Registration Dossier

Physical & Chemical properties

Additional physico-chemical information

Currently viewing:

Administrative data

Link to relevant study record(s)

Description of key information

Removal of Water from Bis-Aminopropyl Diglycol Dimaleate Including Development of an Analytical Method and Determination of Water Content

SPONSOR: Liqwd, Inc, 1482 East Valley Road 701, Santa Barbara, CA 93108, USA

TEST FACILITY: Charles River Laboratories Den Bosch BV, Hambakenwetering 7, 5231 DD ‘s-Hertogenbosch, The Netherlands

Test Facility Study No. 512944

Test period: 6/6/2017 to 23/6/2017

SUMMARY

Bis-aminopropyl diglycol dimaleate was freeze dried. The water content, 4,7,10-trioxa-1,13 -tridecanediamine content and maleic acid content were determined before and after the freeze drying process. The results are given below.

   Water content [% w/w]  4,7,10-trioxa-1,13-tridecanediamine
content [g/kg] 
  Maleic acid content
[g/kg] 
 
     Not corrected for
water content 
Corrected for
water content  
 Not corrected for
water content 
Corrected for
water content  
undried test item (207396/A) 74 1201 4631 1292 4942
dried test item (207396/B) 5.4 4263 4513 2834 3004

(1) Chromatogram is shown in Figure 4 (attached)

(2) Chromatogram is shown in Figure 5 (attached)

(3) Chromatogram is shown in Figure 7 (attached)

(4) Chromatogram is shown in Figure 8 (attached)

Purpose of the study

The purpose of the study was to remove water by freeze-drying process from Bis-Aminopropyl Diglycol Dimaleate. The water content before and after freeze-drying process was determined by Karl Fisher analysis. An analytical method was developed for the determination of identity of the cationic part and anionic part of the test item.

MATERIALS

Test item

Test item information

Test item       207396/A

Identification       Bis-Aminopropyl Diglycol Dimaleate

Appearance              Clear colourless to pale yellow liquid (determined by Charles River Den Bosch)

Batch                     3

Purity/Composition       Not indicated

Test item storage       At room temperature

Stable under storage conditions until       30 September 2016 (expiry date)

Study specific test item information

Purity/composition correction factor       No correction factor required

Test item handling       No specific handling conditions required

Stability at higher temperatures       Yes, maximum temperature: 40°C, maximum duration: 60 minutes

Analytical Standard – AS1645

Container              A1

Identification       4,7,10-Trioxa-1,13-tridecanediamine

CAS Number       4246-51-9

Molecular formula       C10H24N2O3

Molecular weight       220.31

Appearance       Colourless liquid

Batch       STBC8331V

Purity       99.9%

Storage conditions       At room temperature

Stable under storage conditions until       11 April 2017

Supplier              Sigma Aldrich Chemie GmbH, Steinheim, Germany

Article number       369519

Analytical Standard – AS1646

Container       A1

Identification       Maleic acid

CAS Number       110-16-7

Molecular formula       C4H4O4

Molecular weight       116.07

Appearance       White powder

Batch       SLBP7337V

Purity       100.0 %

Storage conditions At room temperature

Stable under storage conditions until       30 November 2021

Supplier              Sigma Aldrich Chemie GmbH, Steinheim, Germany

Article number       M0375

Reference Standard – RS528

Identification number       RS528

Container       B1

Identification       HYDRANAL®-Water Standard 1.0

Appearance       Clear colourless liquid (determined by Charles River Den Bosch)

Batch       SZBD0090V

Water content       1.000 mg/g (expanded uncertainty = 0.003 mg/g)

Storage conditions       At room temperature

Stable under storage conditions until       14 December 2017

Reference Standard – RS529

Identification number       RS529

Container       A1

Identification       HYDRANAL®-Water Standard 0.1

Appearance       Clear colourless liquid

Batch       SZBF1660V

Water content       0.1 mg/g

Storage conditions       In refrigerator (2-8°C)

Stable under storage conditions until       04 June 2017

DRYING PROCESS

Apparatus

Freeze dryer Christ Alpha 1-4 LD plus (Salm & Kipp, Breukelen, The Netherlands)

Performance freeze drying process

An accurately weighed amount of approximately 65 g of test item 207396/A was poured into a glass container and stored at ≤ -80°C for 1 day prior to the freeze drying process. The frozen sample was freeze dried for 6 days. After this period a yellow-orange viscous liquid was obtained. The dried test item was collected, register with test item number / batch number 207396/B and stored at room temperature.

LC-MS ANALYSIS

Reagents

Water              Tap water purified by a Milli-Q water purification system (Millipore, Bedford, MA, USA)

Acetonitrile       Biosolve, Valkenswaard, The Netherlands

Ammonium acetate       Biosolve

Formic acid                    Biosolve

Sodium hydroxide       Merck, Darmstadt, Germany

All reagents were of analytical grade, unless specified otherwise.

Performance of the study

An ultra performance liquid chromatographic method with mass spectrometric detection (UPLC-MS) for the analysis of the cationic part of the test items was developed.

An ultra performance liquid chromatographic method with tandem mass spectrometric detection (UPLC-MS/MS) for the analysis of the anionic part of the test items was developed.

Research performed on method development included, but was not limited to, the selection of a solvent to dissolve the test item, a suitable ionization mode and an appropriate combination of column (e.g. stationary phase, dimensions) and mobile phase. Optimizations were performed to enhance retention-time stability, limit carry-over and improve the peak shape within the selected calibration range.

Method development is archived in the raw data, only the conditions of the final test are issued in the report. The conditions of the final test were approved in the raw data.

The test was performed as follows:

Calibration curves

Calibration solutions were analysed in duplicate. The response of the calibration solutions was correlated with concentration of the analytical standards (AS1645 and AS1646) using regression analysis with a 1/concentration2 weighting factor. Calibration curves with a coefficient of correlation (r) of > 0.99 and back calculated accuracies of the calibration solutions in the range 85-115% were accepted.

Analysis of the test samples

An amount of undried test item (207396/A) and dried test item (207396/B) were accurately weighed, if necessary dissolved in a suitable solvent and analysed by single injection. The concentration of 4,7,10-trioxa-1,13-tridecanediamine and maleic acid were determined for each test sample.

Analytical method

Analytical conditions – 4,7,10-trioxa-1,13-tridecanediamine

Instrument              Acquity UPLC system (Waters, Milford, MA, USA)

Detector       Xevo TQ-S mass spectrometer (Waters)

Column       Acclaim Mixed-Mode Wax-1, 150 mm  4.6 mm i.d., dp = 5 µm (Thermo Scientific, Waltham, MA, USA)

Column temperature       40°C  1°C

Injection volume       5 µL

Mobile phase       0.1% formic acid in 25/75 (v/v) acetonitrile/water

Flow       0.6 mL/min

MS detection

Ionisation source       ESI+

Cone voltage       30 V

Quantification       m/z 221.1

Analytical conditions – maleic acid

Instrument        Acquity UPLC system (Waters, Milford, MA, USA)

Detector            Xevo TQ-S mass spectrometer (Waters)

Column              Acquity UPLC BEH Amide, 50 mm  2.1 mm i.d., dp = 1.7 µm (Waters)

Column temperature     40°C  1°C

Injection volume             5 µL

Mobile phase1                A – 10 mM ammonium acetate in 95/5 (v/v) acetonitrile/water

B - 10 mM ammonium acetate in 50/50 (v/v) acetonitrile/water

1adjusted at pH 9 with sodium hydroxide

Gradient

Time [minutes]

%A

%B

0

99.9

0.1

0.4

99.9

0.1

0.5

60

40

2

30

70

2.1

99.9

0.1

5

99.9

0.1

 

Flow     0.5 mL/min

MS detection

Ionisation source             ESI-

Cone voltage                    40V

Collision energy               11 eV

Quantification                  m/z 115.0  m/z 71.0

 

Preparation of solutions

Stock solutions

Stock solutions of AS1645 were prepared in acetonitrile at concentrations of 1000 and 2000 mg/L.

Stock solutions of AS1646 were prepared in acetonitrile at concentrations of 1000 mg/L.

Calibration solutions

Five calibration solutions in the concentration range of 0.1 – 1 mg/L were prepared from two stock solutions of AS1645. The end solution of the calibration solutions was acetonitrile.

Five calibration solutions in the concentration range of 0.1 – 1 mg/L were prepared from two stock solutions of AS1646. The end solution of the calibration solutions was 10 mM ammonium acetate in 50/50 (v/v) acetonitrile/water.

Test samples

Approximately 100 mg undried test item (207396/A) and dried test item (207396/B) were accurately weighed into volumetric flasks of 100 mL. The flasks were filled up to mark with acetonitrile and further diluted to obtain end solutions of acetonitrile, 10 mM ammonium acetate in 50/50 (v/v) acetonitrile/water and concentrations within the calibration ranges.

 

Formulas

Response (R)                     Peak area of the test item [units]

Response factor (Rf)       Rf= R/CN

where:

CN= nominal concentration [mg/L]

Linear calibration curve R = aCN+ b

here:

a = slope [units x L/mg]

b = intercept [units]

Analysed concentration (CA) = (R – b) / a x (V x d) / (w x (1 - %H2O) = mg/g

where:

w = weight sample [mg]

V = volume volumetric flask [mL]

d = dilution factor

%H2O = water content of the test item [% w/w]

 

Quadratic calibration curve R = aCN2+ bCN+ c

where:

a = quadratic regression factor [units x (L/mg)2]

b = linear regression factor [units x L/mg]

c = intercept [units]

 

Analysed concentration (CA) = (-b + √(b2– 4a x (c-R))) / 2a x (Vd) / (w x (1 – H2O) = mg/g

where:

w = weight sample [mg]

V = volume volumetric flask [mL]

d = dilution factor

%H2O = water content of the test item [% w/w]

 

Results

Calibration curve – AS1645

The calibration line was constructed excluding one data point, since the back calculated accuracy was > 15% from the nominal concentration Figure 1 (attached) illustrates the calibration curve and the table below shows the statistical parameters. There was a quadratic relationship between response and test item concentration in the range of 0.1 – 1 mg/L (in end solution). Since the coefficient of correlation (r) was > 0.99 and the back calculated accuracies of the remaining data points were in the range 85-115% the calibration line was accepted.

Statistical parameters of the calibration curve – AS1645

Quadratic regression factor

 -1.60 x 107

Linear regression factor

3.21 x 107

Intercept

-2.32 x 106

Weighting factor

1/concentration2

r

0.996

 

Calibration curve – AS1646

The calibration line was constructed using all data points Figure 2(attached) illustrates the calibration curve and the table below shows the statistical parameters. There was a linear relationship between response and AS1646 concentration in the range of 0.1 – 1 mg/L (in end solution). Since the coefficient of correlation (r) was > 0.99 and the back calculated accuracies of the data points were in the range 85-115% the calibration line was accepted.

Statistical parameters of the calibration curve – AS1646

Slope

47.0 x 102

 

Intercept

-26.6

 

Weighting factor

1/concentration2

r

0.998

 

Analysis of the test samples

Chromatograms of a AS1645 calibration solution, undried test item (207396/A) and dried test item (207396/B) are shown in Figure 3, Figure 4 and Figure 5, respectively (attached).

Chromatograms of a AS1646 calibration solution, undried test item (207396/A) and dried test item (207396/B) are shown in Figure 6, Figure 7 and Figure 8, respectively (attached).

Content of 4,7,10-trioxa-1,13-tridecanediamine and maleic acid for the undried test item (207396/A) and dried test item (207396/B) are given in Table 3 and Table 4 respectively (attached).

4,7,10-trioxa-1,13-tridecanediamine content

 

Sample weight (mg)

Amount (mg)

Content (g/kg)

 

 Not corrected for
water content

Corrected for
water content
 

 Not corrected for
water content

Corrected for
water content
 

undried test item (207396/A)

138

35.8

16.6

120

463

dried test item (207396/B)

100

94.8

42.7

426

451

Maleic acid content

 

Sample weight (mg)

Amount (mg)

Content (g/kg)

 

 Not corrected for
water content

Corrected for
water content
 

 Not corrected for
water content

Corrected for
water content
 

undried test item (207396/A)

106

27.6

13.7

129

494

dried test item (207396/B)

94.4

89.3

26.8

283

300

 

Conclusion

The 4,7,10-trioxa-1,13-tridecanediamine and maleic acid contents of the undried test item (207396/A) and dried test item (207396/B) were determined by LC-MS(/MS). The results of the study samples after correction of the sample weights for the water content are:

 

 

4,7,10-trioxa-1,13-tridecanediamine
content [g/kg]

Maleic acid content
[g/kg]

 

 Not corrected for
water content

Corrected for
water content
 

 Not corrected for
water content

Corrected for
water content
 

undried test item (207396/A)

120

463

129

494

dried test item (207396/B)

426

451

283

300

 

DETERMINATION OF WATER CONTENT BY KARL FISHER

8.1. Reagents

Hydranal® Coulomat AK                              Sigma Aldrich, Steinheim, Germany

Hydranal® Coulomat CG-K                          Sigma Aldrich, Steinheim, Germany

Methanol anhydrous,                                  VWR International, Leuven, Belgium

All reagents were of analytical grade, unless specified otherwise.

 

8.2. Performance of the test

The water content in the undried and dried test item was determined by a Coulometric Karl Fisher titration using a Metrohm 831 KF Coulometer (Metrohm, Herisau, Switzerland). The undried and dried test item were analysed in duplicate.

The system consisted of a reaction vessel which was equipped with a double platinum electrode used for the instrumental end point indication. In addition to the double platinum electrode, a generator electrode was also present in the reaction vessel. Prior to titration, the reaction vessel was filled with Hydranal® Coulomat AK reagent and the generator electrode was filled with Hydranal® Coulomat CG-K reagent. The generator electrode and the reaction vessel were connected via a porous membrane which facilitates the conduction of electrical current between the two compartments but prevents the exchange of fluids. The reaction vessel was equipped with a septum via which the test solutions were introduced using a syringe. Titration was performed by applying a current to the generator electrode which results in the formation of iodine (I2) in the reaction vessel which is consumed in the Karl Fisher titration process.

On each day of analysis, the water content of the methanol was determined and the water content for the samples diluted with this methanol was corrected for it.

Prior to analysis of a sample, the system was first titrated water free. Subsequently a sample was introduced and titrated. The 831 KF Coulometer directly calculates the water content (in %) of the sample using the sample weight, no external calculations are required. Because the undried test item was dissolved in a solvent prior to measurement, the calculations described in paragraph 8.3 were performed to calculate the water content.

 

8.3. Formulas

Test item fraction of the sample solution fTS= (wTS) / (wM+ wTS) x 100 %

where:

fTS: fraction of test item in the sample solution [%]

wTS: weight of test item in the sample solution [g]

wM: weight of methanol in the sample solution [g]

 

Methanol fraction of the sample solution fM= (wM) / (wM+ wTS) x 100 %

where:

fM: fraction of methanol in the sample solution [%]

 

Water content of test substance acTS= (wSS– wcM(fM/100)) / (fTS) x 100 %

 

where:

wcTS: water content of the test item [%]

wcSS: water content of the sample solution [%]

wcM: water content of methanol [%]

 

Analytical method

Analytical conditions

Instrument                       831 KF Coulometer (Metrohm, Herisau, Switzerland)

Magnetic stirrer               Ti Stand 703 (Metrohm)

 

Calibration

The validity of the system was verified using two water standards with a certified water amount of 0.1 and 1.0 mg/g (Sigma-Aldrich).

Duplicate samples of each standard were weighed and the water content was determined by titration.

 

Sample pre-treatment – undried test item

19.9 to 39.5 mg undried test item was dissolved in approximately 4 g methanol.

A blank titration with methanol was performed to determine the water content in the solvent.

 

Sample pre-treatment – dried test item

17.7 to 24.5 mg dried test item was transferred to a glass piece.

The glass piece was introduced into the titration vessel.

 

Results

The table below shows the calibration results. The system was found to be accurate for the determination of the water content of the test item.

Date of analysis Water

Water standard (mg/g)

Certified water content (mg/g)

Analysed water content (mg/g)

Recovery (%)

Mean recovery (%)

25-May-16

0.1

0.1

0.104

104

105

 

1

1

1

100

100

 

 

 

1.01

101

 

 

The water content results of the undried and dried test item are given in the table below.

Date of analysis

Test item

Weight test item (g)

Analysed water (%)

Water content (%)

Mean water content (%)

25-May-16

undried

19.9

0.328

73.9

74

 

 

39.5

0.69

73.4

 

25-May-16

dried

17.7

5.42

5.42

5.4

 

 

24.5

5.43

5.43

 

 

Conclusion

The water contents of the undried test item (207396/A) and dried test item (207396/B) were determined by a Coulometric Karl Fischer titration.

Water contents of the undried and dried test item are:

Test item

Water content (% w/w)

Undried (207396/A)

74

Dried (207396/B)

5.4

 

Additional information