(iv) Its ultraviolet absorption spectrum is characteristic of a conjugated heptaene and is qualitatively the same as that of the candicidin working standard.

(2) Labeling. It shall be labeled in accordance with the requirements of § 432.5(b) of this chapter.

(3) Requests for certification; samples. In addition to the requirements of § 431.1 of this chapter, each such request shall contain:

(1) Results of tests and assays on the batch for potency, loss on drying, pH, and identity.

(ii) Samples required: 10 packages, each containing approximately 300 milligrams.

(b) Tests and methods of assay—(1) Potency. Proceed as directed in § 436.106 of this chapter, preparing the sample for assay as follows: Dissolve a portion of the sample in sufficient dimethylsulfoxide to yield an estimated concentration of 1,000 micrograms of candicidin activity per milliliter. Further dilute an aliquot with sterile distilled water to the reference concentration of 0.06 microgram of candicidin activity per milliliter (estimated).

(2) Loss on drying. Proceed as directed in § 436.200(b) of this chapter.

(3) pH. Proceed as directed in §436.202 of this chapter, using a 1 percent aqueous suspension.

(4) Identity—(i) Preparation of aqueous alcohol solution. Prepare an aqueous alcohol solution by mixing 53 volumes of ethyl alcohol and 47 volumes of water. (ii) Preparation of standard solution. Grind a small portion of the candicidin working standard to a fine powder with a mortar and pestle. Accurately weigh an amount equivalent to 20,000 micrograms of candicidin activity and transfer it to a 100-milliliter volumetric flask. Add about 50 milliliters of the aqueous alcohol solution and shake to effect complete dissolution. Bring to volume with the aqueous alcohol solution and mix well. Transfer a 25-milliliter aliquot to a 100-milliliter volumetric flask and bring to volume with the aqueous alcohol solution. This solution contains 50 micrograms of candicidin activity per milliliter.

(iii) Preparation of sample solution. Proceed as directed in paragraph (b)(4)(ii) of this section.

(iv) Procedure. Using a suitable recording spectrophotometer, record the absorption spectra of the standard solution and the sample solution between the wavelengths of 330 and 410 nanometers with the aqueous alcohol solution as the reference solution. Compare the absorption spectra of the standard solution and the sample solution. They should exhibit absorption maxima and minima at the same wavelengths, which are approximately 342, 359, 378, and 397 nanometers for the maxima and 348, 366, and 390 nanometers for the minima.

[39 FR 19134, May 30, 1974, as amended at 44 FR 30333, May 25, 1979; 49 FR 2243, Jan. 19, 1984]

(a) Requirements for certification—(1) Standards of identity, strength, quality, and purity. Griseofulvin is a microsize, white to pale-cream compound with the following chemical name: 7-chloro2',4,6-trimethoxy-6'ẞ

methylspiro[benzofuran-2(3H),1′[2]cyclohexene]-3,4′-dione. It is so purified and dried that:

(1) Its griseofulvin content is not less than 900 micrograms and not more than 1,050 micrograms of griseofulvin per milligram.

(ii) [Reserved]

(iii) Its loss on drying is not more than 1.0 percent.

(iv) Its melting point, after drying, is not less than 217° C. and not more than 224° C.

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(v) Its specific rotation dimethylformamide at 25° C. is not less than +348° and not more than +364°.

(vi) Its ultraviolet absorption spectrum in methyl alcohol compares qualitatively with that of the griseofulvin reference standard.

(vii) Its residue on ignition is not more than 0.2 percent.

(viii) Its heavy metals content is not more than 25 parts per million.

(ix) Its specific surface area is not less than 1.3 and not more than 1.7 square meters per gram.

(x) It is crystalline.

(2) Labeling. It shall be labeled in accordance with the requirements of § 432.5(b) of this chapter.

(3) Requests for certification; samples. In addition to the requirements of § 431.1 of this chapter, each such request shall contain:

(i) Results of tests and assays on the batch for griseofulvin content, loss on drying, melting point, specific rotation, identity, residue on ignition, heavy metals, specific surface area, and crystallinity.

(ii) Samples required: 10 packages, each containing not less than 1 gram. (b) Tests and methods of assay—(1) Griseofulvin content (gas liquid chromatography). Proceed as directed

§ 436.321 of this chapter. (2) [Reserved]

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hol to obtain a concentration of 10 micrograms of griseofulvin per milliliter and mix well. (The standard solution can be kept under refrigeration and used for up to 1 month.) Record the ultraviolet absorption spectrum of solutions of the sample and standard from 240 to 320 nanometers. The spectral curves shall be similar, and each shall have a maximum at 292+ 2 nanometers and a minimum at 269± 2 nanometers.

(7) Residue on ignition. Proceed as directed in § 436.207 of this chapter.

(8) Heavy metals. Proceed as directed in § 436.208 of this chapter.

(9) Specific surface area—(i) Procedure. Determine the apparent particle size in microns by the air-permeation method, using a suitable subsieve sizer. Weigh 1.819 grams ±0.001 gram of the sample and transfer to the compression tube of the apparatus. Compact the sample with moderate pressure so that it has a uniform porosity. Pass compressed dry air through the sample and measure the air pressure with a water manometer. Observe the porosity and calculate the apparent particle size from the instrument equation or read it from a chart that has been calculated in accordance with the equation. Repeat the readings at successively higher degrees of compaction until the apparent particle size reaches a minimum. Calculate the observed specific surface area (SSA) in square meters per gram of sample, as follows:

(iii) Calculations. Calculate the corrected specific surface area of the sample as follows:

SSA of sample

(10) Crystallinity. Proceed as directed in § 436.203(a) of this chapter.

[39 FR 19134, May 30, 1974, as amended at 44 FR 20660, Apr. 6, 1979; 50 FR 19920, May 13, 1985]

§ 449.40 Natamycin.

(a) Requirements for certification-(1) Standards of identity, strength, quality, and purity. Natamycin is 22-[(3 - amino 3,6 dideoxy -B-D-mannopyranosyl) oxy] - 1,3,26-trihydroxy- 12-methyl-10oxo-6,11,28

trioxatricyclo [22.3.1.05.7] octacosa8,14,16,18,20

pentaene-25-carboxylic acid. It is an off-white to cream colored powder which may contain up to 3 moles of water. It is practically insoluble in water, slightly soluble in methanol, and soluble in glacial acetic acid and dimethylformamide. It is so purified and dried that:

(i) Its potency is not less than 900 micrograms of natamycin per milligram on an anhydrous basis.

(ii) Its moisture content is not less than 6.0 percent and not more than 9.0 percent.

(iii) Its pH in a 1 percent aqueous suspension is not less than 5.0 and not more than 7.5.

(iv) It passes the identity test. (v) It is crystalline.

(2) Labeling. It shall be labeled in accordance with the requirements of § 432.5 of this chapter.

(3) Requests for certification; samples. In addition to complying with the requirements of § 431.1 of this chapter, each such request shall contain:

(1) Results of tests and assays on the batch for potency, moisture, pH, identity, and crystallinity.

(ii) Samples required: 10 packages, each containing approximately 500 milligrams.

(b) Tests and methods of assay. Dilute solutions of natamycin are very sen

sitive to light and should be kept in the dark as much as possible or substantial decomposition will take place.

(1) Potency. Proceed as directed in § 436.105 of this chapter, preparing the sample for assay as follows: Dissolve an accurately weighed sample in dimethylsulfoxide and further dilute with sufficient dimethylsulfoxide to give a concentration of 100 micrograms milliliter (estiof natamycin per milliliter mated). Further dilute with 0.2M potassium phosphate buffer, pH 10.5 (solution 10), to the reference concentration of 5.0 micrograms of natamycin per milliliter (estimated).

(2) Moisture. Proceed as directed in § 436.201 of this chapter.

(3) pH. Proceed as directed in §436.202 of this chapter, using a 1.0 percent aqueous suspension.

(4) Identity. Accurately weigh approximately 50 milligrams of the sample into a 200-milliliter volumetric flask. Add approximately 5.0 milliliters of distilled water, water, and completely moisten the sample. Then add approximately 100 milliliters of an acid-alcohol solvent (0.1 percent glacial acetic acid in methyl alcohol) and stir or shake mechanically in the dark until solution is complete. Dilute to volume with the acid-alcohol solvent. Transfer 2.0 milliliters of this solution to a 100milliliter volumetric flask and dilute to volume with the acid-alcohol solvent. Using a suitable spectrophotometer with 1-centimeter cells and the acid-alcohol solvent as a blank, record the ultraviolet absorption spectrum from 215 to 330 nanometers. The spectrum compares qualitatively to that of the natamycin working standard similarly treated.

(5) Crystallinity. Proceed as directed in § 436.203(a) of this chapter.

[43 FR 55384, Nov. 28, 1978, as amended at 46 FR 16684, Mar. 13, 1981]

§ 449.50 Nystatin.

(a) Requirements for certification—(1) Standards of identity, strength, quality, and purity. Nystatin is the yellow to light-tan compound of a kind of nystatin or a mixture of two or more such compounds. It is very slightly soluble in water, moderately soluble in methyl alcohol, butyl alcohol, or propyl alcohol. It is so purified and dried that:

(i) Its potency is not less than 4,400 units of nystatin per milligram; except, if it is packaged for extemporaneous preparation of oral suspensions, its potency is not less than 5,000 units of nystatin per milligram.

(ii) [Reserved]

(iii) Its loss on drying is not more than 5.0 percent.

(iv) Its pH in a 3 percent aqueous suspension is not less than 6.5 and not more than 8.0.

(v) It passes the identity test.

(vi) If it is packaged for extemporaneous preparation of oral suspensions, it passes the suspendibility test. (vii) If it is packaged for extemporaneous preparation of oral suspensions, it is crystalline.

(2) Labeling. It shall be labeled in accordance with the requirements of § 432.5(b) of this chapter.

(3) Requests for certification; samples. In addition to the requirements of § 431.1 of this chapter, each such request shall contain:

(i) Results of tests and assays on the batch for potency, loss on drying, pH, and identity. In addition, if it is packaged for extemporaneous preparation of oral suspensions, results of tests and assays on the batch for suspendibility and crystallinity.

(ii) Samples required on the batch: 10 packages, each containing approximately 300 milligrams.

(b) Tests and methods of assay-(1) Potency. Proceed as directed in § 436.105 of this chapter, preparing the sample for assay as follows: Dissolve an accurately weighed sample in sufficient dimethylformamide to give a nystatin concentration of 400 units per milliliter (estimated). Further dilute with 10 percent potassium phosphate buffer, pH 6.0 (solution 6), to the reference concentration of 20 units of nystatin per milliliter (estimated).

(2) [Reserved]

(3) Loss on drying. Proceed as directed in § 436.200(b) of this chapter.

(4) pH. Proceed as directed in § 436.202 of this chapter, using a 3 percent aqueous suspension of the drug.

(5) Identity. Weigh approximately 100 milligrams of the sample into a 200milliliter, glass-stoppered, volumetric flask. Add 50 milliliters of absolute methyl alcohol and 10 milliliters of glacial acetic acid. When the sample has dissolved, dilute to volume with methyl alcohol. Transfer 2 milliliters of this solution to a 100-milliliter volumetric flask and dilute to volume with methyl alcohol. Use the same dilution of acetic acid in methyl alcohol as the blank. Immediately determine the absorption peaks at 230, 291, 305, and 319 nanometers, and the shoulders at 279±2 nanometers, using a suitable ultraviolet spectrophotometer and quartz cells. Set the instrument to 100 percent transmission with the blank. If a recording spectrophotometer is used, record the ultraviolet absorption spectrum from 220 nanometers to 350 nanometers. If a nonrecording spectrophotometer is used, the exact positions of the peaks and shoulder should be determined for the particular instrument used. The ratio of the two absorbances (A230/A279)

should be not less than 0.90 and not more than 1.25.

(6) Suspendibility test. Transfer 200 milligrams of the sample into a 250milliliter beaker containing 200 milliliters of water. Swirl the suspension gently with a stirring rod. Allow the beaker to remain still for 2 minutes and observe the bottom. It passes the test if the powder remains in suspension. If a significant amount of sediment is observed, withdraw an accurately measured aliquot of the undisturbed suspension and assay as directed in §449.150c(b)(1) of this chapter. It passes the test if the suspension contains not less than 90 percent of the number of units of nystatin that it is represented to contain.

(7) Crystallinity. Proceed as directed in § 436.203(a) of this chapter.

[39 FR 19134, May 30, 1974, as amended at 39 FR 43833, Dec. 19, 1974; 49 FR 5098, Feb. 10, 1984; 50 FR 19920, May 13, 1985]

(a) Requirements for certification—(1) Standards of identity, strength, quality, and purity. Amphotericin B oral suspension is a mixture of amphotericin B with one or more suitable and harmless preservatives, colorings, sweetening ingredients, flavorings, buffer substances, lubricants, suspending agents, and sequestrants in an aqueous vehicle. Each milliliter contains 100 milligrams of amphotericin B. Its potency is satisfactory if it is not less than 90 percent and not more than 125 percent of the number of milligrams of amphotericin B that it is represented to contain. Its pH is not less than 4.5 and not more than 6.0. The amphotericin B conforms to the standards prescribed by § 449.4(a)(1).

(2) Labeling. It shall be labeled in accordance with the requirements of § 432.5 of this chapter.

(3) Requests for certification; samples. In addition to complying with the requirements of § 431.1 of this chapter, each such request shall contain:

(1) Results of tests and assays on: (a) The amphotericin B used in making the batch for potency, amphotericin A content, loss on drying, pH, residue on ignition, and identity.

(b) The batch for potency and pH. (ii) Samples required:

(a) The amphotericin B used in making the batch: 10 packages, each containing approximately 500 milligrams.

(b) The batch: A minimum of 5 immediate containers.

(b) Tests and methods of assay-(1) Potency. Proceed as directed in § 436.105 of this chapter, preparing the sample for assay as follows: Place an accurately measured representative portion into a high-speed glass blender with sufficient dimethylsulfoxide to give a stock solution of convenient concentration. Blend for 3 to 5 minutes. Dilute an aliquot of the stock stock solution solution with dimethylsulfoxide to give a concentration of 20 micrograms of amphotericin B per milliliter (estimated). Further dilute an aliquot with 0.2M potassium phosphate buffer, pH 10.5 (solution 10), to the reference concentration of 1.0

§ 449.120a Griseofulvin tablets.

(a) Requirements for certification—(1) Standards of identity, strength, quality, and purity. Griseofulvin tablets are tablets composed of griseofulvin, with or without one or more suitable fillers, colorings, lubricants, and binders. Each tablet contains 125, 250, or 500 milligrams of griseofulvin. The griseofulvin content is satisfactory if it is not less than 90 percent and not more than 115 percent of the number of milligrams of griseofulvin that it is represented to contain. The loss on drying is not more than 5.0 percent. The tablets shall disintegrate within 1 hour. The griseofulvin used conforms to the standards prescribed by §449.20(a)(1).

(2) Labeling. It shall be labeled in accordance with the requirements of § 432.5 of this chapter.

(3) Requests for certification; samples. In addition to the requirements of § 431.1 of this chapter, each such request shall contain:

(1) Results of tests and assays on:

(a) The griseofulvin used in making the batch for griseofulvin content, loss on drying, melting point, specific rotation, identity, residue on ignition, heavy metals, specific surface area, and crystallinity.

(b) The batch for griseofulvin content, loss on drying, and disintegration time.

(ii) Samples required:

(a) The griseofulvin used in making the batch: 10 packages, each containing not less than 1 gram.

(b) The batch for griseofulvin content, loss on drying, and disintegration time.

(b) Tests and methods of assay-(1) Griseofulvin content (gas liquid chromatography). Proceed as directed § 436.321 of this chapter, except:

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(1) Prepare the sample solution as follows: Accurately weigh 20 tablets