The Role of Simcoe Hop in Dry-Hopping: An Analysis of Beer's Physicochemical properties, Flavor Chemistry, and Sensory Evaluation

Detection of the physicochemical properties of beerBefore the analysis of the beer samples, degassing treatment was carried out by filtration. The levels of diacetyl, color, alcohol, bitterness and residual sugars were detected using CDR BeerLab (CDR s.r.l, Florence, Italy)[36,37]. The pH value and total acidity level were measured referring to the ASBC methods [38]. Turbidity was measured by VOS ROTA 90/25 dual-angle turbidity meter (Pentair Group, Minnesota, America), and the reading of H90 was recorded. Total polyphenols (mg/L, gallic acid equivalents) were determined by the Folin-Ciocalteu colorimetric method [39]. Quantification of volatile compounds by headspace-solid-phase microextraction coupled with gas chromatography–mass spectrometry (HS-SPME-GC-MS)Beer samples were subjected to headspace solid-phase microextraction (HS-SPME) at first. The beer sample (10 mL), sodium chloride (2 g) and the internal standard (2-octanol, 50 mg/L ethanol) were mixed well in a 20 mL headspace vial. The headspace vial was heated at 55°C for 10 minutes and extracted with a 50 µm DVB/CAR/PDMS SPME fiber assembly (Supelco, Bellefonte, PA, USA) for 30 minutes. The GC analysis was conducted on a DB-WAX UI column (60mx0.25mmx0.25μm, Thermo Fisher Scientific, Waltham, MA, USA) installed on the GCMS-QP2020NX (Shimadzu Corporation, Kyoto, Japan). The inlet temperature was set to 250°C, helium gas as the carrier gas, and the sample was injected in the splitless mode rate set at 1.5 mL/min. The column temperature was initially maintained at 50°C for 2 minutes, and then it was increased up to 230°C at a rate of 6°C/min.MS analyzes were carried out by electron ionization mass (EI) in the full scan mode, using ionization energy of 70 eV with the ion source temperature set to 230°C, the interface temperature set to 260°C, and the quadrupole temperature was set to 150°C. The mass acquisition range was 29.0-500.0 amu. The resulting full spectrum of volatile substances in the beer sample was used for qualitative and semi-quantitative analysis. Quantification of PFTs by solid-phase extraction coupled with triple quadrupole gas chromatography–mass spectrometry (SPE-GC-MS/MS)The detection methods for 3SH, 3SHA, and 4SMP were adapted from that of Koji Takazumi et al. 40, the ASPEC GX-274 Ag+ solid-phase extraction method was used to extract free PFTs from 50 mL of beer samples. PFTs were quantified by external standardization. The standard mixed solutions of PFTs’ concentrations were 1, 10, 25, 50, and 100 μg/L mixed and gradient-diluted with ethyl acetate. 1-Thioglycerol (10 g/L) was added as an analyte protectant.GC–MS/MS analysis was performed with an Agilent 8890-7000D Triple Quadrupole GC/MS (Agilent Technologies, Palo Alto, CA, USA). An InertCap Pure Wax capillary chromatographic column (30 m × 250μm × 0.25μm, GL Sciences) for separation. Here 2 μ of extract was injected using an automatic sampler Agilent 7693A (Agilent Technologies, Palo Alto, CA, USA). The inlet was operated in pulsed splitless mode (30 psi, 1 min) at 250 °C. The carrier gas was high-purity helium (1 mL/min). The oven was heated from 70°C (held for 1 min) to 250°C at a rate of 20°C/min and then held for 1 minute. The triple quadrupole mass spectrometer was operated in multiple reaction monitoring (MRM) mode.  Sensory evaluationThe beer sensory evaluation panel consisted of 10 professionally trained personnel, aged between 20 and 55 years old, including 6 men and 4 women. They were divided into two groups of 5 each to test each beer sample. Before the evaluation, the beer samples were filled into sealed containers and placed in a water bath to warm up to 15°C. During the formal evaluation, the flavor of the 50 mL beer samples was evaluated and scored. Here, the participants would not know the source of the samples. The attributes assessed included tropical fruit, berry-like, stone fruit, citrus, floral, sweet, and herby. The tasting order was randomized, and each attribute was scored on a scale of 0 to 5 (0 = imperceptible, 1 = very weak, 2 = weak, 3 = moderate, 4 = intense, and 5 = very intense). Additionally, each sample was evaluated three times on different days, and the data was recorded as the average of the three evaluation results. Statistical analysesAll results in this study were expressed as the mean ± SD (standard deviation) from at least three replicates. All data were subjected to analysis of variance (ANOVA) and Multiple Range Test (Tukey’s test) using SPSS 2021 statistics software.