Milk Thistle Tablets
DEFINITION
Milk Thistle Tablets are prepared from Powdered Milk Thistle Extract. They contain NLT 90.0% and NMT 110.0% of the labeled amount of silymarin as silybin (C25H22O10), calculated as the sum of silydianin, silychristin, silybin A, silybin B, isosilybin A, and isosilybin B.
IDENTIFICATION
• A. Thin-Layer Chromatographic Identification Test
Standard solution:
1.0 mg/mL of USP Silydianin RS in methanol
Sample solution:
Equivalent to 5 mg/mL of silymarin from finely powdered Tablets (NLT 20) in methanol. Shake for 1 min, and sonicate for 10 min. Allow to stand for 15 min before use.
Chromatographic system
Mode:
TLC
Adsorbent:
0.25-mm layer of chormatographic silica gel, typically 20 cm long
Application volume:
10 µL
Developing solvent system:
Freshly prepared mixture of chloroform, acetone, and anhydrous formic acid (75: 16.5: 8.5)
Spray reagent A:
10-mg/mL solution of 2-aminoethyl diphenylborinate in methanol
Spray reagent B:
50-mg/mL solution of polyethylene glycol 4000 in alcohol
Analysis
Samples:
Standard solution and Sample solution
Develop the chromatograms until the solvent front has moved about three-fourths of the plate, and dry it for 30 min in a current of cold air. Spray the plate with Spray reagent A, allow to dry, and then spray with Spray reagent B. One h later, examine the plate under long-wavelength UV light.
Acceptance criteria:
The chromatogram of the Sample solution exhibits an intense green-blue fluorescent zone at an RF value of 0.5 (presence of silybin) and exhibits a gray-blue spot at RF value of 0.4, corresponding to a spot observed in the chromatogram of the Standard solution. The chromatogram of the Sample solution may exhibit other colored zones: an intense green-blue zone at an RF value of 0.25 (presence of silychristin) and a red-orange zone at an RF value of 0.3 (presence of taxifolin).
• B. HPLC Identification Test
Analysis:
Proceed as directed for Content of Silymarin.
Acceptance criteria:
The chromatogram of the Sample solution exhibits peaks for silydianin, silychristin, silybin A, silybin B, isosilybin A, and isosilybin B at retention times that correspond to those of the Milk thistle standard solution.
STRENGTH
• Content of Silymarin
Solution A:
Methanol, phosphoric acid, and water (20: 0.5: 80)
Solution B:
Methanol, phosphoric acid, and water (80: 0.5: 20)
Mobile phase:
See Table 1.
Table 1
| Time (min) |
Solution A (%) |
Solution B (%) |
|---|---|---|
| 0 | 0 | 0 |
| 5 | 85 | 15 |
| 20 | 55 | 45 |
| 40 | 55 | 45 |
| 41 | 85 | 15 |
| 55 | 85 | 15 |
Milk thistle standard solution:
0.7 mg/mL of USP Powdered Milk Thistle Extract RS in methanol. Sonicate for 20 min to dissolve.
Silybin standard solutions:
0.20, 0.02, and 0.004 mg/mL of USP Silybin RS in methanol. Pass through a membrane filter having a 0.45-µm or finer pore size.
Sample solution:
Weigh and finely powder NLT 20 Tablets. Transfer an accurately weighed quantity of the powder equivalent to 100 mg of silymarin to a 100-mL volumetric flask, add 90 mL of methanol, and sonicate for 20 min with occasional shaking. Cool to 20
, and dilute with methanol to volume. Filter through a membrane with a 0.45-µm or finer pore size.
, and dilute with methanol to volume. Filter through a membrane with a 0.45-µm or finer pore size.
Chromatographic system
Mode:
LC
Detector:
UV 288 nm
Column:
4.6-mm × 15-cm; 5-µm packing L1
Column temperature:
40
Flow rate:
1 mL/min
Injection size:
10 µL
System suitability
Sample:
Milk thistle standard solution
[Note—For the relative retention times, see Table 2. ]
Table 2
| Name | Relative Retention Time |
|---|---|
| Silychristin | 0.68 |
| Silydianin | 0.73 |
| Silybin A | 1.00 |
| Silybin B | 1.05 |
| Dehydrosilybin | 1.09 |
| Isosilybin A | 1.15 |
| Isosilybin B | 1.19 |
Suitability requirements
Chromatogram similarity:
The chromatogram from the Milk thistle standard solution is similar to the reference chromatogram provided with the lot of USP Powdered Milk Thislte Extract RS being used.
Resolution:
NLT 1.0 between silybin A and silybin B
Tailing factor:
0.8–2.0
Relative standard deviation:
NMT 2.0% for the sum of peak responses due to silybin A and silybin B
Analysis
Samples:
Milk thistle standard solution, each of the Silybin standard solutions, and Sample solution
Identify the peaks due to silychristin, silydianin, silybin A, silybin B, isosilybin A, and isosilybin B by comparison of the chromatogram of the Milk thistle standard solution with the reference chromatogram, and measure the peak areas of the relevant peaks. Plot the areas of the sum of silybin A and silybin B peaks versus the concentration of USP Silybin RS in the Silybin standard solutions, and obtain a regression line for calibration.
Separately calculate the content, in mg, of each relevant component of silymarin as silybin (C25H22O10) in the portion of Tablets taken:
Result = C × V
| C | = | = concentration of the relevant component in the Sample solution as interpolated in the calibration graph (mg/mL) |
| V | = | = volume of the Sample solution (mL) |
Calculate the content of silymarin, as a percentage, in the portion of Tablets taken:
Result = CS × (AWT/W) × (100/L)
| CS | = | = sum of the contents of each relevant component of silymarin in the portion of Tablets taken (mg) |
| AWT | = | = average weight of the Tablets (mg/Tablet) |
| W | = | = weight of the portion of Tablets taken (mg) |
| L | = | = labeled amount of silymarin (mg/Tablet) |
Acceptance criteria:
90.0%–110.0%
PERFORMANCE TESTS
• Disintegration and Dissolution 
2040
:
Meet the requirements for Dissolution
2040:
Meet the requirements for Dissolution
Buffer:
6.9 g/L of monobasic sodium phosphate and 1.52 g/L of sodium hydroxide in water. Adjust to a pH of 7.5.
Medium:
Buffer containing 2% lauryl sulfate; 900 mL
Apparatus 2:
100 rpm
Time:
45 min
Determine the amount of silymarin as silybin (C25H20O10) dissolved by using the method in the test for Content of Silymarin, making any necessary modifications.
Tolerances:
NLT 75% of the labeled amount of silymarin as silybin (C25H22O10) is dissolved.
• Weight Variation 2091:
Meet the requirements
2091:
Meet the requirements
CONTAMINANTS
• Microbial Enumeration Tests 2021:
The total bacterial count does not exceed 104 cfu/g, the total combined molds and yeasts count does not exceed 103 cfu/g, and the enterobacterial count does not exceed 102 cfu/g.
2021:
The total bacterial count does not exceed 104 cfu/g, the total combined molds and yeasts count does not exceed 103 cfu/g, and the enterobacterial count does not exceed 102 cfu/g.
• Absence of Specified Microorganisms 2022:
It meets the requirements of the tests for absence of Salmonella species and Escherichia coli.
2022:
It meets the requirements of the tests for absence of Salmonella species and Escherichia coli.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in tight, light-resistant containers.
• Labeling:
The label states the Latin binomial and, following the official name, the article from which the Tablets were prepared. The label also indicates the content of silymarin, in mg/Tablet.
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Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Natalia Davydova
Scientific Liaison 1-301-816-8328 |
(DS2010) Monographs - Dietary Supplements |
| 2021 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| 2022 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 1390
Pharmacopeial Forum: Volume No. 28(2) Page 422