有關CAM 植物, 燈籠草( Kalachoe pinnata ) 的一些新了解

Newly Understanding of Kalanchoe pinnata, a CAM Plant

Fen-Wan Chen

, Meng-Yuan Huang

, Hsueh-Wen Yeh

, Chi-Ming Yang

*,

Yung-Ta Chang

*

1 _{Reseachrer of Institute of Earth Sciences, Academia Sinica}

Taipei, Taiwan

2 _{Research Center for Biodiversity, Academia Sinica}

Taipei, Taiwan

3 _{Department of Life Science, National Taiwan Normal University}

Taipei, Taiwan

(Received: 8 January 2013, accepted: 27 March 2013) ABSTRACT

Kalanchoe pinnata, a CAM plant was evaluated through the stable carbon isotopic analysis to

distinguish whether an obligate CAM plant or not. The δ13_C

PDB values of Kalanchoe pinnata are range

from –22.0 to –29.1 ‰ thus to be suggested to a facultative CAM plant. Also how environmental factors: wind affect the δ13_C

PDB values of plants, was evaluated through experiments. The δ13CPDB values of Kalanchoe pinnata are range from –23.9 to –29.1 ‰ in a wind-blowing treatment and –22.0 to –27.5 ‰

in the control.

Key words: Kalanchoe pinnata, obligate CAM plant, facultative CAM plant, carbon isotope, δ13_C PDB

environmental changes Introduction

For years photosynthesis has been discussed and research wildly. Photosynthesis pathway can be divided into 3 kinds of pathways: C3 pathway

(Calvin Cycle), C4 pathway, Crassulacean Acid

Metabolism (CAM) pathways. The metabolic mechanism of CAM pathway in succulent plants was wildly discussed from late 1950’s to early 1960’s (Bruinma, 1958; Ranson and Thomas, 1960; Wolf, 1960; Walker, 1962). Furthermore, things are more interesting since the improvements of biochemical techniques in late 1980’s. On the other hand, the techniques of stable isotopic analysis were fully developments for over 20 years. The differences in the carbon isotopic composition of plants can be expressed as δ13_C

PDB values: δ13CPDB

(‰)= [(13C/12C(sample)-13C/12C(standard))/13C/12C(standard))]

×103_{. PDB  stands for Pee Dee Belemnite. The}

standard used to report δ13_{C-values as given above}

is the 13_C/12_{C ratio  of limestone, composed of}

Belemnite (Belemnite Americana)  remains, from the Pee Dee Formation, South Carolina (Craig, 1957).  The techniques of carbon isotopic analysis

became a very useful tool to distinguish C3, C4, and

CAM plants. The δ13CPDB values of C3 plants range

from –22.0 to –38.0 ‰ (Yah and Wang, 2001), C4

plants from –10 to –20 ‰ (Bender, 1971), CAM plants from –13 to –30 ‰. Due to the curiosity of how plants represent the climate in ancient world, and interested in how the carbon assimilation acts and would environmental factors make changes in CAM plant, experiments were done in a series of process.

Materials and Methods

The Kalanchoe pinnata (Lam) Pers. (Crassulaceae), is a succulent-leaved plant (Winter

et al., 1997), suggested to be whether an obligate

Crassulacean Acid Metabolism plant (Zotz &Winter 1993) or a facultative CAM plant (Winter

et al., 1982). Some references (Kondo et al., 1998)

classify it as a PCK (Phosphoenolpyruvate carboxykinase) CAM plant. Life plants were grown from leaf cutting under our indoor environment. They have become 3~4 leave plants after one month, then transplanted to 3-inch pots. After 7

Fen-Wan Chen, Meng-Yuan Huang, Hsueh-Wen Yeh, Chi-Ming Yang, Yung-Ta Chang 

10

days, experiment started with 18 days of time. Our environment maintained in 22-23℃ photoperiod 10hr/14hr, light intensity about 3000 lux, and with irrigation once a week. No fertilization was made through the whole experiment.  Tissue harvested at different day in the control environmental was dried for 48 h at 65°C, weighed, and ground to a fine powder using a mortar and pestle.

Results

The experiment was lasted for 18 days, since Feb. 25, 2000 to April 25, 2000. The sampling was made out with the 11th, 13th, 18th days, and only the mature leaves were harvested. During the experiment, the average temperature was 22.5℃ (control 20.6 ℃ ; wind blowing 22.5 ℃ ), light intensity were 3238 & 3164 lux, wind velocity were 2.19 and 0.1 m/s (Table 1.). Irrigation was made once a week. The estimate that Kalanchoe

pinnata is an obligate CAM plant or a facultative

CAM plant can be clearly understood through the isotopic analysis. After one week putting in an 60℃ oven, the samples were powdered, and being analysis by the stable carbon isotope analysis system (Fig 1). The theorem of isotopic analysis is samples are mixed with CuO in 1100℃, 10-2_{~ 10}-3

conditions, processing redox reaction. After CO2

generate, collect and analyze them with MS: SIRA system (Fig2). The δ13_C

PDB value of Kalanchoe pinnata is varied from –23.9 to –29.1 in a contrast

and –22.0 to –27.5 in the control (Table 2).

Discussion

There are some interesting findings: (1) δ13_C

PDB values of Kalanchoe pinnata range from –

22.0 to –29.1; δ13_C

PDB values of Kalanchoe pinnata

in Bender et al. (1973) are –15.2 to –29.9. It seems that δ13_C

PDB values of Kalanchoe pinnata have a

big difference among them. (2) According to the definition of the obligate CAM plant, it would process C4 pathway and have similar δ13CPDB values

with C4 plants – ranging from –10 to -20, but it

didn’t. The δ13_C

PDB values of Kalanchoe pinnata

fall in the range of C3 plant’s, it suggest that Kalanchoe pinnata process Calvin cycle in our

experimental environment. According to the definition of the facultative CAM plant, Kalanchoe

pinnata belongs to that. Similar suggest in Winter

Figure 1. Vacuum carbon isotopic analysis system in the Institute of Earth Sciences, Academia Sinica.

Figure 2. MS (system SIRA) for carbon isotopic analysis.

et al. (1982). (3) Environmental changes affect the δ13CPDB values of Kalanchoe pinnata: In our

blowing experiment, the differences between control and experimental are 2.0 to 2.3, suggesting blowing in our nature has an effect on δ13_C

PDB

values of Kalanchoe pinnata. Since δ13CPDB values

reflect the contributions of the CO2 fixed in the

dark by PEP carboxylase and in the light by RuBP carboxylase (O’Leary & Osmond, 1980), the reason could due to wind effects the flowing of CO2, and the velocity of CO2 speed up the formula

below toward right indirectly, but still needs to be improved (Adams III & Osmond, 1988). Therefore, more research will continue to go on in order to clarify the environmental changes play what kinds of role in the δ13CPDB values of Kalanchoe pinnata.

Thus we can understand the represent meaning of plant’s fossils.

Table 1. The control environmental conditions of absolute CAM plant (Kalanchoe pinnata) Treatments Wind velocity

Exp.(m/s) Wind velocity Control(m/s) Average temperature Exp.(℃) Average temperature Control(℃) Indoor 7 days 2.3 0.2 21.6 20.7 Indoor 14 days 1.1 0.1 21.7 21.7 Indoor 21 days 2.2 0.1 22.7 22.8 Indoor 28 days 1.3 0.1 23.8 23.6

Woody box 7 days 2.4 0.1 22.0 22.0 Woody box 14 days 2.9 0.1 22.6 21.2 Woody box 21days 2.4 0.2 23.7 23.7 Woody box 28 days 2.4 0.1 23.3 23.3 Woody box-2 11 days 2.6 0.1 23.0 23.0 Woody box-2 13 days 2.6 0.1 23.4 23.4 Woody box-2 18 days 2.1 0.2 22.4 22.4 Table 2. The δ13_C

PDB (‰) of absolute CAM plant

(Kalanchoe pinnata) Treatments δ13_C PDB Control (‰) δ13CPDB Exp.(‰) Indoor 7 days -27.5±0.1 - Indoor 14 days - -29.1 Indoor 21 days - -28.3±0.0(2) Indoor 28 days - -27.3±0.1(2) Woody box 14 days -25.9±0.2(3) -25.7±0.2(3) Woody box 21days -25.5 -26.4 Woody box 28 days -23.4 -23.9 Woody box-2 11 days -22.0±0.0(2) -24.0±0.1(2) Woody box-2 13 days -22.5±0.1(2) -24.8±0.0(2) Woody box-2 18 days -22.5±0.0(2) -24.7±0.1(2)

We thank the technical support from Institute of Earth Sciences, Academia Sinica. Also thanks to the financial assistance from the National Council of Science.

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通信作者：張永達（Yung-Ta Chang）；FAX：886-2-29312904；E-mail：biofv031@ntnu.edu.tw

有關

CAM 植物, 燈籠草( Kalachoe pinnata ) 的一些新了解

陳芬莞

黃盟元

葉學文

楊棋明

* 張永達

*

CAM 植物燈籠草( Kalanchoe pinnata )經由穩定性碳同位素分析方法來分別其是否為絕對 CAM 植物或是非絕對 CAM 植物。實驗結果其 δ13_C PDB值介於-22.0 ‰與-29.1 ‰之間，因此判定為 非絕對性 CAM 植物。另外為了了解環境因子-風，對植物碳同位素值之影響，針對其實驗之結果 顯示，有風處理δ13_C PDB值為-23.9 至-29.1 ‰，無風處理 δ13CPDB值為-22.0 至-27.5 ‰。 關鍵詞：燈籠草、絕對CAM 植物、非絕對 CAM 植物、穩定性碳同位素、環境改變、δ13_C PDB