An alternative and simplified approach to identification and test for minimum content of TCM herbal drugs.

Following a decision of the European Pharmacopoeia (Ph. Eur.) Commission, the Traditional Chinese Medicines (TCM) Working Party started a pilot phase to examine the suitability of a high-performance thin-layer chromatography (HPTLC) minimum content test as an alternative to the classical assay in TCM monographs. This approach was evaluated with two TCM herbal drugs: Fritillaria thunbergii bulbs (FTB) and Corydalis rhizome (CYR). Firstly, the existing HPTLC methods were optimised for both drugs. The new methods were applied to the evaluation of multiple samples, and acceptance criteria for the identification, following Ph. Eur. chapter 2.8.25. High-performance thin-layer chromatography of herbal drugs and herbal drug preparations, were set. The HPTLC test for minimum content of markers was then developed and validated. In this test, the intensity of the marker zone in the fingerprint of the sample is compared to the corresponding zone in the reference solution, which has a concentration giving an intensity equivalent to the acceptance criterion. This test gives a pass or fail result rather than a content and can be performed visually (on the images) or by software (using peak profiles from images; PPI). Reproducibility of the HPTLC methods was evaluated in a collaborative trial including six laboratories. In summary, results for FTB from five laboratories were in agreement. The remaining laboratory did not pass the identification of the samples. For CYR, all laboratories presented the same results for identification. In the test for minimum content, one borderline sample passed in four laboratories and failed in two. All laboratories reached similar conclusions for the other seven samples. The HPTLC methods proposed offer a simplified approach to evaluating identity and minimum content of TCM drugs in a single analysis.

Fast method for the determination of short-chain-length polyhydroxyalkanoates (scl-PHAs) in bacterial samples by In Vial-Thermolysis (IVT).

A new method based on the GC-MS analysis of thermolysis products obtained by treating bacterial samples at a high temperature (above 270°C) has been developed. This method, here named "In-Vial-Thermolysis" (IVT), allowed for the simultaneous determination of short-chain-length polyhydroxyalkanoates (scl-PHA) content and composition. The method was applied to both single strains and microbial mixed cultures (MMC) fed with different carbon sources. The IVT procedure provided similar analytical performances compared to previous Py-GC-MS and Py-GC-FID methods, suggesting a similar application for PHA quantitation in bacterial cells. Results from the IVT procedure and the traditional methanolysis method were compared; the correlation between the two datasets was fit for the purpose, giving a R2 of 0.975. In search of further simplification, the rationale of IVT was exploited for the development of a "field method" based on the titration of thermolyzed samples with sodium hydrogen carbonate to quantify PHA inside bacterial cells. The accuracy of the IVT method was fit for the purpose. These results lead to the possibility for the on-line measurement of PHA productivity. Moreover, they allow for the fast and inexpensive quantification/characterization of PHA for biotechnological process control, as well as investigation over various bacterial communities and/or feeding strategies.

Avalanche photodiodes and quenching circuits for single-photon detection.

Avalanche photodiodes, which operate above the breakdown voltage in Geiger mode connected with avalanche-quenching circuits, can be used to detect single photons and are therefore called singlephoton avalanche diodes SPAD's. Circuit configurations suitable for this operation mode are critically analyzed and their relative merits in photon counting and timing applications are assessed. Simple passive-quenching circuits (PQC's), which are useful for SPAD device testing and selection, have fairly limited application. Suitably designed active-quenching circuits (AQC's) make it possible to exploit the best performance of SPAD's. Thick silicon SPAD's that operate at high voltages (250-450 V) have photon detection efficiency higher than 50% from 540- to 850-nm wavelength and still ~3% at 1064 nm. Thin silicon SPAD's that operate at low voltages (10-50 V) have 45% efficiency at 500 nm, declining to 10% at 830 nm and to as little as 0.1% at 1064 nm. The time resolution achieved in photon timing is 20 ps FWHM with thin SPAD's; it ranges from 350 to 150 ps FWHM with thick SPAD's. The achieved minimum counting dead time and maximum counting rate are 40 ns and 10 Mcps with thick silicon SPAD's, 10 ns and 40 Mcps with thin SPAD's. Germanium and III-V compound semiconductor SPAD's extend the range of photon-counting techniques in the near-infrared region to at least 1600-nm wavelength.