《茅蒼術(shù)工廠化育苗技術(shù)規(guī)程（征求意見稿）》編制說明

江蘇省地方標(biāo)準(zhǔn)《茅蒼術(shù)工廠化育苗技術(shù)規(guī)程》編制說明應(yīng)。但作為道地產(chǎn)區(qū)的江蘇已有近30年沒有形成蒼術(shù)藥材商品了。究其原因，裝運輸、生產(chǎn)檔案等相關(guān)法律、法規(guī)、標(biāo)準(zhǔn)與號）后，2023年9月，南京師范大學(xué)會同南京倍萜源生物科技有限公司、江蘇修改、補充和完善，并對標(biāo)準(zhǔn)內(nèi)容進(jìn)行了修改標(biāo)準(zhǔn)編制組于2023年12月組織召開《茅蒼術(shù)工廠化育苗技術(shù)規(guī)程》地方送審稿，并提交江蘇省市場監(jiān)督管理局和江蘇省農(nóng)業(yè)規(guī)定了茅蒼術(shù)、組培苗、煉苗、工廠化育苗的茅蒼術(shù)無菌組培苗由本課題組參考各類藥用植物無菌組培苗處理方法的基菌處理：取其葉芽，流水沖洗，在超凈臺中將葉芽依次浸泡于75%乙醇、9%次氯酸鈉溶液、75%乙醇，隨后無菌水沖洗3遍。按照該方法處理可獲得經(jīng)多次傳100株茅蒼術(shù)野生苗制備得來的無菌組培苗傳代至圖1茅蒼術(shù)無菌組培苗對茅蒼術(shù)的組培苗干重積累無明顯影響，但影響根總長。CaCl2含量為原含量兩倍時，干重、根數(shù)及根總長均最大。而KNO3的含量在原含量1.5到2倍時，組培苗的干重顯著提高，2倍條件下的植株比1倍含量條件下植株干重高95.7%。NH4NO3含量為原含量的1到1.5倍間，干重與其他含量下的干重有顯著性差異，條件下會影響茅蒼術(shù)總揮發(fā)油及其主成分變化（表2其中CaCl2含量為原含量量最大。而NH4NO3不存在時不利于植株生長，高含量不利于揮發(fā)油的產(chǎn)生。KH2PO4也表現(xiàn)為高濃度促進(jìn)茅蒼術(shù)生長，不利于幾種主成分的積累，雖然最終提高茅蒼術(shù)揮發(fā)油的總含量，通過以上優(yōu)化，茅蒼術(shù)活性成分可提高表1培養(yǎng)基不同大量元素及不同含量對茅蒼術(shù)干重及根部生長的影響MS培養(yǎng)基大量元素含量(×倍數(shù)）根總長(cm)根數(shù)干重(g)折干率(%)CaCl2014.53±1.36a4.33±0.58a0.042±0.002a12.089%±0.43ab0.517.67±1.04abc5.33±0.58a0.055±0.003abc11.63%±0.28a123.63±2.06d6.00±0.82a0.054±0.005d12.62%±0.52b21.81±1.35cd5.00±1.15a0.058±0.005cd11.41%±0.23a245.79±1.30l8.00±1.73b0.062±0.005l11.63%±0.31aNH4NO3017.63±0.87abc7.33±1.53ab0.028±0.003abc16.61%±0.01c0.537.34±4.15gh7.00±1.00ab0.039±0.004gh7.69%±0.01a139.57±4.27hib0.060±0.007hi29.98%±0.03d31.45±1.55f8.00±4.76ab0.067±0.003il12.86%±0.01b224.88±2.21de3.00±1.00c0.025±0.003de13.38%±0.01bKNO3029±2.11ef9.00±1.00ab0.054±0.002ef13.57%±0.01a0.533.01±5.81fg10.67±2.52b0.047±0.001fg14.62%±0.66a129.61±2.20f9.67±1.53ab0.046±0.003f15.03%±0.32a44.75±3.40l7.67±0.58a0.082±0.006m13.75%±0.74a241.96±3.72il10.50±0.58b0.09±0.004n13.23%±0.95aMgSO4015.76±1.53ab6.50±1.91a0.054±0.004ab24.79%±0.03a0.521.02±0.91cd7.33±1.53a0.062±0.004l24.50%±0.03a121.61±1.24cd7.75±1.71a0.050±0.002cd23.57%±0.03aKH2PO421.04±2.11cd5.75±1.71a0.051±0.004cd24.20%±0.03a220.33±1.78bcd5.33±0.58a0.041±0.006bcd21.80%±0.01a017.99±1.75abc3.67±0.58a0.026±0.005abc16.24%±0.21a0.546.6±4.01d9.50±1.29b0.061±0.006ll14.00%±0.72bc123.22±1.35l5.67±1.15c0.051±0.005cd13.39%±1.40c33.41±2.40fg8.67±1.15b0.055±0.003fg14.87%±0.54b228.91±1.99ef7.75±0.96b0.079±0.005m12.87%±0.66c表2不同大量元素不同含量對茅蒼術(shù)組培苗揮發(fā)油主成分含量的影響大量元素濃度(×倍數(shù)）姜畢烯(μg/g)茅術(shù)醇(μg/g)β-桉葉油醇(μg/g)蒼術(shù)酮(μg/g)蒼術(shù)素(μg/g)CaCl20.055.22±5.31a2.17±0.29a7.24±2.11a3.79±0.65a10.34±1.61a0.5191.60±60.65b1.81±0.55a18.94±4.76bc5.04±2.18a70.62±20.93b65.19±11.77b2.16±0.60a17.05±3.63ab5.46±2.02a47.67±1.98c115.95±25.70b2.76±0.16a21.38±5.57cd5.08±1.51a90.20±6.12b2.0106.19±29.87b2.09±0.33a28.25±3.48d5.22±0.16a47.50±13.07cNH4NO30.0122.59±4.24a2.27±0.45a64.54±26.17a4.69±0.08ab86.66±18.30a0.5149.45±14.95a3.06±1.08a55.61±15.75a8.66±0.60c91.67±2.08a80.21±33.26b1.93±0.51a43.43±11.61ab4.24±0.93a35.43±10.73b62.05±7.67b2.27±0.65a42.16±8.80ab5.56±0.78b51.99±10.09b2.047.18±4.41b2.21±0.55a22.69±9.00b4.28±0.13a31.00±7.86bKNO30.0111.48±25.67ab2.87±0.06c34.39±4.07c5.23±0.74a48.85±14.62ab0.5153.49±10.82c2.67±0.41bc18.38±8.67ab7.43±1.82bc68.86±5.68a148.65±49.71bc2.00±0.44ab10.57±0.62a8.52±1.08c59.75±16.80a94.06±6.50a1.91±0.47a23.27±6.57b7.37±0.76bc34.92±8.60b2.0125.52±17.30c2.60±0.40abc35.57±0.99c6.65±0.33ab59.24±10.46aMgSO40.080.5±46.77a2.28±0.67a17.73±7.56a7.08±2.75a20.48±6.47a0.582.29±19.93a2.79±1.15a33.79±11.68b6.64±0.95a40.98±15.00bc3.02±0.85a25.43±10.25ab5.85±0.93a54.89±10.60c91.02±25.22a2.74±0.94a5.41±0.83a33.73±1.67ab2.0135.50±29.00a3.19±1.67a4.79±0.19a28.43±4.08abKH2PO40.065.96±4.45a5.09±1.97a23.35±13.54a6.99±0.49a194.23±89.69a0.596.53±22.26ab2.88±0.23b34.58±8.42a7.54±1.57a44.50±4.27b105.49±6.66abc2.33±0.49b25.30±5.49a6.51±2.56a37.98±9.39b147.34±45.47c1.96±0.16b27.73±6.78a5.71±1.28a45.98±18.53b2.0140.18±16.62bc2.41±0.47b32.52±12.63a6.91±1.54a47.17±14.25b圖2茅蒼術(shù)煉苗與穴盤苗培養(yǎng)建立了各類文獻(xiàn)中公認(rèn)的茅蒼術(shù)中的4種主要活性成分：β-桉葉油醇、茅術(shù)醇、蒼術(shù)酮、蒼術(shù)素的定量分析方法。茅蒼術(shù)種苗揮發(fā)油中有85個組分，其中含量大于1%的有21個（圖3）。從總離子流圖中可以看到，茅術(shù)醇（峰11）、β-圖3茅蒼術(shù)組培苗揮發(fā)油總離子流圖表3茅蒼術(shù)組培苗揮發(fā)油主成分編號保留時間（min）化合物名稱分子式分子量相對含量（%）19.2501-乙基-3-(1-丙烯基)金剛烷（1-etyhl-3-(1-propenyl)adamantane）C15H24204.350.187229.424β-廣藿香烯(β-patchoulene）十氫-4α-甲基-1-亞甲基-7(1-甲基亞乙基)萘C15H24204.350.069939.618（4α-methyl-1-methylene-7-(propan-2-ylidene)decahydronaphthalene）C15H24204.350.8973410.932δ-愈創(chuàng)木烯(δ-guaiene）C15H24204.352.2845511.215反-丁香烯(β-caryophyllene）(1Z,4Z,7Z)-1,5,9,9四-甲基-1,4,7-環(huán)十(三)烯C15H24204.351.9807612.019（(1Z,4Z,7Z)-1,5,9,9-tetramethylcycloundeca-1,4,7-triene）C15H24204.350.7407712.6711-(1,5-二甲基-4-乙烯基)-4-甲基-苯（1-(1,5-dimethyl-4-hexenyl)-4-methyl-benzene）C15H22202.342.8328813.215姜畢烯（zingiberene）C15H24204.356.3303914.031β-倍半水芹烯(β-sesquiphellandrene）C15H24204.351.750515.806氧化石竹烯（caryophylleneoxide）C15H24O220.351.198318.272茅術(shù)醇（hinesol）C15H26O222.370.059118.597β-桉葉油醇(β-eudesmol）C15H26O222.370.010318.835蒼術(shù)酮（atractylone）C15H20O216.321.225519.870β-雪松烯(β-himachalene）C15H24204.350.546122.053蒼術(shù)素（atractylodin）C13H10O182.226.101230.156亞麻酸（(Z,Z,Z)-9,12,15-octadecatrienoicacid）δ-生育酚（2H-1-benzopyran-6-ol,C18H30O2278.4318.475134.4163,4,-dihydro-2,8-dimethyl-2-(4,8,12-trimethyltridecyl-,[2R-[2R*(4R*,8R*)]]）γ-生育酚C27H46O2402.650.671336.364（2,7,8-trimethyl-2-(4,8,12-trimethyltridecyl)chroman-6-ol）C28H48O2416.683.0325表4茅蒼術(shù)種苗內(nèi)8種化合物氣相色譜檢測的線性、檢測范圍和最低檢測限化合物線性范圍回歸方程R2檢測限（mg/mL）反-丁香烯(β-caryophyllene）0.0003-0.03y=24251x-0.81140.99980.0001姜畢烯（zingiberene）0.0003-0.03y=28942x-1.77810.99950.0003β-倍半水芹烯(β-sesquiphellandrene）0.0005-0.03y=19826x-0.99790.99970.0001氧化石竹烯（caryophylleneoxide）0.0005-0.03y=26177x-13.2250.99940.0003茅術(shù)醇（hinesol）0.0005-0.03y=14737x-3.67740.99970.0003β-桉葉油醇(β-eudesmol）0.0005-0.03y=19327x-5.79350.99930.0003蒼術(shù)酮（atractylone）0.003-0.03y=20625x-5.96650.99910.0001蒼術(shù)素（atractylodin）0.0005-0.03y=15730x-3.47190.99980.0003表5茅蒼術(shù)種苗內(nèi)8種化合物氣相色譜檢測的精密度、重復(fù)性、穩(wěn)定性和準(zhǔn)確性化合物精密度a重復(fù)性a穩(wěn)定性a準(zhǔn)確性b反-丁香烯(β-caryophyllene）2.153