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Acta Crystallogr Sect E Struct Rep Online. 2009 December 1; 65(Pt 12): o3027–o3028.
Published online 2009 November 7. doi:  10.1107/S1600536809042512
PMCID: PMC2971979

Redetermination of 2-[4-(2-hydroxy­ethyl)piperazin-1-ium-1-yl]ethanesul­fonate at 100 K

Abstract

The crystal structure of the title compound (common name HEPES), C8H18N2O4S, has been redetermined at 100 K in order to properly elucidate the protonation state of the HEPES molecule. The piperazine ring has a chair conformation and one of the N atoms in the ring is protonated, which was not previously reported [Gao, Yin, Yang, & Xue (2004). Acta Cryst. E60, o1328–o1329]. The change of protonation state of the nitrogen atom significantly affects the intermolecular interactions in the HEPES crystal. The structure is stabilized by N—H(...)O and O—H(...)O hydrogen bonds and ionic inter­actions, as the title compound in solid state is a zwitterion. HEPES mol­ecules pack in layers that are held together by ionic and weak inter­actions, while a hydrogen-bonded network connects the layers.

Related literature

For background to HEPES and analogous compounds, see: Ferguson et al. (1980 [triangle]); Good & Izawa (1972 [triangle]); Good et al. (1966 [triangle]). For the crystal structure of HEPES crystallized from methanol, see: Wouters et al. (1996 [triangle]) and from water, see: Gao et al. (2004 [triangle]). For related structures, see: Kubicki et al. (2007 [triangle]); Chruszcz et al. (2005 [triangle]); Zhao et al. (2006 [triangle]).

An external file that holds a picture, illustration, etc.
Object name is e-65-o3027-scheme1.jpg

Experimental

Crystal data

  • C8H18N2O4S
  • M r = 238.31
  • Orthorhombic, An external file that holds a picture, illustration, etc.
Object name is e-65-o3027-efi1.jpg
  • a = 8.341 (1) Å
  • b = 9.567 (1) Å
  • c = 27.066 (1) Å
  • V = 2159.8 (4) Å3
  • Z = 8
  • Mo Kα radiation
  • μ = 0.30 mm−1
  • T = 100 K
  • 0.50 × 0.50 × 0.23 mm

Data collection

  • Rigaku R-AXIS RAPID diffractometer
  • Absorption correction: multi-scan (Otwinowski et al., 2003 [triangle]) T min = 0.86, T max = 0.93
  • 613697 measured reflections
  • 17694 independent reflections
  • 14854 reflections with I > 2σ(I)
  • R int = 0.036

Refinement

  • R[F 2 > 2σ(F 2)] = 0.031
  • wR(F 2) = 0.099
  • S = 1.04
  • 17694 reflections
  • 208 parameters
  • All H-atom parameters refined
  • Δρmax = 0.83 e Å−3
  • Δρmin = −0.80 e Å−3

Data collection: HKL-2000 (Otwinowski & Minor, 1997 [triangle]); cell refinement: HKL-2000; data reduction: HKL-2000; program(s) used to solve structure: SHELXS97 (Sheldrick, 2008 [triangle]) and HKL-3000SM (Minor et al., 2006 [triangle]); program(s) used to refine structure: SHELXL97 (Sheldrick, 2008 [triangle]) and HKL-3000SM; molecular graphics: HKL-3000SM, ORTEPIII (Burnett & Johnson, 1996 [triangle]), ORTEP-3 (Farrugia, 1997 [triangle]), Mercury (Macrae et al., 2006 [triangle]) and POV-RAY (The POV-RAY Team, 2004 [triangle]); software used to prepare material for publication: HKL-3000SM.

Table 1
Hydrogen-bond geometry (Å, °)

Supplementary Material

Crystal structure: contains datablocks I, global. DOI: 10.1107/S1600536809042512/fl2269sup1.cif

Structure factors: contains datablocks I. DOI: 10.1107/S1600536809042512/fl2269Isup2.hkl

Additional supplementary materials: crystallographic information; 3D view; checkCIF report

Acknowledgments

The authors thank Zbigniew Dauter for helpful discussions. This work was supported by contract GI11496 from HKL Research, Inc.

supplementary crystallographic information

Comment

The title compound, (I), known commonly as HEPES, is a sulfonic compound used as a zwitterionic buffer. HEPES and analogous compounds (known as the Good buffers) are used during the study of biological processes (Good et al., 1966; Good & Izawa, 1972; Ferguson et al., 1980), and very often during crystallization of macromolecules.

The crystal structures of HEPES crystallized from methanol (determined at 293 K - Wouters et al., 1996) and from water (298 K - Gao et al., 2004) have been already reported. Both previously reported structures crystallized in the Pbca space group, but with different unit-cell parameters. Not only the unit-cell parameters, but also the conformations of the HEPES molecules were different. However, our attention was turned to differences in the reported protonation states of the HEPES molecules. In the structure reported by Wouters et al., the HEPES molecule was presented as zwitterionic (II), while in structure reported by Gao et al. HEPES had both piperazine nitrogen atoms non-protonated and a protonated sulfonic group (III). The non-zwitterionic form of HEPES is quite unusual, as all previously determined structures of compounds from this group, e.g. MES [2-(N-morpholino)ethanesulfonic acid] (Kubicki et al., 2007), MOPS [3-(N-morpholino)propanesulfonic acid] (Chruszcz et al., 2005), PBHPS [piperazine-1,4-diylbis(2-hydroxypropanesulfonic acid)] (Zhao et al., 2006), consistently report zwitterionic forms as being observed in the solid state.

In order to localize all hydrogen atoms, the structure determination was performed at 100 K. The crystal structure of the title compound reported here is isomorphous to the structure reported by Gao et al., but a more detailed analysis revealed that the HEPES molecules are zwitterionic and the nitrogen atom (N1) is protonated (Fig. 1). Change of the localization of the hydrogen atom in comparison with the previously reported structure (Gao et al., 2004) significantly affects the hydrogen bond network (Fig. 2, Table 1), which we believed was previously incorrectly determined. To confirm our finding, we also performed a structural analysis of HEPES crystals (crystallized from water or taken directly from the bottle provided by SIGMA) at 293 K. In both cases (100 K and 293 K) the structures had the same protonation state, which excluded the possibility of temperature dependent changes of protonation.

The differing protonation states reported here and in the structure determined by Wouters et al. suggest that the change in protonation state of the HEPES molecule (and/or the change of the conformations of 2-hydroxyethyl and enthanesulfonic moieties) results in generation of polymorphic forms.

The crystal structure of HEPES is stabilized mainly by hydrogen bonds and ionic interactions. Both nitrogen atoms from the piperazine ring, oxygen atom (O4) from the hydroxyl group and one of the oxygen atom (O2) from the sulfonic group are involved in formation of the hydrogen bond network. Hydrogen bonds extend along the [100] direction and details of their geometric parameters are summarized in Table 1.The HEPES molecules pack in layers that are held together by ionic and weak interactions.

Experimental

HEPES was purchased from SIGMA (99.5% purity, lot 036 K5461). The crystal of (I), used for X-ray diffraction study, was obtained by slow evaporation of HEPES solution in water.

Refinement

All hydrogen atoms were localized using the difference density Fourier map. Their positions and isotropic displacement parameters were refined.

Figures

Fig. 1.
The molecular structures of the title compound. Displacement ellipsoids are drawn at the 75% probability level and hydrogen atoms are drawn as spheres of an arbitrary radius.
Fig. 2.
The molecular packing of compound (I) shown along the [010] axis. Hydrogen atoms are bonds are marked in green, and H-bonds are shown as blue, dashed lines.
Fig. 3.
The structures of (I), (II) and (III).

Crystal data

C8H18N2O4SF(000) = 1024.0
Mr = 238.31Dx = 1.466 Mg m3
Orthorhombic, PbcaMo Kα radiation, λ = 0.7107 Å
Hall symbol: -P 2ac 2abCell parameters from 613697 reflections
a = 8.341 (1) Åθ = 2.9–62.9°
b = 9.567 (1) ŵ = 0.30 mm1
c = 27.066 (1) ÅT = 100 K
V = 2159.8 (4) Å3Block, colorless
Z = 80.50 × 0.50 × 0.23 mm

Data collection

Rigaku R-AXIS RAPID diffractometer17694 independent reflections
Radiation source: fine-focus sealed tube14854 reflections with I > 2σ(I)
graphiteRint = 0.036
Detector resolution: 10 pixels mm-1θmax = 62.9°, θmin = 2.9°
ω scan with χ offseth = −20→20
Absorption correction: multi-scan (Otwinowski et al., 2003)k = −23→23
Tmin = 0.86, Tmax = 0.93l = −67→67
613697 measured reflections

Refinement

Refinement on F2Primary atom site location: structure-invariant direct methods
Least-squares matrix: fullSecondary atom site location: difference Fourier map
R[F2 > 2σ(F2)] = 0.031Hydrogen site location: difference Fourier map
wR(F2) = 0.099All H-atom parameters refined
S = 1.04w = 1/[σ2(Fo2) + (0.060P)2 + 0.0821P] where P = (Fo2 + 2Fc2)/3
17694 reflections(Δ/σ)max = 0.013
208 parametersΔρmax = 0.83 e Å3
0 restraintsΔρmin = −0.80 e Å3

Special details

Geometry. All e.s.d.'s (except the e.s.d. in the dihedral angle between two l.s. planes) are estimated using the full covariance matrix. The cell e.s.d.'s are taken into account individually in the estimation of e.s.d.'s in distances, angles and torsion angles; correlations between e.s.d.'s in cell parameters are only used when they are defined by crystal symmetry. An approximate (isotropic) treatment of cell e.s.d.'s is used for estimating e.s.d.'s involving l.s. planes.
Refinement. Refinement of F2 against ALL reflections. The weighted R-factor wR and goodness of fit S are based on F2, conventional R-factors R are based on F, with F set to zero for negative F2. The threshold expression of F2 > σ(F2) is used only for calculating R-factors(gt) etc. and is not relevant to the choice of reflections for refinement. R-factors based on F2 are statistically about twice as large as those based on F, and R- factors based on ALL data will be even larger.

Fractional atomic coordinates and isotropic or equivalent isotropic displacement parameters (Å2)

xyzUiso*/Ueq
S10.449764 (8)0.776637 (6)0.555875 (2)0.01145 (1)
N20.52317 (2)0.69925 (2)0.302785 (7)0.01111 (2)
O20.30713 (2)0.86815 (2)0.555768 (7)0.01413 (3)
N10.53191 (2)0.75196 (2)0.408697 (7)0.01121 (2)
O40.72742 (3)0.83527 (2)0.216353 (8)0.01514 (3)
C10.45506 (3)0.69919 (3)0.495798 (9)0.01321 (3)
C50.49778 (3)0.58710 (2)0.339211 (8)0.01316 (3)
O10.59735 (3)0.85596 (3)0.560910 (9)0.01744 (3)
C30.56026 (3)0.86534 (2)0.371519 (9)0.01330 (3)
C60.42540 (3)0.64240 (2)0.386736 (8)0.01298 (3)
C40.63131 (3)0.80396 (2)0.324618 (8)0.01271 (3)
C20.46765 (3)0.81142 (3)0.456032 (9)0.01364 (3)
C80.59941 (3)0.73733 (3)0.213505 (9)0.01485 (3)
C70.58957 (3)0.63743 (2)0.257306 (8)0.01343 (3)
O30.43278 (4)0.66097 (3)0.590283 (9)0.02055 (4)
H1A0.5416 (10)0.6363 (10)0.4955 (4)0.030 (2)*
H7B0.5234 (9)0.5552 (9)0.2488 (3)0.0238 (17)*
H2A0.5445 (9)0.8874 (10)0.4673 (3)0.0253 (19)*
H4B0.6453 (10)0.8784 (8)0.3001 (3)0.0252 (16)*
H7A0.7005 (8)0.5925 (7)0.2643 (3)0.0184 (14)*
H3B0.4576 (9)0.9063 (10)0.3665 (3)0.0253 (19)*
H6A0.4173 (11)0.5721 (9)0.4086 (3)0.030 (2)*
H6B0.3201 (9)0.6878 (8)0.3822 (3)0.0199 (15)*
H4A0.7367 (10)0.7630 (8)0.3328 (3)0.0193 (15)*
H1O40.8138 (12)0.7897 (9)0.2127 (3)0.038 (2)*
H5A0.6006 (9)0.5394 (8)0.3472 (3)0.0175 (14)*
H3A0.6309 (10)0.9315 (7)0.3860 (3)0.0217 (16)*
H5B0.4242 (10)0.5192 (9)0.3260 (3)0.0249 (18)*
H1B0.3616 (9)0.6429 (8)0.4933 (3)0.0220 (16)*
H8B0.5045 (10)0.7906 (9)0.2102 (3)0.0225 (17)*
H1N0.6197 (10)0.7168 (7)0.4156 (3)0.0209 (16)*
H2B0.3673 (10)0.8534 (8)0.4489 (3)0.0235 (17)*
H8A0.6076 (9)0.6844 (8)0.1846 (3)0.0193 (15)*

Atomic displacement parameters (Å2)

U11U22U33U12U13U23
S10.01147 (2)0.01296 (2)0.00992 (2)0.00039 (1)0.00046 (1)0.00017 (1)
N20.01141 (5)0.01171 (5)0.01023 (5)−0.00043 (4)0.00008 (4)−0.00049 (4)
O20.01144 (6)0.01507 (5)0.01589 (6)0.00088 (4)0.00135 (4)−0.00176 (4)
N10.01141 (5)0.01222 (5)0.01001 (5)0.00009 (4)0.00001 (4)−0.00027 (4)
O40.01385 (6)0.01526 (6)0.01631 (6)0.00036 (5)0.00216 (5)0.00128 (4)
C10.01505 (8)0.01313 (6)0.01144 (6)−0.00002 (5)0.00161 (5)−0.00042 (5)
C50.01598 (8)0.01164 (6)0.01185 (6)−0.00126 (5)0.00163 (5)−0.00039 (4)
O10.01166 (6)0.02271 (8)0.01794 (7)−0.00175 (5)−0.00219 (5)−0.00297 (6)
C30.01666 (8)0.01192 (6)0.01131 (6)−0.00133 (5)0.00028 (5)−0.00032 (4)
C60.01328 (7)0.01395 (6)0.01170 (6)−0.00217 (5)0.00141 (5)−0.00079 (5)
C40.01330 (7)0.01364 (6)0.01120 (6)−0.00253 (5)0.00046 (5)−0.00035 (5)
C20.01686 (8)0.01329 (6)0.01078 (6)0.00146 (6)0.00107 (5)−0.00046 (5)
C80.01395 (8)0.01909 (8)0.01151 (6)−0.00078 (6)0.00002 (5)0.00110 (5)
C70.01524 (8)0.01335 (6)0.01171 (6)−0.00001 (5)0.00150 (5)−0.00108 (5)
O30.03015 (11)0.01750 (7)0.01399 (6)0.00248 (7)0.00373 (6)0.00455 (5)

Geometric parameters (Å, °)

S1—O11.4525 (3)C5—H5A0.995 (8)
S1—O31.4532 (2)C5—H5B0.963 (8)
S1—O21.4771 (2)C3—C41.5190 (3)
S1—C11.7874 (3)C3—H3B0.951 (8)
N2—C41.4719 (3)C3—H3A0.949 (8)
N2—C51.4724 (3)C6—H6A0.898 (9)
N2—C71.4736 (3)C6—H6B0.987 (8)
N1—C61.4971 (3)C4—H4B0.981 (8)
N1—C31.4984 (3)C4—H4A0.988 (8)
N1—C21.5008 (3)C2—H2A1.016 (9)
N1—H1N0.827 (8)C2—H2B0.948 (8)
O4—C81.4226 (4)C8—C71.5250 (4)
O4—H1O40.848 (10)C8—H8B0.946 (9)
C1—C21.5239 (4)C8—H8A0.935 (8)
C1—H1A0.939 (9)C7—H7B0.988 (8)
C1—H1B0.950 (8)C7—H7A1.037 (7)
C5—C61.5162 (3)
O1—S1—O3114.856 (17)C4—C3—H3A111.1 (5)
O1—S1—O2111.907 (17)H3B—C3—H3A110.1 (7)
O3—S1—O2111.966 (15)N1—C6—C5110.17 (2)
O1—S1—C1106.317 (13)N1—C6—H6A107.9 (6)
O3—S1—C1105.650 (15)C5—C6—H6A109.1 (6)
O2—S1—C1105.296 (12)N1—C6—H6B105.6 (4)
C4—N2—C5108.376 (18)C5—C6—H6B113.7 (4)
C4—N2—C7112.22 (2)H6A—C6—H6B110.1 (7)
C5—N2—C7108.719 (19)N2—C4—C3111.07 (2)
C6—N1—C3109.504 (18)N2—C4—H4B107.2 (5)
C6—N1—C2113.10 (2)C3—C4—H4B109.3 (5)
C3—N1—C2110.809 (19)N2—C4—H4A111.4 (4)
C6—N1—H1N109.3 (5)C3—C4—H4A108.3 (4)
C3—N1—H1N107.8 (5)H4B—C4—H4A109.5 (6)
C2—N1—H1N106.1 (5)N1—C2—C1111.129 (19)
C8—O4—H1O4107.0 (6)N1—C2—H2A107.6 (5)
C2—C1—S1110.620 (17)C1—C2—H2A109.6 (5)
C2—C1—H1A113.1 (6)N1—C2—H2B107.6 (5)
S1—C1—H1A106.9 (6)C1—C2—H2B112.4 (5)
C2—C1—H1B113.9 (5)H2A—C2—H2B108.4 (7)
S1—C1—H1B106.2 (5)O4—C8—C7114.27 (2)
H1A—C1—H1B105.5 (8)O4—C8—H8B106.2 (5)
N2—C5—C6111.782 (19)C7—C8—H8B111.5 (5)
N2—C5—H5A110.8 (4)O4—C8—H8A110.3 (5)
C6—C5—H5A108.7 (4)C7—C8—H8A108.4 (5)
N2—C5—H5B109.5 (5)H8B—C8—H8A105.8 (7)
C6—C5—H5B107.3 (5)N2—C7—C8114.71 (2)
H5A—C5—H5B108.7 (7)N2—C7—H7B107.7 (5)
N1—C3—C4110.055 (19)C8—C7—H7B110.3 (5)
N1—C3—H3B104.6 (5)N2—C7—H7A110.5 (4)
C4—C3—H3B113.0 (5)C8—C7—H7A110.7 (4)
N1—C3—H3A107.7 (4)H7B—C7—H7A102.1 (5)
O1—S1—C1—C2−59.26 (2)C5—N2—C4—C3−59.66 (2)
O3—S1—C1—C2178.27 (2)C7—N2—C4—C3−179.726 (19)
O2—S1—C1—C259.63 (2)N1—C3—C4—N259.63 (3)
C4—N2—C5—C659.21 (3)C6—N1—C2—C162.57 (3)
C7—N2—C5—C6−178.55 (2)C3—N1—C2—C1−174.03 (2)
C6—N1—C3—C4−56.75 (3)S1—C1—C2—N1159.105 (18)
C2—N1—C3—C4177.80 (2)C4—N2—C7—C8−68.69 (3)
C3—N1—C6—C555.97 (3)C5—N2—C7—C8171.44 (2)
C2—N1—C6—C5−179.911 (19)O4—C8—C7—N276.07 (3)
N2—C5—C6—N1−58.33 (3)

Hydrogen-bond geometry (Å, °)

D—H···AD—HH···AD···AD—H···A
O4—H1O4···N2i0.85 (1)1.99 (1)2.8368 (4)173 (1)
N1—H1N···O2ii0.83 (1)1.92 (1)2.7414 (4)169 (1)

Symmetry codes: (i) x+1/2, y, −z+1/2; (ii) x+1/2, −y+3/2, −z+1.

Footnotes

Supplementary data and figures for this paper are available from the IUCr electronic archives (Reference: FL2269).

References

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