Accessed from 128.83.63.

20 by nEwp0rt1 on Wed Nov 30 20:46:11 EST 2011

1190 Andrographis / Dietary Supplements USP 35

Relative ADDITIONAL REQUIREMENTS

Analyte Retention Time • PACKAGING AND STORAGE: Preserve in well-closed
14-Deoxy-11,12-didehydroan- 1.31 containers, protected from light and moisture, and store
drographolide at room temperature.
Andrograpanin 1.50 • LABELING: The label states the Latin binomial and,
following the official name, the parts of the plant
Separately calculate the percentages of andrographo- contained in the article.
lide, neoandrographolide, 14-deoxy-11,12- • USP REFERENCE STANDARDS 〈11〉
didehydroandrographolide, and andrograpanin in the USP Andrographolide RS
portion of Powdered Andrographis taken: USP Powdered Andrographis Extract RS

Result = (CS/W) × (rU/rS) × 10F

rU = peak response for each diterpene lactone from .

the Sample solution Powdered Andrographis Extract

rS = peak response for andrographolide from
Standard solution A DEFINITION
CS = concentration of USP Andrographolide RS in Powdered Andrographis Extract is prepared from An-
Standard solution A (mg/mL) drographis by extraction with methanol or alcohol. The
W = weight of Powdered Andrographis taken to ratio of plant material to extract is between 15:1 and
prepare the Sample solution (g) 10:1. It contains NLT 90.0% and NMT 110.0% of the
F = conversion factor for each analyte (1.00 for labeled amount of diterpene lactones, calculated on the
andrographolide, 3.90 for dried basis as the sum of andrographolide, neoandro-
neoandrographolide, 1.45 for 14-deoxy- grapholide, 14-deoxy-11,12-didehydroandrographolide,
11,12-didehydroandrographolide, and 2.65 and andrograpanin. The content of 14-deoxy-11,12-
for andrograpanin) didehydroandrographolide is NMT 15% of the total
Acceptance criteria: NLT 1.0%, on the dried basis, of diterpene lactones. It may contain suitable added sub-
the sum of the percentages of andrographolide, stances as carriers.
neoandrographolide, 14-deoxy-11,12-
didehydroandrographolide, and andrograpanin IDENTIFICATION
• A. THIN-LAYER CHROMATOGRAPHIC IDENTIFICATION TEST
IMPURITIES 〈201〉
Inorganic Impurities Standard solution 1: Use Standard solution A, prepared
• ARTICLES OF BOTANICAL ORIGIN, Acid-Insoluble Ash 〈561〉: as directed in the test for Content of Diterpene Lactones.
NMT 3.0% Standard solution 2: Sonicate for 10–15 min a quantity
• HEAVY METALS, Method II 〈231〉: NMT 20 ppm of USP Powdered Andrographis Extract RS, equivalent to
Organic Impurities about 15 mg of diterpene lactones, in 25 mL of metha-
• PROCEDURE: ARTICLES OF BOTANICAL ORIGIN, General nol. Centrifuge, and use the supernatant.
Method for Pesticide Residues Analysis 〈561〉: Meets the Sample solution: Sonicate for 10–15 min a quantity of
requirements Powdered Andrographis Extract, equivalent to about 15
SPECIFIC TESTS mg of diterpene lactones, in 25 mL of methanol. Centri-
• BOTANIC CHARACTERISTICS fuge, and use the supernatant.
Macroscopic: It is a grayish brown powder. Adsorbent: Chromatographic silica gel mixture with an
Histology average particle size of 10–15 µm (TLC plates)
Microscopic examination: It reveals cells of the upper Application volume: 10 µL, as 5–10 mm bands
and lower epidermis of the leaves, some cells Developing solvent system: Chloroform, acetone, and
containing large cystoliths, up to 36 µm in diameter toluene (2:2:1)
and 180 µm long, with a hilum-shaped scar in the large Spray reagent: A mixture containing 1% vanillin in alco-
end; 1–4 celled nonglandular hairs; disk-shaped hol and 10% sulfuric acid in alcohol (1:1)
glandular hairs, 8-celled head and very short stalk; Analysis
diacytic stomata mostly on the lower epidermis; stem Samples: Standard solution 1, Standard solution 2, and
epidermal cells, some cells containing cystoliths, Sample solution
stomata, nonglandular hairs and glandular hairs similar Use a saturated chamber. Develop until the solvent
to those of the leaves; thin-walled parenchyma cells; front has moved up about 90% of the plate. Remove
collenchyma cells; acicular phloem fibers; tracheids; the plate from the chamber. Dry, and spray with Spray
vessels, with spiral and scalariform thickening. reagent. Heat for 5–10 min at 100°, and examine
• LOSS ON DRYING 〈731〉: Dry 1.0 g of Powdered under visible light.
Andrographis at 105° for 3 h: it loses NMT 12.0% of its Acceptance criteria: The Sample solution exhibits three
weight. main grayish blue zones with RF values of approximately
• ARTICLES OF BOTANICAL ORIGIN, Total Ash 〈561〉: NMT 0.4, 0.6, and 0.8 that correspond in position and color
15%, determined on 1.0 g of Powdered Andrographis to zones in Standard solution 2. Standard solution 1 ex-
• ARTICLES OF BOTANICAL ORIGIN, Alcohol-Soluble Extractives, hibits a grayish blue zone due to andrographolide at an
Method 2 〈561〉: NLT 8.0% RF of about 0.4. The Sample solution exhibits a zone simi-
• MICROBIAL ENUMERATION TESTS 〈2021〉: The total aerobic lar in color and RF value to that due to andrographolide
bacterial count does not exceed 105 cfu/g; the total in Standard solution 1.
combined molds and yeasts count does not exceed 103 • B. The Sample solution in the test for Content of Diterpene
cfu/g; and the bile-tolerant Gram-negative bacteria does Lactones shows a main peak at a retention time corre-
not exceed 103 cfu/g. sponding to that of andrographolide in Standard solution
• MICROBIOLOGICAL PROCEDURES FOR ABSENCE OF SPECIFIED A. Identify other diterpene lactone peaks in the Sample
MICROORGANISMS 〈2022〉: Meets the requirements of the solution by comparison with Standard solution B and the
tests for absence of Salmonella species and Escherichia coli reference chromatogram provided with the lot of USP
Powdered Andrographis Extract RS. The Sample solution
shows additional peaks corresponding to neoandrograph-
olide, 14-deoxy-11,12-didehydroandrographolide, and
andrograpanin.

Official from May 1, 2012

Copyright (c) 2011 The United States Pharmacopeial Convention. All rights reserved.
Accessed from [Link] by nEwp0rt1 on Wed Nov 30 20:46:11 EST 2011

USP 35 Dietary Supplements / Arginine 1191

COMPOSITION Relative
• CONTENT OF DITERPENE LACTONES Analyte Retention Time
Solution A: Dissolve 0.14 g of potassium dihydrogen Andrographolide 1.00
phosphate in 900 mL of water, add 0.5 mL of phos- Neoandrographolide 1.16
phoric acid, dilute with water to 1000 mL, mix, filter, 14-Deoxy-11,12-didehydroan- 1.31
and degas. drographolide
Solution B: Use filtered and degassed acetonitrile.
Standard solution A: Dissolve a weighed quantity of Andrograpanin 1.50
USP Andrographolide RS in methanol to obtain a solu- Separately calculate the percentages of andrographo-
tion having a known concentration of about 1.0 mg/mL. lide, neoandrographolide, 14-deoxy-11,12-
Transfer 5.0 mL of this solution to a 10-mL volumetric didehydroandrographolide, and andrograpanin in the
flask, dilute with acetonitrile to volume, and mix. portion of Powdered Andrographis Extract taken:
Standard solution B: Transfer an amount of USP Pow-
dered Andrographis Extract RS, equivalent to about 25 Result = (rU/rS) × (CS/W) × 5F
mg of diterpene lactones, to a 50-mL volumetric flask,
add 25 mL of methanol, heat gently for 15–20 min, di- rU = peak response for each diterpene lactone from
lute with acetonitrile to volume, and mix. Before injec- the Sample solution
tion, pass through a membrane filter of 0.45-µm or finer rS = peak response for andrographolide from
pore size, discarding the first 5 mL of the filtrate. Standard solution A
Sample solution: Transfer a weighed amount of Pow- CS = concentration of USP Andrographolide RS in
dered Andrographis Extract, equivalent to about 25 mg Standard solution A (mg/mL)
of diterpene lactones, to a 50-mL volumetric flask, add W = weight of Powdered Andrographis Extract
25 mL of methanol, heat gently for 15–20 min, dilute taken to prepare the Sample solution (g)
with acetonitrile to volume, and mix. Before injection, F = conversion factor for each analyte (1.00 for
pass through a membrane filter of 0.45-µm or finer pore andrographolide, 3.90 for
size, discarding the first 5 mL of the filtrate. neoandrographolide, 1.45 for 14-deoxy-
Mobile phase: See the gradient table below. 11,12-didehydroandrographolide, and 2.65
for andrograpanin)
Time Solution A Solution B Acceptance criteria: 90.0%–110.0%, on the dried
(min) (%) (%) basis, of the labeled amount of diterpene lactones
0 95 5 calculated as the sum of the percentages of
18 55 45 andrographolide, neoandrographolide, 14-deoxy-11,12-
didehydroandrographolide, and andrograpanin
25 20 80
28 20 80 IMPURITIES
35 55 45 Inorganic Impurities
40 95 5 • HEAVY METALS, Method II 〈231〉: NMT 20 ppm
45 95 5
Organic Impurities
• PROCEDURE: ARTICLES OF BOTANICAL ORIGIN, General
Chromatographic system Method for Pesticide Residues Analysis 〈561〉: Meets the
(See Chromatography 〈621〉, System Suitability.) requirements
Mode: LC SPECIFIC TESTS
Detector: UV 223 nm • LOSS ON DRYING 〈731〉: Dry 2.0 g at 105° for 3 h: it loses
Column: 4.6-mm × 25-cm; 5-µm packing L1 NMT 5.0% of its weight.
Flow rate: 1.5 mL/min • MICROBIAL ENUMERATION TESTS 〈2021〉: The total aerobic
Injection size: 20 µL microbial count does not exceed 104 cfu/g. The total
System suitability combined yeasts and molds count does not exceed 103
Samples: Standard solution A and Standard solution B cfu/g.
Suitability requirements • MICROBIOLOGICAL PROCEDURES FOR ABSENCE OF SPECIFIED
The chromatogram from Standard solution B is similar to MICROORGANISMS 〈2022〉: It meets the requirements of
the reference chromatogram provided with the lot of the tests for absence of Salmonella species and Escherichia
USP Powdered Andrographis Extract RS. coli.
Column efficiency: NLT 5000 theoretical plates,
Standard solution A ADDITIONAL REQUIREMENTS
Tailing factor: NMT 1.5 for the andrographolide • PACKAGING AND STORAGE: Preserve in well-closed
peak, Standard solution A containers, protected from light and moisture, and store
Relative standard deviation: NMT 2.0%, determined at controlled room temperature.
from the andrographolide peak for replicate injec- • LABELING: The label states the Latin binomial and,
tions, Standard solution A following the official name, the part of the plant from
Resolution: NLT 5 between the neoandrographolide which the article was prepared. It meets other labeling
and 14-deoxy-11,12-didehydroandrographolide requirements under Botanical Extracts 〈565〉.
peaks, Standard solution B • USP REFERENCE STANDARDS 〈11〉
Analysis USP Andrographolide RS
Samples: Standard solution A, Standard solution B, and USP Powdered Andrographis Extract RS
Sample solution
Using Standard solution A, Standard solution B, and the
reference chromatogram provided with the lot of USP
Powdered Andrographis Extract RS being used, identify
the retention times of the peaks corresponding to dif-
ferent diterpene lactones. The approximate relative re- Arginine—see Arginine General Monographs
tention times of the different diterpene lactones are
provided in the following table:

Official from May 1, 2012

Copyright (c) 2011 The United States Pharmacopeial Convention. All rights reserved.
 Accessed from [Link] by nEwp0rt1 on Wed Nov 30 20:46:11 EST 2011

1192 Arginine / Dietary Supplements USP 35

Arginine Hydrochloride—see Arginine rU = peak response from the Sample solution

rS = peak response from the Standard solution
Hydrochloride General Monographs CS = concentration of the Standard solution
(mg/mL)
CU = nominal concentration of Arginine in the
Sample solution (mg/mL)
Acceptance criteria: 90.0%–110.0% of the labeled
amount of arginine (C6H14N4O2)
Arginine Capsules
.

PERFORMANCE TESTS
DEFINITION • DISINTEGRATION AND DISSOLUTION OF DIETARY SUPPLEMENTS
Arginine Capsules contain NLT 90.0% and NMT 110.0% of 〈2040〉: Meet the requirements for Dissolution
the labeled amount of arginine or arginine hydrochloride Medium: 0.1 N hydrochloric acid; 900 mL
in an amount equivalent to arginine (C6H14N4O2). Apparatus 2: 100 rpm
Time: 60 min
IDENTIFICATION Standard solution: Proceed as directed in the Procedure
• A. THIN-LAYER CHROMATOGRAPHIC IDENTIFICATION TEST for Strength.
〈201〉 Sample solution: Sample per Disintegration and
Standard solution: 1.5 mg/mL of USP L-Arginine RS or Dissolution of Dietary Supplements 〈2040〉. Dilute with
USP Arginine Hydrochloride RS in water Medium to a concentration similar to that of the
Sample solution: Weigh the content of NLT 20 Cap- Standard solution.
sules, mix, and transfer a portion of the content, equiva- Analysis: Determine the amount of arginine dissolved in
lent to about 150 mg of arginine, to a 100-mL volumet- the Procedure for Strength, making any necessary
ric flask, add 80 mL of water, sonicate for 15 min, dilute modifications.
with water to volume, mix, and filter. Tolerances: NLT 75% of the labeled amount of arginine
Application volume: 5 µL (C6H14N4O2) is dissolved.
Developing solvent system: Isopropyl alcohol and am- • WEIGHT VARIATION OF DIETARY SUPPLEMENTS 〈2091〉: Meet
monium hydroxide (7:3) the requirements
Spray reagent: 2 mg/mL of ninhydrin in a mixture of
butyl alcohol and 2 N acetic acid (95:5) ADDITIONAL REQUIREMENTS
Analysis: Proceed as directed for Chromatography 〈621〉, • PACKAGING AND STORAGE: Preserve in tight, light-resistant
Thin-Layer Chromatography. Dry the plate at 100°–105° containers.
until the ammonia disappears completely. Spray with the • LABELING: The label states the form of arginine that is
Spray reagent, and heat at 100°–105° for about 15 min. used and the equivalent amount of arginine.
Examine the plate under white light. • USP REFERENCE STANDARDS 〈11〉
Acceptance criteria: The principal spot from the Sample USP L-Arginine RS
solution corresponds in appearance and RF value to that USP Arginine Hydrochloride RS
of the Standard solution.
• B. The retention time of the major peak of the Sample
solution corresponds to that from the Standard solution,
as obtained in the Strength.
.

Arginine Tablets
STRENGTH
• PROCEDURE DEFINITION
Buffer: 6.9 mg/mL of monobasic sodium phosphate in Arginine Tablets contain NLT 90.0% and NMT 110.0% of
water. Adjust with phosphoric acid to a pH of 3.5. the labeled amount of arginine or arginine hydrochloride
Solution A: 0.5 mg/mL of 1-octanesulfonic acid sodium in an amount equivalent to arginine (C6H14N4O2).
salt in Buffer
Mobile phase: Solution A and acetonitrile (95:5) IDENTIFICATION
Standard solution: 1.5 mg/mL of USP L-Arginine RS or • A. THIN-LAYER CHROMATOGRAPHIC IDENTIFICATION TEST
USP Arginine Hydrochloride RS in Buffer 〈201〉
Sample solution: Weigh the content of NLT 20 Cap- Standard solution: 1.5 mg/mL of USP L-Arginine RS or
sules, mix, and transfer a portion of the content, equiva- USP Arginine Hydrochloride RS in water
lent to about 150 mg of arginine, to a 100-mL volumet- Sample solution: Weigh and finely powder NLT 20 Tab-
ric flask, add 80 mL of Buffer, sonicate for 15 min, dilute lets, mix, and transfer a portion of the powder, equiva-
with Buffer to volume, mix, and filter. lent to about 150 mg of arginine, to a 100-mL volumet-
Chromatographic system ric flask, add 80 mL of water, sonicate for 15 min, dilute
(See Chromatography 〈621〉, System Suitability.) with water to volume, mix, and filter.
Mode: LC Application volume: 5 µL
Detector: UV 215 nm Developing solvent system: Isopropyl alcohol and am-
Column: 4.6-mm × 25-cm; packing L7 monium hydroxide (7:3)
Flow rate: 0.8 mL/min Spray reagent: 2 mg/mL of ninhydrin in a mixture of
Injection size: 10 µL butyl alcohol and 2 N acetic acid (95:5)
System suitability Analysis: Proceed as directed for Chromatography 〈621〉,
Sample: Standard solution Thin-Layer Chromatography. Dry the plate at 100°–105°
Suitability requirements until the ammonia disappears completely. Spray with the
Column efficiency: NLT 1500 theoretical plates Spray reagent, and heat at 100°–105° for about 15 min.
Relative standard deviation: NMT 2.0% from the ar- Examine the plate under white light.
ginine peak, in repeated injections Acceptance criteria: The principal spot from the Sample
Analysis solution corresponds in appearance and RF value to that
Samples: Standard solution and Sample solution from the Standard solution.
Calculate the percentage of the labeled amount of argi- • B. The retention time of the major peak of the Sample
nine in the portion of Capsules taken: solution corresponds to that of the Standard solution, as
obtained in Strength.
Result = (rU/rS) × (CS/CU) × 100

Official from May 1, 2012

Copyright (c) 2011 The United States Pharmacopeial Convention. All rights reserved.