Electronic analytical balance

Mice were euthanized by overdose of chloral hydrate and killed 3 days after ICH. Brains were removed immediately and separated into left and right hemispheres, and cerebellum. Each hemisphere was cut into 4 mm thickness block around the needle track. Brain samples were immediately weighted on an electronic analytical balance (Mettler Toledo, Columbus, OH, USA) to obtain the wet weight. Brain samples were dried at 100 °C for 24 hours to obtain the dry weight. Brain water content was calculated as: (wet weight − dry weight)/wet weight × 100%.

Microfuge 22R desktop refrigerated centrifuge (Beckman, Germany), ultra-low temperature refrigerator (Anhui Zhongke Duling Co., Ltd.), electric thermostatic water bath (Changzhou Putian Instrument Manufacturing Co., Ltd.), electronic analytical balance [Mettler-Toledo Instruments (Shanghai) Co., Ltd.], electric heating constant temperature blast drying oven (Shanghai Yuejin Medical Equipment Factory), physiological saline (Shijiazhuang Siyao Co., Ltd.), BSA24S-CW electronic analytical balance (Sartorius, Germany), paraformaldehyde fixative solution (Wuhan Sewell Biotechnology Co., Ltd.); TissueLyser-24 (Shanghai Jingxin Industrial Development Co., Ltd.), BCA protein concentration assay kit (Beijing Solarbio company), NE-PER™ Nuclear and Cytoplasmic Extraction Reagents (ThermoFisher Scientific, United States), Mouse IFN-γ enzyme-linked immunosorbent assay kit (Jianglai Biotechnology), Mouse TNF-α ELISA kit (Jianglai Biotechnology), Mouse IL-6 ELISA kit (Jianglai Biotechnology), Anti-Claudin 1 (Abcam, United States); Anti-Occludin (Abcam, United States), Anti-ZO1 (Abcam, United States); Anti-HIF-1α (Affinity Biosciences, United States); Anti-NF-κB p65 (Abcam, United States); Anti-STAT1 (Proteintech).

Vanquish Flex Ultra-performance Liquid Chromatography (UPLC) and TSQ Altis Triple Quadrupole Mass Spectrometer (MS) (Thermo Fisher Scientific, United States), nitrogen blowing concentrator (Beijing Politech Instrument Co., Ltd., China), High-speed Refrigerated Centrifuge (Yancheng Kait Experimental Equipment Co., Ltd. China), Vortex Meter (Wiggens, Germany), Milli-Q Purification System (Millipore, United States), KQ-500E Ultrasonic Cleaner (Kunshan Ultrasonic Instrument Co., Ltd. China), and Electronic analytical balance (Mettler Toledo, United States).

The sample powder weighed using the electronic analytical balance (one in 10,000, Germany Mettler Toledo) was placed in a conical flask (50 mL) with stopper and 25 mL of methanol were added. After soaking for 4 h, ultrasound assisted solvent extraction 20 min (UASE, KQ-500DB, Kunshan, China) was carried out using ultrasound bath at 60°C, power 150 w, frequency 20 kHz. Aliquots of methanol were used to make up for the mass of solvent lost through evaporation. The test solution was centrifuged (220R centrifuge, Germany Hettich) and obtained by filtering the prepared solution through a 0.45 µm membrane filter.
The sample masses of the root powder for each variety were: “Diana” 0.42 g, “Adolescent Boy” 0.22 g, “Zhu Guang” 0.17 g, “Cameo” 0.16 g, “Mo Zi Ling” 0.14 g, “Qiao Ling” 0.24 g, “L—Bowl of Beauty” 0.16 g, “Zhong Sheng Fen” 0.15 g, “Karl” 0.13 g, control medicinal varieties “Zi Hai” 0.4 g, and wild medicinal P. lactiflora. artificially transplantedare variety 0.25 g.

Average seed weight was tested through weighing mature dry seeds in batches of 100 using an electronic analytical balance (Mettler, Toledo). Measurements of cotyledon area were made through scanning these organs to form a digital image and then calculating area using Image J software. Mature seeds were photographed under relevant magnification using a HIROX three-dimensional video microscope.
Mature dried seeds were imbibed for 60–100 min and dissected under microscope for isolating mature embryos. The embryos were incubated overnight in buffer (30 mM sodium phosphate, pH 7.0, 10 mM EDTA, 1% Triton X-100 and 1% DMSO) at 37°C, fixed for 1 h in buffer (FAA with 10% formalin, 5% acetic acid and 45% ethanol) and 0.01% Triton X-100, and dehydrated with an ethanol series, as described by Ohto et al. (2005) (link). The embryos were then treated for 1–2 h in Hoyer’s buffer (3:0.8:0.4 of chloral hydrate: water: glycerol). A HIROX three-dimensional video microscope, under relevant magnification, was used to observe the treated embryos. The cell size of cotyledon embryos and the cell size of globular stage embryos were measured using Image J software.