Institute of Biological Chemistry and Biochemistry/Biophysics Program, Washington State University, Pullman, Washington 99164-6340.
Plant Physiol. 1991 May;96(1):38-43. doi: 10.1104/pp.96.1.38.
Cell-free extracts from Pinus ponderosa Lawson (ponderosa pine) and Pinus sylvestris L. (Scotch pine) wood exhibited high levels of monoterpene synthase (cyclase) activity, whereas bark extracts of these species contained no detectable activity, and they inhibited cyclase activity when added to extracts from wood, unless polyvinylpyrrolidone was included in the preparation. The molecular mass of the polyvinylpyrrolidone added was of little consequence; however, polyvinylpolypyrrolidone (a cross-linked insoluble form of the polymer) was ineffective in protecting enzyme activity. Based on these observations, methods were developed for the efficient extraction and assay of monoterpene cyclase activity from conifer stem (wood and bark) tissue. The level of monoterpene cyclase activity for a given conifer species was shown to correlate closely with the monoterpene content of the oleoresin and with the degree of anatomical complexity of the specialized resin-secreting structures. Cyclase activity and monoterpene content were lowest in the stems of species containing only isolated resin cells, such as western red cedar (Thuja plicata D. Don). Increasing levels of cyclase activity and oleoresin monoterpenes were observed in advancing from species with multicellular resin blisters (true firs [Abies]) to those with organized resin passages, such as western larch (Larix occidentalis Nutt.), Colorado blue spruce (Picea pungens Engelm.) and Douglas-fir (Pseudotsuga menziesii [Mirb.] Franco). The highest levels of cyclase activity and oleoresin monoterpenes were noted in Pinus species that contain the most highly developed resin duct systems. The relationship between biosynthetic capacity, as measured by cyclase activity, monoterpene content, and the degree of organization of the secretory structures for a given species, may reflect the total number of specialized resin-producing cells per unit mass of stem tissue.
从黄松（Pinus ponderosa Lawson）和欧洲赤松（Pinus sylvestris L.）的木材中提取的无细胞提取物表现出高水平的单萜合酶（环化酶）活性，而这些物种的树皮提取物则没有检测到活性，并且除非在制备过程中加入聚乙烯吡咯烷酮（PVP），否则它们会抑制从木材中提取的环化酶活性。添加的 PVP 分子量大小影响不大；然而，聚乙烯基聚吡咯烷酮（聚合物的交联不溶性形式）在保护酶活性方面无效。基于这些观察结果，开发了从针叶树茎（木材和树皮）组织中有效提取和测定单萜环化酶活性的方法。证明给定针叶树物种的单萜环化酶活性水平与松脂中的单萜含量以及专门的树脂分泌结构的解剖复杂性程度密切相关。仅含有孤立树脂细胞的物种（如西部红柏（Thuja plicata D. Don））的茎中，环化酶活性和单萜含量最低。从具有多细胞树脂泡（真冷杉属）的物种到具有有组织的树脂通道的物种，如西部落叶松（Larix occidentalis Nutt.）、科罗拉多蓝云杉（Picea pungens Engelm.）和花旗松（Pseudotsuga menziesii [Mirb.] Franco），观察到环化酶活性和松脂单萜不断增加。在包含最发达树脂导管系统的松树物种中，环化酶活性和松脂单萜含量最高。以环化酶活性、单萜含量和给定物种分泌结构的组织程度来衡量的生物合成能力与每单位质量的茎组织中专门的树脂产生细胞的总数之间可能存在相关性。
