Mobile phase— Prepare a filtered and degassed mixture of hexanes and tetrahydrofuran (97.75:2.25).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Pyrethrum Extract RSin hexanes,and dilute stepwise with hexanes to obtain a solution having a known concentration of about 0.5mg of pyrethrins per mL.

Assay preparation— Transfer an accurately weighed quantity of Extract,equivalent to about 50mg of pyrethrins,to a 100-mLvolumetric flask,dilute with hexanes to volume,and mix.

Chromatographic system (see Chromatography á621ñ)— The liquid chromatograph is equipped with a 240-nm detector and a 4.6-mm ×25-cm column that contains 5-µm packing L10.The flow rate is about 1.5mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the resolution,R,between the individual pyrethrin peaks is not less than 2.0;and the relative standard deviation for replicate injections is not more than 1.0%.

Procedure— Separately inject equal volumes (about 10µL)of the Standard preparationand the Assay preparationinto the chromatograph,record the chromatograms,and measure the responses for the six major peaks.The relative retention times are about 0.78to 0.80for jasmolin I;0.86to 0.87for cinerin I;1.0for pyrethrin I;1.95to 2.15for jasmolin II;2.15to 2.4for cinerin II;and 2.55to 2.88for pyrethrin II.Calculate the quantity,in mg,of pyrethrins in the portion of Extract taken by the formula: in which CSiis the concentration,in mg per mL,of the individual pyrethrin of interest in the Standard preparation;and rUiand rSiare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.Calculate the ratio of Pyrethrins Ito Pyrethrins IIin the portion of the Extract taken by the formula: in which Aand Bare the sums of the quantities of Pyrethrins Iand Pyrethrins II,respectively.