4.6 ± 0.2	58.8 ± 2.5		11.3 ± 0.5	101 ± <1
	24		4.5 ± 0.2	58.8 ± 2.5		11.5 ± 0.5	101 ± < 1

Simulated interstitial fluid 

Solution composition 

The measured metal concentration of the reference compound and MC in the interstitial fluid is given in Table 7. The MC has very low solubility in the interstitial fluid, after 12h the percentage dissolved material of the MC at 1 g compound/L is less than 1%, which is lower than in pure water and in the gastric medium. The reference compound MnCl2.4H2O at a concentration of 30 μM Mn is completely dissolved (> 100%) in the interstitial fluid. At a higher concentration of 3000 μM Mn the reference salt precipitated in the interstitial fluid, and is therefore not included in the analysis. 

Table 7 Measured total metal concentration (C_M) dissolved during 12h in the simulated interstitial fluid and percent solubility (f_sol) ± standard deviation. For the metal carboxylate (MC), the nominal concentrations are the expected concentrations in solution based on the metal content of the compound and using the exact weighted mass of the compounds. 

	Measured	Nominal
Compound	CM	CM	fsol
	μM	μM	%
Reference salt
MnCl2.4H2O	32.7 ± 2.3	30.0	109
Metal Carboxylate
Tall oil, Mn	4.46 ± 0.11	1296	< 1%

Free metal ion concentration measurement 

For the reference salt (MnCl₂.4H₂O) the Mn concentration in solution decreased to about 20% of the initial concentration after 1h due to sorption of the metal on the resin (Table 8). The measured Mn concentration in all diluted test solutions is above the quantification limit (LOQ) of the ICP-MS analysis. The concentration in solution is the free metal in equilibrium with the resin. Binding of Mn to the resin is instantaneous since no differences are observed between measurements at 1h, 6h and 24h. The binding of the metal to the resin in the interstitial fluid is less strong than in the acid gastric medium due to competition between protons and metal cations to bind to the resin, that binding is also well predicted with speciation calculations using Visual Minteq (Table 8). The model predicts an equilibrium Mn concentration in solution that corresponds well with the measured concentration. At the pH and composition of the interstitial medium the reference MnCl₂.4H₂O sample with the higher concentration (3000 μM Mn) precipitated, and is therefore not included in the analysis. 

The Mn concentration in solution of the MC decreased to about 20% of the initial concentration after 1h due to sorption of the metal on the resin. The MC undergoes a dissociation reaction and exists in equilibrium with its dissociation products. The dissociated metal reacts with the resin. The metal concentration in solution of the MC is thus, in theory, the free metal in equilibrium with the zero-sink and undissociated MC. The predicted free metal concentration after 1h is 0.90 ± 0.04 μM Mn, this is equal to the measured Mn concentration of 0.82 ± 0.04 μM Mn. After 1h, the calculated dissociated fraction of metal carboxylate is 102 ± 2.Therefore, this test indicates that the MC dissociates completely in the test media representing interstitial fluid. The metal concentration in solution for the reference salt and MC remained constant in time after 1h which means that the dissociation is instantaneously (< 1h) in the test media relevant for interstitial fluid. 

Table 8 Measured metal concentration in solution at the start (C_M,i) and after equilibration (C_M,e) and concentration sorbed on the zero sink (C_M,s) ± standard deviation at different time points for the reference salts and the metal carboxylate (MC) dissolved in the interstitial medium. For the MC, the equilibrium free metal concentration C_M²⁺ is calculated from the reference compounds and used to calculate the dissociated fraction (f_diss). The C_M²⁺ values of the reference compounds are also calculated with ion exchange principles using Visual Minteq. 

		Measured	Measured	Calculated	Predicted	Calculated
Compound	time	CM,i	CM,e	CM,s	CM^2+	CM^2+	fdiss
	h	μM	μM	μmol/g	μM	μM	%
Reference salt	0	32.7 ± 2.3
MnCl2.4H2O	1		6.44 ± 0.18	1.31 ± 0.11	8.53
30 μM	6		6.28 ± 0.22	1.32 ± 0.11	8.53
	24		6.24 ± 0.27	1.32 ± 0.10	8.53
Metal Carboxylate	0	4.46 ± 0.11
Tall oil, Mn	1		0.82 ± 0.04	0.18 ± 0.01		0.90 ± 0.04	102 ± 2
	6		0.80 ± 0.01	0.18 ± < 0.01		0.88 ± 0.02	102 ± <1
	24		0.81 ± 0.08	0.18 ± < 0.01		0.87 ± 0.01	101 ± 2

Conclusions:

Tall oil, manganese salt dissociates completely (>= 101%) in the test media representing gastric fluid and interstitial fluid. The manganese concentration in solution for the reference salt MnCl2.4H2O and for tall oil, manganese salt remained constant in time after 1h which means that the dissociation is instantaneously (< 1h) in both test media. 

Reference 2

Endpoint:

dissociation constant

Remarks:

dissociation of salts into individual ions

Type of information:

experimental study

Adequacy of study:

key study

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:

according to guideline

Guideline:

OECD Guideline 112 (Dissociation Constants in Water)

GLP compliance:

no

Dissociating properties:

yes

No.:

#1

Temp.:

25 °C

Remarks on result:

other: 127±3% dissociation after 3 weeks in water

Solution composition 

The measured concentration of metal and organic carbon in solution of the reference compounds and metal carboxylate (MC) is given in Table 2. The percentage dissolved material of the MC prepared at 1 g/L is low (< 1%) after 1h. The evolution of the concentration of metal and organic carbon in time shows that the compound slowly dissolves. Because of the low concentration, an extra time point after 3 weeks was included to increase detection for the conductivity measurements. After 3 weeks about 10% of the MC is dissolved, the measured Mn concentration in solution (170 ± 7 μM Mn) is about half of the expected concentration of 410 μM Mn based on the solubility of the metal carboxylate (265 ± 86 mg MC/L, on a Mn content of 8.5 %). The molar metal/organic carbon ratio in solution ranged 0.041-0.045 throughout the experiment. Multi-element analysis with ICP-MS did not show important concentration of other elements in solution, which could interfere with the electrical conductivity measurements. Both reference compounds are completely dissolved after 1h (f_sol > 100%). 

Table 2 Measured total concentration metal (C_M,T) or organic carbon (C_OC,T) at 1 g/L and percent solubility (f_sol) ± standard deviation at different samplings. For the metal carboxylate (MC), the nominal concentrations are the expected concentrations in solution based on the metal content of the compound and using the exact weighted mass of the compounds. 

		Measured	Measured	Nominal
Compound	time	CM,T	COC,T	CT*	fsol
	h	mM	mM	mM	%
Reference salt
MnCl2.4H2O	1	5.42 ± 0.05		4.94 ± 0.19	110 ± 5
	6	5.67 ± 0.01			115 ± 4
	24	5.61 ± 0.03			114 ± 4
Reference salt
NaC8H16O2	1	50.2 ± 3.7		49.0 ± 2.7	102 ± 2
	6	49.3 ± 2.2			101 ± 1
	24	49.6 ± 3.0			101 ± <1
Metal Carboxylate		μM	μM	μM
Tall oil, Mn (UVCB)	1	2.66 ± 1.21	65.1 ± 8.0	1648 ± 30	< 1
	6	7.06 ± 2.50	170 ± 51		<1
	24	17.0 ± 4.5	381 ± 41		1 ± <1
	3 weeks	170 ± 7			10 ± <1

* For MnCl₂.4H₂O and the MC, the nominal concentration is the metal concentration, for NaC₈H₁₆O₂ the nominal concentration is the organic carbon concentration. 

Electrical conductivity measurements 

The measured electrical conductivity and concentration of metal and ligand for the MC and reference compounds in function of time is given in Table 3. The dissociated fraction is calculated from the measured electrical conductivity using the tabulated diffusion coefficients. The dissociated fraction of reference compounds is expected to approach 100%, the deviation for both reference compounds (<100% for MnCl₂.4H₂O and >100% for NaC₈H₁₆O₂) might be related to inaccuracies of the model, i.e. the theoretical λ0, values. For the MC tall oil, the exact composition is not known or variable (UVCB) which results in uncertainty for the method. The diffusion coefficient for tall oil in Table 1 is approximately calculated from the composition of tall oil (mainly oleic and linoleic acid) using the online epa tool to calculate diffusion coefficients based on composition. An exact reference compound for the carboxylate is not included, instead the C8 2-ethylhexanoate is included as reference. The dissociation in the reference compounds is instantaneously as expected. For the MC, both the electrical conductivity and metal concentration increased with time. The conductivity measured between 1h and 24h is below the detection limit of the electrode used, therefore an extra measurement after 3 weeks is included. The results after 3 weeks suggest that the MC is fully dissociated (127 ± 3%) at that time. 

Table 3 Measured electrical conductivity (σ) and concentration of metal (C_M,T) and carboxylate ligand (C_L,T) of reference salts and MC (in duplicate) at different points in time. The dissociated metal concentration (C_Mⁿ⁺) and dissociated fraction (f_diss) ± standard deviation are calculated using the diffusion coefficients in Table 1. 

Compound		Measured	Measured	Measured	Measured	Measured	Measured	Calculated	Calculated
		Rep. 1	Rep. 1	Rep. 1	Rep. 2	Rep. 2	Rep. 2
	time	σ	CM,T	CL,T	σ	CM,T	CL,T	CM^n+	fdiss
	h	μS/cm	mM	mM	μS/cm	mM	mM	mM	%
Reference salt
MnCl2.4H2O	1	1184	5.46		1187	5.38		4.57 ± 0. 01	84 ± 1
	6	1219	5.66		1218	5.67		4.69 ± < 0. 1	83 ± < 1
	24	1173	5.59		1184	5.63		4.54 ± < 0.03	81 ± < 1
Reference salt
NaC8H16O2	1	517		5.95	551		6.61	7.03 ± 0.32	112 ± 3
	6	519		5.97	553		6.36	7.05 ± 0.32	114 ± < 1
	24	503		5.93	535		6.46	6.83 ± 0.20	110 ± 2
Metal carboxylate	time	σ	CM,T	Coc,T	σ	CM,T	Coc,T	CMn+	fdiss
	h	μS/cm	μM	μM	μS/cm	μM	μM	μM	%
Tall oil, Mn	1	0	3.5	59	0	1.8	71	< DL
	6	4	8.8	206	1	5.3	134	< DL
	24	11	20	410	7	14	352	< DL
	3 weeks	32	175		29	164		218 ± 15	127 ± 3

Conclusions:

The conductivity measured between 1h and 24h is below the detection limit of the electrode used, therefore an extra measurement after 3 weeks is included. The results after 3 weeks suggest that tall oil, manganese salt is fully dissociated (127 ± 3%) at that time.

Description of key information

The electrical conductivity test in pure water and the zero-sink test simulated gastric medium (pH 1.5) and simulated interstitial medium (pH 7.4) all conclude that the dissociation of tall oil, manganese salt is complete (≥ 101%). The zero-sink test indicated that the dissociation is instantaneous (within 1h). Due to the limited solubility of the substance, the electrical conductivity test did not provide results on the dissociation rate within 24h.