全 文 ：Antibacterial Effect of Waste Liquor of Essence
Oil Extraction from Sabina vulgaris Ant in Foods
Haifang ZHANG1, Xiaohong SHI2, Linhe WANG3, Guosheng ZHANG3, Liqin ZHAO2*
1. School of Material Engineering, Inner Mongolia Vocational College of Chemical Engineering, Huhhot 010010, China;
2. College of Food Science and Engineering, Inner Mongolia Agrcultural University, Huhhot 010010, China;
3. College of Ecology and Environmental Science, Inner Mongolia Agrcultural University, Huhhot 010010, China
Supported by Special Fund for Forest-scientific Research in the Public Interest of China
(201104061).
*Corresponding author. E-mail: zhaoliqin12@sohu.com
Received: November 20, 2015 Accepted: January 17, 2016A
Agricultural Science & Technology, 2016, 17(2): 414-416
Copyright訫 2016, Information Institute of HAAS. All rights reserved Storage and Processing
S abina vulgaris Ant (Cupres-saceae: Sabina) is a creepingevergreen shrub. S. vulgaris
has ecological function, and its branch-
es, leaves and seeds can be used as
medicine, treating a variety of dis-
eases[1]. S. vulgaris is an important na-
tive species in arid and semi-arid
desert areas of northwest China, and it
plays an important role in maintaining
the ecology. S. vulgaris is also a new
tree species in the garden construction
of northern cities. Particularly impor-
tant, S. vulgaris is an excellent insecti-
cidal plant first discovered and report-
ed in China, and it has great develop-
ment and utilization prospects [2]. With
the advance of science and technolo-
gy and development of production
technology, production processes of
food industry have become increas-
ingly mechanized. The disinfection as-
pect of production needs more safe
disinfectants, and the application of
chemical disinfectants in food produc-
tion has been criticized. Therefore, it is
urgent to find a green and sate disin-
fection product. S. vulgaris essence oil
is a kind of natural plant oil extracted
from S. vulgaris. It has been reported
that S. vulgaris essence oil not only
has insecticidal activity[3-4], but also has
significant bactericidal effect with in-
hibitory effects on most pathogenic
bacteria[5-6]. In this study, the bacterio-
static effect of waste liquor of S. vul-
garis essential oil extraction was inves-
tigated to determine whether the
waste liquor had bacteriostatic effect,
as well as the intensity of the bacterio-
static effect, thereby providing a new
synergistic approach for essence oil
extraction enterprises and a theoreti-
cal basis for the development of S.
vulgaris essential oil extraction waste
liquor as a food disinfectant.
Materials and Methods
Strains
The strains of E. coli, S. aureus,
Salmonella, B. subtilis, B. magaterium
and B. cereus were all provided by the
microbiological lab of College of Food
Science and Engineering, Inner Mon-
golia Agricultural University. The indi-
cator strains were basically all
Abstract The antibacterial effects of waste liquor of essence oil extraction from
Sabina vulgaris Ant on Escherichia coli, Staphylococcus aureus, Salmonella, Bacillus
subtilis, Bacillus magaterium and Bacillus cereus in foods were evaluated by paper
disk method. The results showed that the waste liquor of essence oil extraction
from S. vulgaris had inhibitory effects on E. coli, S. aureus and Salmonella with bet-
ter inhibitory effects on E. coli and Salmonella; the bacteriostatic effect of water
steam distillation waste liquor was better than that of water extraction waste liquor.
The more times the waste liquor was distilled, the higher the active ingredients
content was, and the better the bacteriostatic effect was.
Key words Sabina vulgaris Ant; Contamination bacteria; Bacteriostatic effect
臭柏精油提取废液对食品污染
细菌的抑制作用
张海芳 1，石晓红 2，王林和 3，张国盛 3，赵丽芹 2*
（1.内蒙古化工职业学院材料工程系，呼和浩特
010010；2. 内蒙古农业大学食品科学与工程学
院，内蒙古呼和浩特 010010；3.内蒙古农业大学
生态环境学院，内蒙古呼和浩特 010010）
摘 要 研究以大肠杆菌 （Escherichia coli）、
金黄色葡萄球菌（Staphylococcus aureus）、沙门
氏菌 （Salmonella）、 枯草芽孢杆菌 （Bacillus
subtilis）、巨大芽孢杆菌 （Bacillus magaterium）、
蜡状芽孢杆菌（Bacillus magaterium）为指示菌，
采用纸片法评估了臭柏 （Sabina vulgaris Ant）
精油废液对食品污染细菌的抑菌效果。 结果表
明，精油废液对大肠杆菌、金黄色葡萄球菌、沙
门氏菌有抑制效果，且对大肠杆菌和沙门氏菌
的抑菌效果较好； 水蒸气蒸馏后的废液比共水
蒸馏后的废液抑菌效果好。 水蒸气蒸馏的废液
随着蒸馏次数的增加，所含的有效成分越多，抑
菌效果越好。
关键词 臭柏（Sabina vulgaris Ant）精油废液；
食品污染细菌；抑菌效果
基金项目 国家林业公益性行业科研专项
（201104061）。
作者简介 张海芳 （1978-），女 ，内蒙古丰镇
人，硕士，副教授，主要从事农产品加工与贮藏
工程研究 ，E-mail: zhanghaifang1999@sina.com。
*通讯作者，教授，主要从事农产品加工与贮藏
工程研究，E-mail: zhaoliqin12@sohu.com。
收稿日期 2015-11-20
修回日期 2016-01-17
DOI:10.16175/j.cnki.1009-4229.2016.02.038
Agricultural Science & Technology2016
Table 1 Antibacterial effects of S. vulgaris essence oil extraction waste liquor on the
indicator strains
Indicator strain
Diameter of inhibition zone//mm
Water extraction waste liquor Water steam distillation wasteliquor (3 times)
Escherichia coli 10.54 12.00
Staphylococcus aureus 8.53 9.63
Salmonella 10.02 11.55
Bacillus subtilis - -
Bacillus magaterium - -
Bacillus cereus - -
pathogenic bacteria.
Medium
The formula of beef extract pep-
tone medium was as follows: beef ex-
tract 3 g, peptone 5 g, NaCl 5 g, agar
15-20 g, deionized water 1 000 ml and
pH 7.4-7.6. After the preparation, the
beef extract peptone medium was
sterilized at 121℃ for 30 min.
Waste liquor of essence oil extrac-
tion from S. vulgaris
In this study, there were two kinds
of waste liquor of essence oil extrac-
tion from S. vulgaris (referred to as
waste liquor), water steam distillation
waste liquor and water extraction
waste liquor. When the plant coarse
powder was soaked in water and then
heated directly, the water and essence
oil were distilled out; after cooled, the
essence oil was separated. This was
defined as water extraction. When the
plant coarse powder was placed in a
distiller and then distilled with water,
the essence soil, along with water
steam, was distilled out. This was de-
fined as water steam distillation. The
waste liquor was filtered with filter pa-
per. The distillation pots were num-
bered, sealed and preserved at 4 ℃
for use.
Preparation of bacterial suspen-
sions
The indicator strains were picked
out from agar slants and inoculated in
liquid medium, respectively. After 24-h
culture, a certain amount (200 μl) of
each bacterial suspension was trans-
ferred to a conical flask filled with 10-
ml sterile saline or sterile water and os-
cillated for 10 min. The concentration
of the bacterial suspensions was about
105 CFU/ml.
Determination of antibacterial ac-
tivities
The antibacterial activities were
determined by paper disk method [7]. A
certain number of small round pieces
of paper with diameter of 6 mm were
prepared by a hole puncher. They
were dry-heat sterilized and soaked in
different waste liquors. In the control
group, the small round pieces of paper
were soaked in sterile saline. The
cooled (to about 50 ℃ ) beef extract
peptone agar medium was poured into
sterile plates, which were placed hori-
zontally. After the medium was solidi-
fied, a certain amount (200 μl) of each
bacterial suspension was added to the
plates, and coated evenly with a sterile
triangle bar. In each of the plates, total
3 pieces of paper were tiled evenly.
Among them, 2 pieces were soaked in
the same waste liquor, while the re-
maining one was soaked in sterile
saline (control). The plates, along with
filter papers, were placed in an incuba-
tor (37 ℃ , 24 h). Finally, the sizes
(horizontal and vertical diameters) of
the inhibition zones were measured by
a vernier caliper. The test was repeat-
ed three times. The final data were ex-
pressed as the means of the 3 repli-
cates[8].
Results and Analysis
Antibacterial effects of S. vulgaris
essence oil extraction waste liquor
on indicator strains
As shown in Table 1, the water
steam distillation waste liquor and wa-
ter extraction waste liquor showed an-
tibacterial effects on E. coli, S. aureus
and Salmonella in varying degrees, in-
stead of B. subtilis, B. magaterium and
B. cereus. Among the 3 strains, the
antibacterial capacities of S. vulgaris
essence oil extraction waste liquor
ranked as E. coli > Salmonella > S.
aureus. The inhibition zones of S. vul-
garis essence oil extraction waste
liquor for E. coli and Salmonella were
larger, i.e., better inhibitory effects on
E. coli and Salmonella. However, the
inhibition zone of S. vulgaris essence
oil extraction waste liquor for S. aureus
was smaller, i.e., poor inhibitory effect
on S. aureus. The bacteriostatic ca-
pacity of S. vulgaris essence oil ex-
traction waste liquor differed between
different extraction methods. The bac-
teriostatic capacity of water extraction
waste liquor was weaker than that of
water steam distillation waste liquor.
Comparison of antibacterial effects
of S. vulgaris essence oil extraction
waste liquor on indicator strains
between different extraction meth-
ods
The S. vulgaris essence oil ex-
traction waste liquor was divided into
water steam distillation waste liquor
and water extraction waste liquor. For
water steam distillation waste liquor,
under the condition of same-pot water,
the active ingredients content in waste
liquor was different for different distilla-
tion time. As shown in Table 2, the ac-
tive ingredients content in water steam
distillation waste liquor was increased
with the increased distillation times,
thus the bacteriostatic activity of the
waste liquor was increased with the
increased distillation times; the active
ingredients content in water extraction
waste liquor was lower, so its antibac-
terial capacity was weaker.
Conclusions
The S. vulgaris essence oil ex-
traction waste liquor had better an-
tibacterial activities for E. coli and
Salmonella and relatively low antibac-
Table 2 Antibacterial activities of S. vulgaris essence oil extraction waste liquor for the
indicator strains under different extraction methods
Indicator strain
Diameter of inhibition zone//mm
Water
extraction
waste liquor
Water steam
distillation waste
liquor (1 time)
Water steam
distillation waste
liquor (2 times)
Water steam
distillation waste
liquor (3 times)
Escherichia coli 10.54 9.64 11.20 12.00
Staphylococcus
aureus 8.53 8.04 9.22 9.63
Salmonella 10.02 9.12 10.76 11.55
415
Agricultural Science & Technology 2016
Responsible editor: Tingting XU Responsible proofreader: Xiaoyan WU
one person in the same day, and the
survey methods and tools should be
consistent, to achieve timely, careful,
detailed, and objective survey.
Records of the field survey include
meteorological conditions, farming op-
eration, crop growth and development
situation, and plant diseases and in-
sect pests. It is required to make a
truthful record of special climatic condi-
tions in the process of experiment, un-
expected natural and human factors,
and circumstances that have serious
influence on experiment, such as
floods, droughts, and human and live-
stock hazards. Abandonment of exper-
iment (if any) should be reported in
written form in a timely manner.
Harvesting and seed evaluation
Harvesting in suitable time
Maize can be harvested when
maize bracts become loose and yellow
and there are black layers in the base
of bracts.
Field record and sampling
Before harvesting of a block, first-
ly reap the guard row, then take field
survey on actually harvested rows:
quantity of lack of plants, actually har-
vested plants, quantity of empty ears,
quantity of plants with two ears, quan-
tity of decayed ears, and quality of
maize grains. For harvested maize,
weigh the fresh ears and take sam-
ples, pack samples of each block into
mesh bags and hang the bags out-
doors for drying and seed evaluation. It
is required to make strict harvesting,
shelling, drying, and yield calculation
by blocks.
Indoor seed evaluation
For indoor seed evaluation, it is
required to select precise instrument to
measure and make a record of ear
shape, ear length, ear thickness, tip-
barren length, ear row number, quan-
tity of row grains, grain type, grain col-
or, and ear axis color, and dry maize
by blocks. After maize is dried to con-
stant weight, measure moisture con-
tent of each block with a moisture ana-
lyzer (measuring three times and cal-
culating the average value), and con-
vert into the yield of regional experi-
ment as per 14% maize standard
moisture content.
Conclusions
Finally, it is required to draft for-
mats and requirements according to
the experiment scheme, make truthful,
objective, reliable and comprehensive
evaluation on agronomic traits such as
adaptability, yielding ability, stress, dis-
ease-resistance, and external charac-
teristics, and submit standardized ex-
periment report to competent authori-
ties.
References
[1] LIU M (刘敏), MOU Y (牟愔), LI XQ (李
绪清 ), et al. How to do maize regional
experiment (如何做好玉米区域试验工
作 ) [J] Bulletin of Agricultural Science
and Technology (农业技术通讯). 2011
(5): 93-95.
[2] ZHAO CX (赵存仙). Technologies and
problems in maize regional experiment
for mountainous areas (山区玉米品种区
域试验中应注意的技术和问题) [J] Bul-
letin of Agricultural Science and Tech-
nology (农业技术通讯 ). 2012(7): 142-
144.
[3] AN F (安芬). Methods and field manage-
ment for maize regional experiment (玉
米区域试验方法及田间管理) [J] Seed
World (种子世界). 2008 (4): 46.
[4] LIU HM (刘红梅), LIU JW (刘江卫). Key
technical points for maize regional ex-
periment (玉米品种区域试验工作的技
术要点 ) [J] Seed World (种子世界 ).
2011 (11): 34-35.
[5] BAI WJ (白万金), YANG P (杨鹏). Sev-
eral Understandings about Maize Re-
gional Experiment (搞好玉米品种区域
试验工作的几点体会) [J] Seed Science
& Technology (种子科技). 2008(5): 32-
33.
Responsible editor: Tingting XU Responsible proofreader: Xiaoyan WU
terial activity for S. aureus, but had no
bacteriostatic effects on B. subtilis, B.
magaterium and B. cereus.
The antibacterial capacity of S.
vulgaris essence oil extraction waste
liquor differed between different ex-
traction methods, and the antibacterial
capacity of water extraction waste
liquor was weaker than that of water
steam distillation waste liquor. The
more times the water steam distillation
waste liquor was distilled, the higher
the active ingredients content was, and
the better the bacteriostatic effect was.
References
[1] Editorial Committee of Inner Mongolia
Flora (内蒙古植物志编辑委员会). Inner
Mongolia flora (Vol. I) (内蒙古植物志(第
一 卷 )) [M]. Hohhot: Inner Mongolia
Peoples Publishing House (呼和浩特:
内蒙古人民出版社), 1998.
[2] ZHANG X (张兴), FU CB (付昌斌), GAO
CF (高聪芬), et al. Research develop-
ment of a novel insecticidal plant savin
juniper (Sabina vulgaris) (新杀虫植物砂
地柏研究进展) [J]. Journal of Northwest
Agriculture & Forestry University (Nat-
ural Sciences) (西北农林科技大学学报
(自然科学版)), 1995, 23(4): 53-57.
[3] LI GZ (李广泽), HE J (何军), YAN HY
(闫海燕), et al. Advances in the studies
on chemical constituents and bioactivi-
ties of Sabina vulgaris Ant (砂地柏化学
成分及生物活性研究进展) [J]. Journal
of Northwest Agriculture & Forestry Uni-
versity (Natural Sciences) (西北农林科
技大学学报 (自然科学版 )), 2006, 34
(10): 133-139.
[4] ZHANG BH (张保华), JIANG ZL (江志
利), CHEN AL (陈安良), et al. Studies
on the 0.75% essential oils from Sabina
vulgaris Ant. Emulsifiable concentrate
and its insecticidal activities (0.75%砂地
柏精油乳油的研制及其杀虫活性测定 )
[J]. Acta Agriculturae Boreali-occiden-
talis Sinica (西北农业学报 ), 2005, 14
(3): 29-31.
[5] GUO XY (郭秀艳), BAO LH (包丽华),
ZHANG GS (张国盛), et al. In vitro an-
tibacterial activity of the essential oil of
Sabina vulgaris branch leaves (臭柏枝
叶精油的体外抑菌活性) [J]. Journal of
Anhui Agricultural Sciences (安徽农业
科学), 2009, 37(4): 1607-1610.
[6] ZULFIYA YUNUS (祖丽皮亚·玉努斯).
Antibacterial effect of the ethanol ex-
traction from Sabina vulgaris Ant bark
(新疆圆柏枝皮醇提物抗菌活性的测定)
[J]. Food Science (食品科学), 2006, 27
(7): 55-58.
[7] ZHOU BJ (周帮靖). Antibacterial effects
of commonly-used Chinese medicines
and measurement methods (常用中药
的 抗 菌 作 用 及 其 测 定 方 法 ) [M].
Chongqing: Chongqing Branch of Sci-
ence and Technology Literature Pub-
lishing House (重庆: 科学技术文献出版
社重庆分社), 1987.
[8] GUO XY (郭秀艳 ). The extraction and
antibacterial effect of Sabina vulgaris
Ant. and Pinus tabuliformis Carr. (臭柏、
油松精油的提取及抑菌活性) [D]. Huh-
hot: Inner Mongolia Agricultural Univer-
sity (呼和浩特: 内蒙古农业大学), 2009.
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