Chronic toxic effect of Acenapthene on diverse microalgaes

Table 2: Two Way Analysis of variance (ANOVA) of C. vulgaris, D. subspicatus and Scytonema sp. with reference to

biochemical parameters (pigments, metabolites and enzymes) of control and three graded concentrations of

Acenapthene after 16 days of incubation.

Parameters

Chlorella vulgaris

Desmodesmus subspicatus

Scytonema sp.

F(cal)

P(F<=F(cal))

F(0.05)

F(cal)

P(F<=F(cal)) F(0.05) F(cal) P(F<=F(cal)) F(0.05)

Total Chlorophyll

0.81

N.S. (P>0.05)

0.51

0.83

N.S. (P>0.05)

0.50

0.56

N.S. (P>0.05)

0.65

Carotenoids

0.11

N.S. (P>0.05)

0.95

0.01

N.S. (P>0.05)

0.99

0.03

N.S. (P>0.05)

0.98

Phycobilliproteins

2.07

* (P<=0.05)

0.05

2.39

* (P<=0.05)

0.02

2.89

* (P<=0.05)

0.01

Carbohydrate

0.70

N.S. (P>0.05)

0.57

0.48

N.S. (P>0.05)

0.70

0.31

N.S. (P>0.05)

0.81

Protein

0.12

N.S. (P>0.05)

0.95

0.19

N.S. (P>0.05)

0.90

0.003

N.S. (P>0.05)

0.99

Amino Acid

1.44

N.S. (P>0.05)

0.28

0.01

N.S. (P>0.05)

0.99

0.04

N.S. (P>0.05)

0.98

Phenol

3.23

N.S. (P>0.05)

0.06

6.04

** (P<=0.01)

0.009

3.18

N.S. (P>0.05)

0.08

Nitrate Reductase

0.15

N.S. (P>0.05)

0.93

0.03

N.S. (P>0.05)

0.99

0.42

N.S. (P>0.05)

0.73

Glutamine

0.19

N.S. (P>0.05)

0.90

0.63

N.S. (P>0.05)

0.61

0.06

N.S. (P>0.05)

0.97

Synthetase

Succinate

0.56

N.S. (P>0.05)

0.65

1.34

N.S. (P>0.05)

0.30

0.43

N.S. (P>0.05)

0.73

Dehydrogenase

1975). The cells were further suspended in 50 mM

Estimation of Carbon and Nitrogen

potassium phosphate buffer (pH 7.0) and the levels of

Carbon and Nitrogen were determined using the elemental

phycobiliproteins, phycocyanin, allophycocyanin, and

analyzer PE2400 Series II CHNS/O. (Mohammady et al.

phycoerythrene were measured spectrophotometrically

2005).

at 562, 615, and 652 nm, respectively, after repeated

freezing and thawing (Bennett and Bogorad, 1973).

Enzymatic Assays

Biochemical Analysis

Estimation of in-vivo nitrate reductase activity was

made based on total nitrite formation (Sempruch, 2008).

Biochemical studies included an estimation of the levels

Ammonia-assimilating glutamine synthetase (GS) activity

of carbohydrates, proteins, amino acids, and phenols. The

was determined through a reading of Mn2+ γ-glutamyl

culture medium was discarded through centrifugation

transferase activity (Pamiljans et al., 1962). Succinate

and the cells were thoroughly crushed in a mortar and

dehydrogenase (SDH), a major enzyme in the TCA cycle

pestle with 80% ethanol. The supernatant obtained after

catalyzing the conversion of succinates to fumarates, was

centrifugation was used for biochemical analysis. The

measured (Kun and Abood, 1949).

Anthrone method (Roe, 1955) was applied for total

carbohydrate estimate on using glucose as a standard.

Statistical Analysis

Total soluble proteins were determined using bovine

serum albumin as the standard (Lowry et al. , 1951).

Two way Analysis of variance was carried out using

Amino acid content was estimated through the Ninhydrin

Ky-Plot software. Results were tested by multivariate

method (Lee and Takahasi, 1996) and phenol levels

analysis to correlate between chlorophyll-a, carotenoids,

were determined by the Folin-Coicalteau reagent method

phycocyanin, allophycocyanin, phycoerythrin, nitrate

(Malick and Singh, 1980).

reductase, glutamate synthetase, succinate dehydrogenase,

469