2. Promotion effect on the phosphate esters hydrolysis

Usually, phosphate ester in water is quite stable. As an example, hydrolysis Glucose 6-phosphate (G6P), a very common phosphate ester in nature, is a slow process without enzyme in the medium of deionized water (DIW), and becomes even slower in the fresh nanomolar inorganic iron solutions. Inorganic orthophosphate (Pi) in the DIW with the addition of 100 μM G6P at room temperature (22 � 2°C) was initially 0.90 � 0.04 μM, which became 4.86 � 0.26 and 10.35 � 1.19 μM at 4 and 12 days, respectively. The corresponding Pi in the fresh nanomole inorganic iron solutions (0.5–50 nM Fe(NO3)3) were 1.35 � 0.09 and 2.55 � 0.15 μM.

After G6P was added into an aged 14-month 16.5 nM Fe(NO3)3 solution (pH 6.30) at room temperature, made by acid-forced hydrolysis [27], the Pi was rapidly released (e.g. the initial 20 μM G6P, as presented in Figure 1). Like metal ions as well as natural and biomimetic enzymes, the kinetics of G6P hydrolysis in the aged iron solution can be described as a pseudo-first-order reaction for a fixed concentration of G6P [29–36]. For the initial 20 μM G6P, the decrease in G6P concentration, [G6P]t, due to its hydrolysis can be expressed as a function of hydrolysis time, t, as

$$\log\left[\text{G\'{G}P}\right]\_t = -1.31 \times 10^{-5}t - 4.718\left(r^2 = 0.999\right) \tag{1}$$

where [G6P]<sup>t</sup> is in M and t is in second.

#### Figure 1.

Hydrolysis of 20 μM G6P in a 16.5 nM Fe(NO3)3 solution aged 14 months at room temperature (22 � 2°C). (a) Time courses of formation of phosphorantimonylmolybdenum blue complex from phosphate released from hydrolysis of 20 μM G6P at times of 0, 1, 3, and 6 h, respectively; (b) concentration of Pi and G6P during G6P hydrolysis; and (c) pseudo first-order reaction kinetics of G6P.

#### Iron Oxide Nanoparticles: An Inorganic Phosphatase DOI: http://dx.doi.org/10.5772/intechopen.82650

several studies have already demonstrated that the phosphate ester can be hydrolyzed with the interaction of minerals in the aqueous environments [5, 20–25]. Here, the results of laboratory study on the hydrolysis of phosphorus esters, promoted by the iron oxide nanoparticles in water, including the aged nanomolar inorganic iron ion solutions [26–28], were summarized. Additionally, the potential role of inorganic iron oxide nanoparticle for the phosphorus cycles due to the intrinsic

Usually, phosphate ester in water is quite stable. As an example, hydrolysis Glucose 6-phosphate (G6P), a very common phosphate ester in nature, is a slow process without enzyme in the medium of deionized water (DIW), and becomes even slower in the fresh nanomolar inorganic iron solutions. Inorganic orthophosphate (Pi) in the DIW with the addition of 100 μM G6P at room temperature (22 � 2°C) was initially 0.90 � 0.04 μM, which became 4.86 � 0.26 and

10.35 � 1.19 μM at 4 and 12 days, respectively. The corresponding Pi in the fresh nanomole inorganic iron solutions (0.5–50 nM Fe(NO3)3) were 1.35 � 0.09 and

After G6P was added into an aged 14-month 16.5 nM Fe(NO3)3 solution (pH 6.30) at room temperature, made by acid-forced hydrolysis [27], the Pi was rapidly released (e.g. the initial 20 μM G6P, as presented in Figure 1). Like metal ions as well as natural and biomimetic enzymes, the kinetics of G6P hydrolysis in the aged iron solution can be described as a pseudo-first-order reaction for a fixed concentration of G6P [29–36]. For the initial 20 μM G6P, the decrease in G6P concentration, [G6P]t, due to its hydrolysis can be expressed as a function of

Hydrolysis of 20 μM G6P in a 16.5 nM Fe(NO3)3 solution aged 14 months at room temperature (22 � 2°C). (a) Time courses of formation of phosphorantimonylmolybdenum blue complex from phosphate released from hydrolysis of 20 μM G6P at times of 0, 1, 3, and 6 h, respectively; (b) concentration of Pi and G6P during G6P

t � 4:718 r

<sup>2</sup> <sup>¼</sup> <sup>0</sup>:<sup>999</sup> (1)

phosphoesterase activity is postulated.

2.55 � 0.15 μM.

Nanocatalysts

hydrolysis time, t, as

Figure 1.

98

2. Promotion effect on the phosphate esters hydrolysis

log G6P ½ �<sup>t</sup> ¼ �1:<sup>31</sup> � <sup>10</sup>�<sup>5</sup>

where [G6P]<sup>t</sup> is in M and t is in second.

hydrolysis; and (c) pseudo first-order reaction kinetics of G6P.

The corresponding reaction rate constant (k) was 3.02 <sup>10</sup><sup>6</sup> <sup>s</sup> 1 , and the half-life (t1/2) was 6.38 h. Similar to the initial 20 μM of G6P, the Pi concentration of a initial 100 μM of G6P in the aged inorganic iron solution at 1, 3 and 6.7 h was 4.95, 10.74 and 20.62 <sup>μ</sup>M, respectively. The corresponding <sup>k</sup> was 8.83 <sup>10</sup><sup>6</sup> <sup>s</sup> 1 , and the t1/2 was 21.8 h. It is highlighted that these k in the aged iron solution were much higher than the previously reported rates in the presence of the fresh unaged nanomolar inorganic iron [26] and millimole metals [30–32] solutions.

Like aged inorganic iron solution, the concentration of phosphate esters and condensed inorganic phosphate decreased, and inorganic orthophosphate (Pi) increased in a solution bearing iron oxide (IO) nanoparticles, which consists of a dialysis membrane tube (DMT, e.g., Spectra/Por 1 membranes, molecular weight cut-off (MWCO) 6000–8000 Da) filled with iron oxide (DMT-IO). The iron oxide (D) was synthesized by Fe(NO3)3 following the basic protocol of Atkinson [37] and aged at 80°C [28]. The k for 100 μM G6P, Glycerol-2-phosphate (3-carbon, G2P), and three energy metabolism compounds, i.e., adenosine monophosphate (AMP), adenosine diphosphate (ADP), adenosine triphosphate (ATP), as well as two inorganic condensed phosphates, i.e., polyphosphate (poly-Pi), and pyrophosphate (PPi) at room temperature (22°C) was 2.69 <sup>10</sup><sup>5</sup> , 1.68 <sup>10</sup><sup>5</sup> , 1.54 <sup>10</sup><sup>5</sup> , 5.73 <sup>10</sup><sup>6</sup> , 5.76 <sup>10</sup><sup>6</sup> , 3.8 <sup>10</sup><sup>6</sup> , and 5.09 <sup>10</sup><sup>6</sup> <sup>s</sup> 1 , respectively. The corresponding t1/2 of these phosphorus esters and inorganic condensed phosphates was 7.1, 11.5, 12.5, 33.6, 33.4, 50.7, and 37.8 h, respectively.

Measured k of the initial 20 μM G6P with different sources of iron, either the aged inorganic iron solutions or the solutions bearing inorganic iron oxide nanoparticles (DMT-IO), are listed in Table 1. The half-life for aged 4-month Fe (NO3)3 (16.5 nM), FeCl3 (10 nM) and Fe(ClO4)3(10 nM) was 37.8, 58.6 and 78.4 h, respectively, whereas the half-life of IO from the same source (Fe(NO3)3, JT Baker), aged at 5–80°C was 11, 2.7, 3.2 and 2.8 h, though they were in the same order of magnitude. The same patterns were observed for the ATP as well [28]. These results further indicate, as expected, that the behavior of catalysis depends on the sources of iron oxides nanoparticles in solutions—whether FeCl3, or Fe(NO3)3, and even on the different manufacturers, as well as with the different aging temperatures for IO (5–80°C) [37, 38]. No clear relationships between ferric ion (III) sources, age processing, and catalytic activity, with the hydrolysis rate constant, were observed.

These inorganic iron solutions also have the same promotion effects on hydrolysis of different sugar phosphates, including G2P, ribose-5-phosphate (5-carbon, R5P), and fructose 1-phosphate (6-carbon, F1P) (Table 2). As expected, the promotion effect was also found on the hydrolysis of AMP, ADP and ATP, and inorganic condense phosphates (poly-Pi and PPi) as well as the RNA model compound (4-nitrophenyl phosphate ester, pNPP). However, no promotion effects were observed for the hydrolysis of phosphonates (C-P bonded compounds, e.g., 2-aminoethylphosphonic acid, phosphono-formic acid) and inositol hexakisphosphate (IP6) (data not shown).

As expected, the catalytic activity is related to the soaked time of DIW with DMT-IO and the nature of IO, which can be described by the hydrolysis reaction rate constant. The kinetics of k of 100 μM G6P and ATP in three different IOs is presented in Figure 2. These results can be explained by the changes of the nanoparticles concentration in the water. It was expected that the concentration of the IO nanoparticles in these solutions would initially increase up to 10 days and then reach equilibrium. However, the total dissolved iron concentrations in these solutions were still beneath the detection limits of iron (0.1 nM) [39].

#### Nanocatalysts


Phosphate ester Fe source Initial OP

Iron Oxide Nanoparticles: An Inorganic Phosphatase DOI: http://dx.doi.org/10.5772/intechopen.82650

> IO-D (made by Fe(NO3)3, aged a week at 80°C, soak 1 month at 22°C)

> IO-D (made by Fe(NO3)3, aged a week at 80°C, soak 1 month at 22°C)

> IO-D (made by Fe(NO3)3, aged a week at 80°C, soak 1 month at 22°C)

> IO-D (made by Fe(NO3)3, aged a week at 80°C, soak 1 month at 22°C)

IO-A(made by Fe(NO3)3, aged a week at 5°C, soak 1 month at 22°C)

IO-B (made by Fe(NO3)3, aged a week at 22° C, soak 1 month at 22°C)

IO-G (made by FeCl3, aged a week at 80°C, soak 1 month at 22°C)

IO-D (made by Fe(NO3)3, aged a week at 80°C, soak 1 month at 22°C)

80°C, soak 1 month at 22°C)

Pyrophosphate (PPi) IO-D (made by Fe(NO3)3, aged a week at

Phosphate ester hydrolysis in the different inorganic iron solutions.

Glycerol-2-phosphate

Ribose-5-phosphate

Fuctose-1-phosphate

(R5P)

(F1P)

Adenosine monophosphate (AMP)

Adenosine diphosphate (ADP)

Adenosine triphosphate (ATP)

Polyphosphate (poly-Pi)

Table 2.

101

(G2P)

(μM)

Fe standard solution, 7.5 nM, 4 mo. 10 12.69 15.2

Fe(NO3)3, 1000 nM, 6 mo. 20 30.12 6.4 FeCl3, 2 nM, 16 mo. 20 5.29 36.4 Fe(NH4)2(SO4)2, 16.5 nM, 16 mo. 20 11.44 16.8

Fe standard solution, 7.5 nM, 4 mo. 10 13.92 13.8

Fe(NO3)3, 1000 nM, 6 mo. 20 25.19 7.6 FeCl3, 2 nM, 16 mo. 20 7.24 26.6 Fe(NH4)2(SO4)2, 16.5 nM, 16 mo. 20 16.16 11.9

Fe standard solution, 7.5 nM, 4 mo. 10 8.66 22.2

Fe(NO3)3, 1000 nM, 6 mo. 20 17.08 11.3 FeCl3, 2 nM, 16 mo. 20 5.29 36.4 Fe(NH4)2(SO4)2, 16.5 nM, 16 mo. 20 8.5 22.6

Rate constant k (10<sup>6</sup> s 1 )

20 6.4 30.1 50 3.45 55.8 500 0.51 374.3

10 85.02 2.26 50 26.91 7.15 100 16.39 11.74

20 8.15 23.6

20 5.25 36.4

10 79 2.44 100 15.4 12.5 250 6.2 31.1

10 134 1.44 100 5.73 33.6 250 1.59 121

10 61.5 3.13 25 30.2 6.38 100 4.4 43.8

20 19.6 9.8 100 10.8 17.9

20 40.4 4.8 100 8.46 22.7

20 38.7 5.0 100 3.55 54.3

10 67.5 2.85 100 3.8 50.7 250 0.39 492

10 162 1.19 100 5.09 37.8 250 0.71 270

Half-life t0.5 (h)

#### Table 1.

Hydrolysis rate constant of 20 μM G6P in inorganic iron solutions.<sup>a</sup>
