A safety study of a B-class CpG ODN in Sprague-Dawley rats.

Oligodeoxynucleotides containing CpG motifs (CpG ODNs) are potent immune activators and are being tested as anti-tumor, antimicrobial agents and as adjuvants in vaccines. Little has been reported, however, about the systematic and comprehensive safety evaluation on repeated CpG ODN administration. To investigate the safety profile of a newly developed CpG ODN, CpG 684, we conducted a 28-day repeated dose toxicity study in rats, at dose levels of 5, 20 and 150 µg CpG 684 per rat. No abnormalities in clinical observations, growth, urinalysis and bone marrow cell counts were found in CpG 684 treated rats. CpG 684 was proved biologically active, capable of up-regulating the expressions of CD40 and CD86 molecules. The monocyte numbers were increased at the dose levels of 20 and 150 µg per rat. The spleen weights were increased in female rats at the dose level of 150 µg per rat. Microscopically, 5, 20 and 150 µg per rat CpG 684 caused local inflammatory cell infiltration and hyperplasia of fibrous tissue at injection sites; the treatment of 5 and 150 µg per rat CpG 684 induced enhanced inflammatory reaction in inguinal lymphoid tissue, and the dose of 150 µg per rat induced cell hyperplasia in white pulp of spleen and white pulp expansion. CpG 684 at 150 µg per rat led to decreases in peripheral lymphocyte, serum globulin, glucose, alkaline phosphatase and K+ levels in female rats, and induced the decrease in serum albumin and total protein in rats of both sexes. The data from this study will provide an important reference for developing CpG 684 as an adjuvant for vaccines of human use.

Determination of echinacoside in rat serum by reversed-phase high-performance liquid chromatography with ultraviolet detection and its application to pharmacokinetics and bioavailability.

A rapid and simple high-performance liquid chromatographic (HPLC) method has been developed and validated for the determination of echinacoside (ECH) in rat serum. After protein precipitation of serum sample with trichloroacetic acid, the supernatant was directly injected and analyzed on a C(18) CapcellACR analytical column (150 mm x 4.6mm I.D. 5 microm) with a mobile phase consisting of acetonitrile-0.5% acetic acid (15.5:84.5, v/v). The UV detector was set at 330 nm. The lower limit of detection and quantification were 9 and 29.2 ng/mL, respectively, and the calibration curves were linear over the concentration range of 29.2-18250 ng/mL. The assay method was successfully applied to the study of the pharmacokinetics and bioavailability of ECH in rat.

SPE-HPLC method for the determination and pharmacokinetic studies on paeoniflorin in rat serum after oral administration of traditional Chinese medicinal preparation Guan-Xin-Er-Hao decoction.

A new HPLC method for the determination of paeoniflorin in rat serum with solid-phase extraction (SPE) for preconcentration is introduced. Paeoniflorin and an internal standard (pentoxifylline) were extracted from serum by means of SPE using cartridges with octadecyl chemically bound phase. The HPLC separation was then performed on a reversed-phase C(18) column using acetonitrile-water (18:82, v/v) as eluting solvent system, and UV detection at 230 nm to measure the analyte with a limit of quantitation about 10 ng ml(-1). The calibration curve for paeoniflorin was linear (r=0.9938) in the concentration range of 10-1200 ng ml(-1), both intra- and inter-day precision of the paeoniflorin were determined and their coefficience of variation did not exceed 10%. The validated method has been successfully applied for pharmacokinetic studies of paeoniflorin from rat serum after oral administration of Guan-Xin-Er-Hao decoction.